As noted in Section 3.3.1, a QA/QC program plan should be developed that documents internal QA/QC procedures (defined under Section 3.3.1) to be implemented by the participating laboratory for each analyte; this plan should provide a list identifying each instrument used in making analyte determinations.
| | A QA/QC status report should be written bimonthly for each analyte. In this report, the results of the QC program during the reporting period should be reported for each analyte in the following manner: The number (N) of QC samples analyzed during the period; a table of the target levels defined for each sample and the corresponding measured values; the mean of F/T value (as defined
below) for the set of QC samples run during the period; and, use of X + 2 S (as defined below) for the set of QC samples run during the period as a measure of precision. | |
| | As noted in Section 2, an F/T value for a QC sample is the ratio of the measured concentration of analyte to the established (i.e., reference) concentration of analyte for that QC sample. The equation below describes the derivation of the mean for F/T values, X: | |
| | The standard deviation, S, for these measurements is derived using the following equation (note that 2 S is twice this value): | |
| | The nonmandatory QA/QC protocol (see Attachment 3) indicates that QC samples should be divided into several discrete pools, and a separate estimate of precision for each pools then should be derived. Several precision estimates
should be provided for concentrations which differ in average value. These precision measures may be used to document improvements in performance with regard to the combined pool. | |
| | Participating laboratories should use the CTQ proficiency program for each analyte. Results of the this program will be sent by CTQ directly to physicians designated by the participating laboratories. Proficiency results from the CTQ program are used to establish the accuracy of results from each participating laboratory, and should be provided to responsible physicians for use in trend analysis. A proficiency report consisting of these proficiency results should accompany data reports as an attachment. | |
| | For each analyte, the proficiency report should include the results from the 6 previous proficiency rounds in the following format: | |
1. Number (N) of samples analyzed;
2. mean of the target levels, (1/N) S T i, with T i being a consensus mean for the sample;
3. mean of the measurements, (1/N) S M i, with M i being a sample
measurement;
4. a measure of error defined by:
(1/N) the sum of (Ti-Mi)2
Analytical data reports should be submitted to responsible physicians directly. For each sample, report the following information: The date the sample was received; the date the sample was analyzed; appropriate chain-of-custody information; the type(s) of analyses performed; and, the results of the analyses. This information should be reported on a form similar to the form provided an appropriate form. The most recent proficiency program report should accompany the analytical data reports (as an attachment).
Confidence intervals for the analytical results should be reported as X + 2 S, with X being the measured value and 2 S the standard deviation calculated as described above.
For CDU or B2MU results, which are combined with CRTU measurements for proper reporting, the 95% confidence limits are derived from the limits for CDU or
B2MU, (p), and the limits for CRTU, (q), as follows:
X / 1 \ 1
--- + or - |----- | (Y2 x p2 + X2 x q2) ---
Y \ Y2 / 2
For these calculations, X + p is the measurement and confidence limits for CDU or B2MU, and Y + q is the measurement and confidence limit for CRTU.
Participating laboratories should notify responsible physicians as soon as they receive information indicating a change in their accreditation status with the CTQ or the CAP. These physicians should not be expected to wait until formal notice of a status change has been received from the CTQ or the CAP.
3.4 Instructions to Physicians
Physicians responsible for the medical monitoring of cadmium-exposed workers must collect the biological samples from workers; they then should select laboratories to perform the required analyses, and should interpret the analytic results.
3.4.1 Sample Collection and Holding Procedures
Blood Samples. The following procedures are recommended for the collection, shipment and storage of blood samples for CDB analysis to reduce analytical variablility; these recommendations were obtained primarily through personal communications with J.P. Weber of the CTQ (1991), and from reports by the Centers for Disease Control (CDC, 1986) and Stoeppler and Brandt (1980).
To the extent possible, blood samples should be collected from workers at the same time of day. Workers should shower or thoroughly wash their hands and arms before blood samples are drawn. The following materials are needed for blood sample collection: Alcohol wipes; sterile gauze sponges; band-aids; 20- gauge, 1.5-in. stainless steel needles (sterile); preprinted labels; tourniquets; vacutainer holders; 3-ml "metal free" vacutainer tubes (i.e., dark-blue caps), with EDTA as an anti- coagulant; and, styrofoam vacutainer shipping containers.
Whole blood samples are taken by venipuncture. Each blue- capped tube should be labeled or coded for the worker and company before the sample is drawn. (Blue-capped tubes are recommended instead of red-capped tubes because the latter may consist of red coloring pigment containing cadmium, which could contaminate the samples.) Immediately after sampling, the vacutainer tubes must be thoroughly mixed by inverting the tubes at least 10 times manually or mechanically using a
Vortex device (for 15 sec). Samples should be refrigerated immediately or stored on ice until they can be packed for shipment to the participating laboratory for analysis.
The CDC recommends that blood samples be shipped with a "cool pak" to keep the samples cold during shipment. However, the CTQ routinely ships and receives blood samples for cadmium analysis that have not been kept cool during shipment. The CTQ has found no deterioration of cadmium in biological fluids that were shipped via parcel post without a cooling agent, even though these deliveries often take 2 weeks to reach their destination.
Urine Samples. The following are recommended procedures for the collection, shipment and storage of urine for CDU and B2MU analyses, and were obtained primarily through personal communications with J.P. Weber of the CTQ (1991), and from reports by the CDC (1986) and Stoeppler and Brandt (1980).
Single "spot" samples are recommended. As B2M can degrade in the bladder, workers should first empty their bladder and then drink a large glass of water at the start of the visit. Urine samples then should be collected within 1 hour. Separate samples should be collected for CDU and B2MU using the following materials: Sterile urine collection cups (250 ml); small sealable plastic bags;
preprinted labels; 15-ml polypropylene or polyethylene screw-cap tubes; lab gloves ( "metal free"); and, preservatives (as indicated).
The sealed collection cup should be kept in the plastic bag until collection time. The workers should wash their hands with soap and water before receiving the collection cup. The collection cup should not be opened until just before voiding and the cup should be sealed immediately after filling. It is important that the inside of the container and cap are not touched by, or come into contact with, the body, clothing or other surfaces.
For CDU analyzes, the cup is swirled gently to resuspend any solids, and the 15-ml tube is filled with 10-12 ml urine. The CDC recommends the addition of 100 m l concentrated HNO 3 as a preservative before sealing the tube and then freezing the sample. The CTQ recommends minimal handling and does not acidify their interlaboratory urine reference materials prior to shipment, nor do they freeze the sample for shipment. At the CTQ, if the urine sample has much sediment, the sample is acidified in the lab to free any cadmium in the precipitate.
For B2M, the urine sample should be collected directly into a polyethylene bottle previously washed with dilute nitric acid. The pH of the urine should be
measured and adjusted to 8.0 with 0.1 N NaOH immediately following collection. Samples should be frozen and stored at -20 C until testing is performed. The B2M in the samples should be stable for 2 days when stored at 2-8 C, and for at least 2 months at -20 C. Repeated freezing and thawing should be avoided to prevent denaturing the B2M (Pharmacia 1990).
3.4.2 Recommendations for Evaluating Laboratories
Using standard error data and the results of proficiency testing obtained from CTQ, responsible physicians can make an informed choice of which laboratory to select to analyze biological samples. In general, laboratories with small standard errors and little disparity between target and measured values tend to make precise and accurate sample determinations. Estimates of precision provided to the physicians with each set of monitoring results can be compared to previously-reported proficiency and precision estimates. The latest precision estimates should be at least as small as the standard error reported previously by the laboratory. Moreover, there should be no indication that precision is deteriorating (i.e., increasing values for the precision estimates). If precision is deteriorating, physicians may decide to use another laboratory for these analyses. QA/QC information provided by the participating laboratories to physicians can, therefore, assist physicians in evaluating
laboratory performance.
3.4.3 Use and Interpretation of Results
When the responsible physician has received the CDB, CDU and/or B2MU results, these results must be compared to the action levels discussed in the final rule for cadmium. The comparison of the sample results to action levels is straightforward. The measured value reported from the laboratory can be compared directly to the action levels; if the reported value exceeds an action level, the required actions must be initiated.
4.0 BACKGROUND
Cadmium is a naturally-occurring environmental contaminant to which humans are continually exposed in food, water, and air. The average daily intake of cadmium by the U.S. population is estimated to ben 10-20 <<mu>>g/day. Most of this intake is via ingestion, for which absorption is estimated at 4-7% (Kowal et al. 1979). An additional nonoccupational source of cadmium is smoking tobacco; smoking a pack of cigarettes a day adds an additional 2-4 <<mu>>g cadmium to the daily intake, assuming absorption via inhalation of 25-35% (Nordberg and Nordberg 1988; Friberg and Elinder 1988; Travis and Haddock
1980).
Exposure to cadmium fumes and dusts in an occupational setting where air concentrations are 20-50 <<mu>>g/m3 results in an additional daily intake of several hundred micrograms (Friberg and Elinder 1988, p. 563). In such a setting, occupational exposure to cadmium occurs primarily via inhalation, although additional exposure may occur through the ingestion of material via contaminated hands if workers eat or smoke without first washing. Some of the particles that are inhaled initially may be ingested when the material is deposited in the upper respiratory tract, where it may be cleared by mucociliary transport and subsequently swallowed.
Cadmium introduced into the body through inhalation or ingestion is transported by the albumin fraction of the blood plasma to the liver, where it accumulates and is stored principally as a bound form complexed with the protein metallothionein. Metallothionein-bound cadmium is the main form of cadmium subsequently transported to the kidney; it is these 2 organs, the liver and kidney, in which the majority of the cadmium body burden accumulates. As much as one half of the total body burden of cadmium may be found in the kidneys (Nordberg and Nordberg 1988).
Once cadmium has entered the body, elimination is slow; about 0.02% of the body burden is excreted per day via urinary/fecal elimination. The whole-body half-life of cadmium is 10-35 years, decreasing slightly with increasing age (Travis and Haddock 1980).
The continual accumulation of cadmium is the basis for its chronic noncarcinogenic toxicity. This accumulation makes the kidney the target organ in which cadmium toxicity usually is first observed (Piscator 1964). Renal damage may occur when cadmium levels in the kidney cortex approach 200 m g/g wet tissue-weight (Travis and Haddock 1980).
The kinetics and internal distribution of cadmium in the body are complex, and depend on whether occupational exposure to cadmium is ongoing or has terminated. In general, cadmium in blood is related principally to recent cadmium exposure, while cadmium in urine reflects cumulative exposure (i.e., total body burden)(Lauwerys et al. 1976; Friberg and Elinder 1988).
4.1 Health Effects
Studies of workers in a variety of industries indicate that chronic exposure to cadmium may be linked to several adverse health effects including kidney
dysfunction, reduced pulmonary function, chronic lung disease and cancer (Federal Register 1990). The primary sites for cadmium-associated cancer appear to be the lung and the prostate.
Cancer. Evidence for an association between cancer and cadmium exposure comes from both epidemiological studies and animal experiments. Pott (1965) found a statistically significant elevation in the incidence of prostate cancer among a cohort of cadmium workers. Other epidemiology studies also report an elevated incidence of prostate cancer; however, the increases observed in these other studies were not statistically significant (Meridian Research, Inc. 1989).
One study (Thun et al. 1985) contains sufficiently quantitative estimates of cadmium exposure to allow evaluation of dose-response relationships between cadmium exposure and lung cancer. A statistically significant excess of lung cancer attributed to cadmium exposure was found in this study, even after accounting for confounding variables such as coexposure to arsenic and smoking habits (Meridian Research, Inc. 1989).
Evidence for quantifying a link between lung cancer and cadmium exposure comes from a single study (Takenaka et al. 1983). In this study, dose-response relationships developed from animal data were extrapolated to humans using a
variety of models. OSHA chose the multistage risk model for estimating the risk of cancer for humans using these animal data. Animal injection studies also suggest an association between cadmium exposure and cancer, particularly observations of an increased incidence of tumors at sites remote from the point of injection. The International Agency for Research on Cancer (IARC) (Supplement 7, 1987) indicates that this, and related, evidence is sufficient to classify cadmium as an animal carcinogen. However, the results of these injection studies cannot be used to quantify risks attendant to human occupational exposures due to differences in routes of exposure (Meridian Research, Inc. 1989).
Based on the above-cited studies, the U.S. Environmental Protection Agency (EPA) classifies cadmium as "B1," a probable human carcinogen (USEPA 1985). IARC in 1987 recommended that cadmium be listed as a probable human carcinogen.
Kidney Dysfunction. The most prevalent nonmalignant effect observed among workers chronically exposed to cadmium is kidney dysfunction. Initially, such dysfunction is manifested by proteinuria (Meridian Research, Inc. 1989; Roth Associates, Inc. 1989). Proteinuria associated with cadmium exposure is most commonly characterized by excretion of low-molecular weight proteins (15,000- 40,000 MW), accompanied by loss of electrolytes, uric acid, calcium, amino
acids, and phosphate. Proteins commonly excreted include b -2-microglobulin (B2M), retinol-binding protein (RBP), immunoglobulin light chains, and lysozyme. Excretion of low molecular weight proteins is characteristic of damage to the proximal tubules of the kidney (Iwao et al. 1980).
Exposure to cadmium also may lead to urinary excretion of high-molecular weight proteins such as albumin, immunoglobulin G, and glycoproteins (Meridian Research, Inc. 1989; Roth Associates, Inc. 1989). Excretion of high-molecular weight proteins is indicative of damage to the glomeruli of the kidney. Bernard et al. (1979) suggest that cadmium-associated damage to the glomeruli and damage to the proximal tubules of the kidney develop independently of each other, but may occur in the same individual.
Several studies indicate that the onset of low-molecular weight proteinuria is a sign of irreversible kidney damage (Friberg et al. 1974; Roels et al. 1982; Piscator 1984; Elinder et al. 1985; Smith et al. 1986). For many workers, once sufficiently elevated levels of B2M are observed in association with cadmium exposure, such levels do not appear to return to normal even when cadmium exposure is eliminated by removal of the worker from the cadmium-contaminated work environment (Friberg, exhibit 29, 1990).
Some studies indicate that cadmium-induced proteinuria may be progressive; levels of B2MU increase even after cadmium exposure has ceased (Elinder et al. 1985). Other researchers have reached similar conclusions (Frieburg testimony, OSHA docket exhibit 29, Elinder testimony, OSHA docket exhibit 55, and OSHA docket exhibits 8-86B). Such observations are not universal, however (Smith et al. 1986; Tsuchiya 1976). Studies in which proteinuria has not been observed, however, may have initiated the reassessment too early (Meridian Research, Inc.1989; Roth Associates, Inc. 1989; Roels 1989).
| | A quantitative assessment of the risks of developing kidney dysfunction as a result of cadmium exposure was performed using the data from Ellis et al. (1984) and Falck et al. (1983). Meridian Research, Inc. (1989) and Roth Associates, Inc. (1989) employed several mathematical models to evaluate the data from the 2 studies, and the results indicate that cumulative cadmium exposure levels between 5 and 100 m g-years/m 3 correspond with a one-in-a-thousand probability of developing kidney dysfunction. | |
| | When cadmium exposure continues past the onset of early kidney damage (manifested as proteinuria), chronic nephrotoxicity may occur (Meridian Research, Inc. 1989; Roth Associates, Inc. 1989). Uremia, which is the loss of the glomerulus' ability to adequately filter blood, may result. This condition
leads to severe disturbance of electrolyte concentrations, which may result in various clinical complications including atherosclerosis, hypertension, pericarditis, anemia, hemorrhagic tendencies, deficient cellular immunity, bone changes, and other problems. Progression of the disease may require dialysis or a kidney transplant. | |
| | Studies in which animals are chronically exposed to cadmium confirm the renal effects observed in humans (Friberg et al. 1986). Animal studies also confirm cadmium-related problems with calcium metabolism and associated skeletal effects, which also have been observed among humans. Other effects commonly reported in chronic animal studies include anemia, changes in liver morphology, immunosuppression and hypertension. Some of these effects may be associated with cofactors; hypertension, for example, appears to be associated with diet, as well as with cadmium exposure. Animals injected with cadmium also have shown testicular necrosis. | |
4.2 Objectives for Medical Monitoring
In keeping with the observation that renal disease tends to be the earliest clinical manifestation of cadmium toxicity, the final cadmium standard mandates that eligible workers must be medically monitored to prevent this condition (as
well as cadmium-induced cancer). The objectives of medical-monitoring, therefore, are to: Identify workers at significant risk of adverse health effects from excess, chronic exposure to cadmium; prevent future cases of cadmium-induced disease; detect and minimize existing cadmium-induced disease; and, identify workers most in need of medical intervention.
The overall goal of the medical monitoring program is to protect workers who may be exposed continuously to cadmium over a 45-year occupational lifespan. Consistent with this goal, the medical monitoring program should assure that:
1. Current exposure levels remain sufficiently low to prevent the accumulation of cadmium body burdens sufficient to cause disease in the future by monitoring CDB as an indicator of recent cadmium exposure;
2. cumulative body burdens, especially among workers with undefined historical exposures, remain below levels potentially capable of leading to damage and disease by assessing CDU as an indicator of cumulative exposure to cadmium; and,
3. health effects are not occurring among exposed workers by determining B2MU as an early indicator of the onset of cadmium- induced kidney disease.
4.3 Indicators of Cadmium Exposure and Disease
Cadmium is present in whole blood bound to albumin, in erythrocytes, and as a metallothionein-cadmium complex. The metallothionein-cadmium complex that represents the primary transport mechanism for cadmium delivery to the kidney. CDB concentrations in the general, nonexposed population average 1 m g Cd/l whole blood, with smokers exhibiting higher levels (see Section 5.1.6). Data presented in Section 5.1.6 shows that 95% of the general population not occupationally exposed to cadmium have CDB levels less than 5 m g Cd/l.
If total body burdens of cadmium remain low, CDB concentrations indicate recent exposure (i.e., daily intake). This conclusion is based on data showing that cigarette smokers exhibit CDB concentrations of 2-7 m g/l depending on the number of cigarettes smoked per day (Nordberg and Nordberg 1988), while CDB levels for those who quit smoking return to general population values (approximately 1 m g/l) within several weeks (Lauwerys et al. 1976). Based on these observations, Lauwerys et al. (1976) concluded that CDB has a biological half-life of a few weeks to less than 3 months. As indicated in Section 3.1.6, the upper 95th percentile for CDB levels observed among those who are not occupationally exposed to cadmium is 5 m g/l, which suggests that the absolute
upper limit to the range reported for smokers by Nordberg and Nordberg may have been affected by an extreme value (i.e., beyond 2 S above the mean).
Among occupationally-exposed workers, the occupational history of exposure to cadmium must be evaluated to interpret CDB levels. New workers, or workers with low exposures to cadmium, exhibit CDB levels that are representative of recent exposures, similar to the general population. However, for workers with a history of chronic exposure to cadmium, who have accumulated significant stores of cadmium in the kidneys/liver, part of the CDB concentrations appear to indicate body burden. If such workers are removed from cadmium exposure, their CDB levels remain elevated, possibly for years, reflecting prior long-term accumulation of cadmium in body tissues. This condition tends to occur, however, only beyond some threshold exposure value, and possibly indicates the capacity of body tissues to accumulate cadmium which cannot be excreted readily (Friberg and Elinder 1988; Nordberg and Nordberg 1988).
CDU is widely used as an indicator of cadmium body burdens (Nordberg and Nordberg 1988). CDU is the major route of elimination and, when CDU is measured, it is commonly expressed either as m g Cd/l urine (unadjusted), m g Cd/l urine (adjusted for specific gravity), or m g Cd/g CRTU (see Section 5.2.1). The metabolic model for CDU is less complicated than CDB, since CDU is
dependent in large part on the body (i.e., kidney) burden of cadmium. However, a small proportion of CDU still be attributed to recent cadmium exposure, particularly if exposure to high airborne concentrations of cadmium occurred. Note that CDU is subject to larger interindividual and day-to-day variations than CDB, so repeated measurements are recommended for CDU evaluations.
CDU is bound principally to metallothionein, regardless of whether the cadmium originates from metallothionein in plasma or from the cadmium pool accumulated in the renal tubules. Therefore, measurement of metallothionein in urine may provide information similar to CDU, while avoiding the contamination problems that may occur during collection and handling urine for cadmium analysis (Nordberg and Nordberg 1988). However, a commercial method for the determination of metallothionein at the sensitivity levels required under the final cadmium rule is not currently available; therefore, analysis of CDU is recommended.
Among the general population not occupationally exposed to cadmium, CDU levels average less than 1 m g/l (see Section 5.2.7). Normalized for creatinine (CRTU), the average CDU concentration of the general population is less than 1 m g/g CRTU. As cadmium accumulates over the lifespan, CDU increases with age. Also, cigarette smokers may eventually accumulate twice the cadmium body burden
of nonsmokers, CDU is slightly higher in smokers than in nonsmokers, even several years after smoking cessation (Nordberg and Nordberg 1988). Despite variations due to age and smoking habits, 95% of those not occupationally exposed to cadmium exhibit levels of CDU less than 3 m g/g CRTU (based on the data presented in Section 5.2.7).
About 0.02% of the cadmium body burden is excreted daily in urine. When the critical cadmium concentration (about 200 ppm) in the kidney is reached, or if there is sufficient cadmium-induced kidney dysfunction, dramatic increases in CDU are observed (Nordberg and Nordberg 1988). Above 200 ppm, therefore, CDU concentrations cease to be an indicator of cadmium body burden, and are instead an index of kidney failure.
Proteinuria is an index of kidney dysfunction, and is defined by OSHA to be a material impairment. Several small proteins may be monitored as markers for proteinuria. Below levels indicative of proteinuria, these small proteins may be early indicators of increased risk of cadmium-induced renal tubular disease. Analytes useful for monitoring cadmium-induced renal tubular damage include:
1. b -2-Microglobulin (B2M), currently the most widely used assay for detecting kidney dysfunction, is the best characterized analyte available (Iwao et al.
1980; Chia et al. 1989);
2. Retinol Binding Protein (RBP) is more stable than B2M in acidic urine (i.e., B2M breakdown occurs if urinary pH is less than 5.5; such breakdown may result in false [i.e., low] B2M values [Bernard and Lauwerys, 1990]);
3. N-Acetyl-B-Glucosaminidase (NAG) is the analyte of an assay that is simple, inexpensive, reliable, and correlates with cadmium levels under 10 m g/g CRTU, but the assay is less sensitive than RBP or B2M (Kawada et al. 1989);
4. Metallothionein (MT) correlates with cadmium and B2M levels, and may be a better predictor of cadmium exposure than CDU and B2M (Kawada et al. 1989);
5. Tamm-Horsfall Glycoprotein (THG) increases slightly with elevated cadmium levels, but this elevation is small compared to increases in urinary albumin, RBP, or B2M (Bernard and Lauwerys 1990);
6. Albumin (ALB), determined by the biuret method, is not sufficiently sensitive to serve as an early indicator of the onset of renal disease (Piscator 1962);
7. Albumin (ALB), determined by the Amido Black method, is sensitive and reproducible, but involves a time-consuming procedure (Piscator 1962);
8. Glycosaminoglycan (GAG) increases among cadmium workers, but the significance of this effect is unknown because no relationship has been found between elevated GAG and other indices of tubular damage (Bernard and Lauwerys 1990);
9. Trehalase seems to increase earlier than B2M during cadmium exposure, but the procedure for analysis is complicated and unreliable (Iwata et al. 1988); and,
10. Kallikrein is observed at lower concentrations among cadmium-exposed workers than among normal controls (Roels et al. 1990).
Of the above analytes, B2M appears to be the most widely used and best characterized analyte to evaluate the presence/absence, as well as the extent of, cadmium-induced renal tubular damage (Kawada, Koyama, and Suzuki 1989; Shaikh and Smith 1984; Nogawa 1984). However, it is important that samples be collected and handled so as to minimize B2M degradation under acidic urine conditions.
The threshold value of B2MU commonly used to indicate the presence of kidney damage 300 m g/g CRTU (Kjellstrom et al. 1977a; Buchet et al. 1980; and Kowal and Zirkes 1983). This value represents the upper 95th or 97.5th percentile level of urinary excretion observed among those without tubular dysfunction (Elinder, exbt L-140-45, OSHA docket H057A). In agreement with these conclusions, the data presented in Section 5.3.7 of this protocol generally indicate that the level of 300 m g/g CRTU appears to define the boundary for kidney dysfuncion. It is not clear, however, that this level represents the upper 95th percentile of values observed among those who fail to demonstrate proteinuria effects.
Although elevated B2MU levels appear to be a fairly specific indicator of disease associated with cadmium exposure, other conditions that may lead to elevated B2MU levels include high fevers from influenza, extensive physical exercise, renal disease unrelated to cadmium exposure, lymphomas, and AIDS (Iwao et al. 1980; Schardun and van Epps 1987). Elevated B2M levels observed in association with high fevers from influenza or from extensive physical exercise are transient, and will return to normal levels once the fever has abated or metabolic rates return to baseline values following exercise. The other conditions linked to elevated B2M levels can be diagnosed as part of a
properly- designed medical examination. Consequently, monitoring B2M, when accompanied by regular medical examinations and CDB and CDU determinations (as indicators of present and past cadmium exposure), may serve as a specific, early indicator of cadmium- induced kidney damage.
4.4 Criteria for Medical Monitoring of Cadmium Workers
Medical monitoring mandated by the final cadmium rule includes a combination of regular medical examinations and periodic monitoring of 3 analytes: CDB, CDU and B2MU. As indicated above, CDB is monitored as an indicator of current cadmium exposure, while CDU serves as an indicator of the cadmium body burden; B2MU) is assessed as an early marker of irreversible kidney damage and disease.
The final cadmium rule defines a series of action levels that have been developed for each of the 3 analytes to be monitored. These action levels serve to guide the responsible physician through a decision-making process. For each action level that is exceeded, a specific response is mandated. The sequence of action levels, and the attendant actions, are described in detail in the final cadmium rule.
Other criteria used in the medical decision-making process relate to tests
performed during the medical examination (including a determination of the ability of a worker to wear a respirator). These criteria, however, are not affected by the results of the analyte determinations addressed in the above paragraphs and, consequently, will not be considered further in these guidelines.
4.5 Defining to Quality and Proficiency of the Analyte Determinations
As noted above in Sections 2 and 3, the quality of a measurement should be defined along with its value to properly interpret the results. Generally, it is necessary to know the accuracy and the precision of a measurement before it can be properly evaluated. The precision of the data from a specific laboratory indicates the extent to which the repeated measurements of the same sample vary within that laboratory. The accuracy of the data provides an indication of the extent to which these results deviate from average results determined from many laboratories performing the same measurement (i.e., in the absence of an independent determination of the true value of a measurement). Note that terms are defined operationally relative to the manner in which they will be used in this protocol. Formal definitions for the terms in italics used in this section can be found in the list of definitions (Section 2).
Another data quality criterion required to properly evaluate measurement results is the limit of detection of that measurement. For measurements to be useful, the range of the measurement which is of interest for biological monitoring purposes must lie entirely above the limit of detection defined for that measurement.
The overall quality of a laboratory's results is termed the performance of that laboratory. The degree to which a laboratory satisfies a minimum performance level is referred to as the proficiency of the laboratory. A successful medical monitoring program, therefore, should include procedures developed for monitoring and recording laboratory performance; these procedures can be used to identify the most proficient laboratories.
5.0 Overview of Medical Monitoring Tests for CDB, CDU, B2MU and CRTU
To evaluate whether available methods for assessing CDB, CDU, B2MU and CRTU are adequate for determining the parameters defined by the proposed action levels, it is necessary to review procedures available for sample collection, preparation and analysis. A variety of techniques for these purposes have been used historically for the determination of cadmium in biological matrices (including CDB and CDU), and for the determination of specific proteins in
biological matrices (including B2MU). However, only the most recent techniques are capable of satisfying the required accuracy, precision and sensitivity (i.e., limit of detection) for monitoring at the levels mandated in the final cadmium rule, while still facilitating automated analysis and rapid processing.
5.1 Measuring Cadmium in Blood (CDB)
Analysis of biological samples for cadmium requires strict analytical discipline regarding collection and handling of samples. In addition to occupational settings, where cadmium contamination would be apparent, cadmium is a ubiquitous environmental contaminant, and much care should be exercised to ensure that samples are not contaminated during collection, preparation or analysis. Many common chemical reagents are contaminated with cadmium at concentrations that will interfere with cadmium analysis; because of the widespread use of cadmium compounds as colored pigments in plastics and coatings, the analyst should continually monitor each manufacturer's chemical reagents and collection containers to prevent contamination of samples.
Guarding against cadmium contamination of biological samples is particularly important when analyzing blood samples because cadmium concentrations in blood samples from nonexposed populations are generally less than 2 m g/l (2 ng/ml),
while occupationally-exposed workers can be at medical risk to cadmium toxicity if blood concentrations exceed 5 m g/l (ACGIH 1991 and 1992). This narrow margin between exposed and unexposed samples requires that exceptional care be used in performing analytic determinations for biological monitoring for occupational cadmium exposure.
Methods for quantifying cadmium in blood have improved over the last 40 years primarily because of improvements in analytical instrumentation. Also, due to improvements in analytical techniques, there is less need to perform extensive multi-step sample preparations prior to analysis. Complex sample preparation was previously required to enhance method sensitivity (for cadmium), and to reduce interference by other metals or components of the sample.
5.1.1 Analytical Techniques Used to Monitor Cadmium in Biological Matrices
TABLE 3 - COMPARISON OF ANALYTICAL PROCEDURES/INSTRUMENTATION FOR
DETERMINATION OF CADMIUM IN BIOLOGICAL SAMPLES
________________________________________________________________________
| | | |
Analytical | Limit |Specified | |
procedure |of det-|biological| Reference | Comments
|ection |matrix | |
|[ng/(g | | |
|or ml)]| | |
______________|_______|__________|_____________|_________________________
| | | |
Flame Atomic |> or = |Any Matrix| Perkin- |Not sensitive enough for
Absorption | 1.0 | | Elmer | biomonitoring without
Spectroscopy | | | (1982)... | extensive sample
(FAAS). | | | | digestion metal
| | | | chelation and organic
| | | | solvent extraction.
Graphite | 0.04 |Urine ... | Pruszkowska |Methods of choice for
Furnace | | |et al (1983) | routine cadmium analysis.
Atomic | | | |
Absorption |> or = |Blood ... | Stoeppler |
Spectroscopy | 0.20 | | and Brandt |
(GFAAS). | | | |
| | | (1980)..... |
Inductively- | 2.0 |Any matrix| NIOSH |Requires extensive sample
Coupled Argon| | | (1984A).... | preparation and
Plasma Atomic| | | | concentration of metal
Emission | | | | with chelating resin.
Spectroscopy | | | | Advantage is simltaneous
(ICAP AES). | | | | analyses for as many as
| | | | 10 metals from 1 sample.
Neutron | 1.5 |In vivo |Ellis et al. |Only available in vivo
Activation | | (liver)..| (1983)..... | method for direct
Gamma | | | | determination of cadmium
Spectroscopy | | | | body tissue burdens;
(NA). | | | | expensive; absolute
| | | | determination of cadmium
| | | | in reference materials.
Isotope |< 1.0 |Any matrix|Michiels and |Suitable for absolute
Dilution | | | DeBievre | determination of cadmium
Mass | | | (1986)..... | in reference materials;
Spectroscopy | | | | expensive.
(IDMS). | | | |
Differential |< 1.0 |Any matrix|Stoeppler |Suitable for absolute
Pulse Anodic | | | and Brandt | determination of cadmium
Stripping | | | (1980)..... | in reference materials;
Voltammetry | | | | efficient method to check
(DPASV). | | | | accuracy of analytical
| | | | method.
______________|_______|__________|_____________|___________________________
| | A number of analytical techniques have been used for determining cadmium concentrations in biological materials. A summary of the characteristics of the most widely employed techniques is presented in Table 3. The technique most
suitable for medical monitoring for cadmium is atomic absorption spectroscopy (AAS). | |
| | To obtain a measurement using AAS, a light source (i.e., hollow cathode or lectrode-free discharge lamp) containing the element of interest as the cathode, is energized and the lamp emits a spectrum that is unique for that element. This light source is focused through a sample cell, and a selected wavelength is monitored by a monochrometer and photodetector cell. Any ground state atoms in the sample that match those of the lamp element and are in the path of the emitted light may absorb some of the light and decrease the amount of light that reaches the photodetector cell. The amount of light absorbed at each characteristic wavelength is proportional to the number of ground state atoms of the corresponding element that are in the pathway of the light between the source and detector. | |
| | To determine the amount of a specific metallic element in a sample using AAS, the sample is dissolved in a solvent and aspirated into a high-temperature flame as an aerosol. At high temperatures, the solvent is rapidly evaporated or decomposed and the solute is initially solidified; the majority of the sample elements then are transformed into an atomic vapor. Next, a light beam is focused above the flame and the amount of metal in the sample can be determined
by measuring the degree of absorbance of the atoms of the target element released by the flame at a characteristic wavelength. | |
| | A more refined atomic absorption technique, flameless AAS, substitutes an electrothermal, graphite furnace for the flame. An aliquot (10-100 m l) of the sample is pipetted into the cold furnace, which is then heated rapidly to generate an atomic vapor of the element. | |
| | AAS is a sensitive and specific method for the elemental analysis of metals; its main drawback is nonspecific background absorbtion and scattering of the light beam by particles of the sample as it decomposes at high temperatures; nonspecific absorbance reduces the sensitivity of the analytical method. The problem of nonspecific absorbance and scattering can be reduced by extensive sample pretreatment, such as ashing and/or acid digestion of the sample to reduce its organic content. | |
| | Current AAS instruments employ background correction devices to adjust electronically for background absorbtion and scattering. A common method to correct for background effects is to use a deuterium arc lamp as a second light source. A continuum light source, such as the deuterium lamp, emits a broad spectrum of wavelengths instead of specific wavelengths characteristic of a
particular element, as with the hollow cathode tube. With this system, light from the primary source and the continuum source are passed alternately through the sample cell. The target element effectively absorbs light only from the primary source (which is much brighter than the continuum source at the characteristic wavelengths), while the background matrix absorbs and scatters light from both sources equally. Therefore, when the ratio of the two beams is measured electronically, the effect of nonspecific background absorption and scattering is eliminated. A less common, but more sophisticated, background correction system is based on the Zeeman effect, which uses a magnetically-activated light polarizer to compensate electronically for nonspecific absorbtion and scattering. | |
| | Atomic emission spectroscopy with inductively-coupled argon plasma (AES-ICAP) is widely used to analyze for metals. With this instrument, the sample is aspirated into an extremely hot argon plasma flame, which excites the metal atoms; emission spectra specific for the sample element then are generated. The quanta of emitted light passing through a monochrometer are amplified by photomultiplier tubes and measured by a photodetector to determine the amount of metal in the sample. An advantage of AES- ICAP over AAS is that multi-elemental analyses of a sample can be performed by simultaneously measuring specific elemental emission energies. However, AES-ICAP lacks the sensitivity
of AAS, exhibiting a limit of detection which is higher than the limit of detection for graphite-furnace AAS (Table 3). | |
| | Neutron activation (NA) analysis and isotope dilution mass spectrometry (IDMS) are 2 additional, but highly specialized, methods that have been used for cadmium determinations. These methods are expensive because they require elaborate and sophisticated instrumentation. | |
| | NA analysis has the distinct advantage over other analytical methods of being able to determine cadmium body burdens in specific organs (e.g., liver, kidney) in vivo (Ellis et al. 1983). Neutron bombardment of the target transforms cadmium- 13 to cadmium-114, which promptly decays (< 10 -14 sec) to its ground state, emitting gamma rays that are measured using large gamma detectors; appropriate shielding and instrumentation are required when using this method. | |
| | IDMS analysis, a definitive but laborious method, is based on the change in the ratio of 2 isotopes of cadmium (cadmium 111 and 112) that occurs when a known amount of the element (with an artificially altered ratio of the same isotopes [i.e., a cadmium 111 "spike"] is added to a weighed aliquot of the sample (Michiels and De Bievre 1986). | |
5.1.2 Methods Developed for CDB Determinations
| | A variety of methods have been used for preparing and analyzing CDB samples; most of these methods rely on one of the analytical techniques described above. Among the earliest reports, Princi (1947) and Smith et al. (1955) employed a colorimetric procedure to analyze for CDB and CDU. Samples were dried and digested through several cycles with concentrated mineral acids (HNO 3 and H 2 SO 4) and hydrogen peroxide (H 2 O 2). The digest was neutralized, and the cadmium was complexed with diphenylthiocarbazone and extracted with chloroform. The dithizone-cadmium complex then was quantified using a spectrometer. | |
| | Colorimetric procedures for cadmium analyses were replaced by methods based on atomic absorption spectroscopy (AAS) in the early 1960s, but many of the complex sample preparation procedures were retained. Kjellstrom (1979) reports that in Japanese, American and Swedish laboratories during the early 1970s, blood samples were wet ashed with mineral acids or ashed at high temperature and wetted with nitric acid. The cadmium in the digest was complexed with metal chelators including diethyl dithiocarbamate (DDTC), ammonium pyrrolidine dithiocarbamate (APDC) or diphenylthiocarbazone (dithizone) in ammonia-citrate
buffer and extracted with methyl isobutyl ketone (MIBK). The resulting solution then was analyzed by flame AAS or graphite- furnace AAS for cadmium determinations using deuterium-lamp background correction. | |
| | In the late 1970s, researchers began developing simpler preparation procedures. Roels et al. (1978) and Roberts and Clark (1986) developed simplified digestion procedures. Using the Roberts and Clark method, a 0.5 ml aliquot of blood is collected and transferred to a digestion tube containing 1 ml concentrated HNO 3. The blood is then digested at 110 ° C for 4 hours. The sample is reduced in volume by continued heating, and 0.5 ml 30% H 2 O 2 is added as the sample dries. The residue is dissolved in 5 ml dilute (1%) HNO 3, and 20 m l of sample is then analyzed by graphite-furnace AAS with deuterium-background correction. | |
| | The current trend in the preparation of blood samples is to dilute the sample and add matrix modifiers to reduce background interference, rather than digesting the sample to reduce organic content. The method of Stoeppler and Brandt (1980), and the abbreviated procedure published in the American Public Health Association's (APHA) Methods for Biological Monitoring (1988), are straightforward and are nearly identical. For the APHA method, a small aliquot (50-300 m l) of whole blood that has been stabilized with
ethylenediaminetetraacetate (EDTA) is added to 1.0 ml 1M HNO 3, vigorously shaken and centrifuged. Aliquots (10-25 m l) of the supernatant then are then analyzed by graphite-furnace AAS with appropriate background correction. | |
| | Using the method of Stoeppler and Brandt (1980), aliquots (50-200 m l) of whole blood that have been stabilized with EDTA are pipetted into clean polystyrene tubes and mixed with 150-600 m l of 1 M HNO 3. After vigorous shaking, the solution is centrifuged and a 10-25 m l aliquot of the supernatant then is analyzed by graphite-furnace AAS with appropriate background correction. | |
| | Claeys-Thoreau (1982) and DeBenzo et al. (1990) diluted blood samples at a ratio of 1:10 with a matrix modifier (0.2% Triton X-100, a wetting agent) for direct determinations of CDB. DeBenzo et al. also demonstrated that aqueous standards of cadmium, instead of spiked, whole-blood samples, could be used to establish calibration curves if standards and samples are treated with additional small volumes of matrix modifiers (i.e., 1% HNO 3, 0.2% ammonium hydrogenphosphate and 1 mg/ml magnesium salts.) | |
| | These direct dilution procedures for CDB analysis are simple and rapid. Laboratories can process more than 100 samples a day using a dedicated graphite-furnace AAS, an auto-sampler, and either a Zeeman- or a deuterium-
background correction system. Several authors emphasize using optimum settings for graphite-furnace temperatures during the drying, charring, and atomization processes associated with the flameless AAS method, and the need to run frequent QC samples when performing automated analysis. | |
5.1.3 Sample Collection and Handling
Sample collection procedures are addressed primarily to identify ways to minimize the degree of variability that may be introduced by sample collection during medical monitoring. It is unclear at this point the extent to which collection procedures contribute to variability among CDB samples. Sources of variation that may result from sampling procedures include time-of-day effects and introduction of external contamination during the collection process. To minimize these sources, strict adherence to a sample collection protocol is recommended. Such a protocol must include provisions for thorough cleaning of the site from which blood will be extracted; also, every effort should be made to collect samples near the same time of day. It is also important to recognize that under the recent OSHA blood-borne pathogens standard (29 CFR 1910.1030), blood samples and certain body fluids must be handled and treated as if they are infectious.
5.1.4 Best Achievable Performance
The best achievable performance using a particular method for CDB determinations is assumed to be equivalent to the performance reported by research laboratories in which the method was developed.
For their method, Roberts and Clark (1986) demonstrated a limit of detection of 0.4 m g Cd/l in whole blood, with a linear response curve from 0.4 to 16.0 m g Cd/l. They report a coefficient of variation (CV) of 6.7% at 8.0 m g/l.
The APHA (1988) reports a range of 1.0-25 m g/l, with a CV of 7.3% (concentration not stated). Insufficient documentation was available to critique this method.
Stoeppler and Brandt (1980) achieved a detection limit of 0.2 m g Cd/l whole blood, with a linear range of 0.4-12.0 m g Cd/l, and a CV of 15-30%, for samples at 1.0aeg m l. Improved precision (CV of 3.8%) was reported for CDB concentrations at 9.3 m g/l.
5.1.5 General Method Performance
For any particular method, the performance expected from commercial laboratories may be somewhat lower than that reported by the research laboratory in which the method was developed. With participation in appropriate proficiency programs and use of a proper in-house QA/QC program incorporating provisions for regular corrective actions, the performance of commercial laboratories is expected to approach that reported by research laboratories. Also, the results reported for existing proficiency programs serve as a gauge of the likely level of performance that currently can be expected from commercial laboratories offering these analyses.
Weber (1988) reports on the results of the proficiency program run by the Centre de Toxicologie du Quebec (CTQ). As indicated previously, participants in that program receive 18 blood samples per year having cadmium concentrations ranging from 0.2-20 m g/l. Currently, 76 laboratories are participating in this program. The program is established for several analytes in addition to cadmium, and not all of these laboratories participate in the cadmium proficiency-testing program.
Under the CTQ program, cadmium results from individual laboratories are compared against the consensus mean derived for each sample. Results indicate that after receiving 60 samples (i.e., after participation for approximately
three years), 60% of the laboratories in the program are able to report results that fall within + 1 m g/l or 15% of the mean, whichever is greater. (For this procedure, the 15% criterion was applied to concentrations exceeding 7 m g/l.) On any single sample of the last 20 samples, the percentage of laboratories falling within the specified range is between 55 and 80%.
The CTQ also evaluates the performance of participating laboratories against a less severe standard: + 2 m g/l or 15% of the mean, whichever is greater (Weber 1988); 90% of participating laboratories are able to satisfy this standard after approximately 3 years in the program. (The 15% criterion is used for concentrations in excess of 13 m g/l.) On any single sample of the last 15 samples, the percentage of laboratories falling within the specified range is between 80 and 95% (except for a single test for which only 60% of the laboratories achieved the desired performance).
Based on the data presented in Weber (1988), the CV for analysis of CDB is nearly constant at 20% for cadmium concentrations exceeding 5 m g/l, and increases for cadmium concentrations below 5 m g/l. At 2 m g/l, the reported CV rises to approximately 40%. At 1 m g/l, the reported CV is approximately 60%.
Participating laboratories also tend to overestimate concentrations for samples
exhibiting concentrations less than 2 m g/l (see Figure 11 of Weber 1988). This problem is due in part to the proficiency evaluation criterion that allows reporting a minimum + 2.0 m g/l for evaluated CDB samples. There is currently little economic or regulatory incentive for laboratories participating in the CTQ program to achieve greater accuracy for CDB samples containing cadmium at concentrations less than 2.0 m g/l, even if the laboratory has the experience and competency to distinguish among lower concentrations in the samples obtained from the CTQ.
The collective experience of international agencies and investigators demonstrate the need for a vigorous QC program to ensure that CDB values reported by participating laboratories are indeed reasonably accurate. As Friberg (1988) stated:
"Information about the quality of published data has often been lacking. This is of concern as assessment of metals in trace concentrations in biological media are fraught with difficulties from the collection, handling, and storage of samples to the chemical analyses. This has been proven over and over again from the results of interlaboratory testing and quality control exercises. Large variations in results were reported even from "experienced" laboratories."
The UNEP/WHO global study of cadmium biological monitoring set a limit for CDB accuracy using the maximum allowable deviation method at Y=X + (0.1X+1) for a targeted concentration of 10 m g Cd/l (Friberg and Vahter 1983). The performance of participating laboratories over a concentration range of 1.5-12 m g/l was reported by Lind et al. (1987). Of the 3 QC runs conducted during 1982 and 1983, 1 or 2 of the 6 laboratories failed each run. For the years 1983 and 1985, between zero and 2 laboratories failed each of the consecutive QC runs.
In another study (Vahter and Friberg 1988), QC samples consisting of both external (unknown) and internal (stated) concentrations were distributed to laboratories participating in the epidemiology research. In this study, the maximum acceptable deviation between the regression analysis of reported results and reference values was set at Y=X + (0.05X+0.2) for a concentration range of 0.3-5.0 m g Cd/l. It is reported that only 2 of 5 laboratories had acceptable data after the first QC set, and only 1of 5 laboratories had acceptable data after the second QC set. By the fourth QC set, however, all 5 laboratories were judged proficient.
The need for high quality CDB monitoring is apparent when the toxicological and
biological characteristics of this metal are considered; an increase in CDB from 2 to 4 m g/l could cause a doubling of the cadmium accumulation in the kidney, a critical target tissue for selective cadmium accumulation (Nordberg and Nordberg 1988).
Historically, the CDC's internal QC program for CDB cadmium monitoring program has found achievable accuracy to be + 10% of the true value at CDB concentrations > 5.0 m g/l (Paschal 1990). Data on the performance of laboratories participating in this program currently are not available.
5.1.6 Observed CDB Concentrations
As stated in Section 4.3, CDB concentrations are representative of ongoing levels of exposure to cadmium. Among those who have been exposed chronically to cadmium for extended periods, however, CDB may contain a component attributable to the general cadmium body burden.
5.1.6.1 CDB concentrations among unexposed samples
Numerous studies have been conducted examining CDB concentrations in the general population, and in control groups used for comparison with cadmium-
exposed workers. A number of reports have been published that present erroneously high values of CDB (Nordberg and Nordberg 1988). This problem was due to contamination of samples during sampling and analysis, and to errors in analysis. Early AAS methods were not sufficiently sensitive to accurately estimate CDB concentrations.
Table 4 presents results of recent studies reporting CDB levels for the general U.S. population not exposed occupationally to cadmium. Other surveys of tissue cadmium using U.S. samples and conducted as part of a cooperative effort among Japan, Sweden and the U.S., did not collect CDB data because standard analytical methodologies were unavailable, and because of analytic problems (Kjellstrom 1979; SWRI 1978).
TABLE 4. -- BLOOD CADMIUM CONCENTRATIONS OF U.S. POPULATION
NOT OCCUPATIONALLY EXPOSED TO CADMIUM(a)
_________________________________________________________________
| | | | | | |
| No. | | | | Arithmetic | Absolute |
Study | in | | | Smoking | mean | range |
No. | study | Sex | Age | habits | (+/-S.D.) | or |
| (n) | | | (b) | (c) | (95% Cl) |
| | | | | | (d) |
_______|_______|_____|_________|_________|____________|___________|
| | | | | | |
1 .... | 80 | M | 4 to 69 | NS,S | 1.13 | 0.35-3.3 |
| 88 | F | 4 to 69 | NS, S | 1.03 | 0.21-3.3 |
| 115 | M/F | 4 to 69 | NS, | 0.95 | 0.21-3.3 |
| 31 | M/F | 4 to 69 | S | 1.54 | 0.4-3.3 |
2 .... | 10 | M | Adults. | (?) | 2.0+/-2.1 | (0.5-5.0) |
| | | | | | |
3 .... | 24 | M | Adults. | NS | | |
| | | | | | |
| | | | | | |
| 20 | M | Adults. | S | | |
| 64 | F | Adults. | NS | | |
| 39 | F | Adults. | S | | |
4 .... | 32 | M | Adults. | S,NS | | |
| | | | | | |
5 .... | 35 | M | Adults. | (?) | 2.1+/-2.1 | (0.5-7.3) |
| | | | | | |
_______|_______|_____|_________|_________|____________|___________|
TABLE 4. -- BLOOD CADMIUM CONCENTRATIONS OF U.S. POPULATION
NOT OCCUPATIONALLY EXPOSED TO CADMIUM(a)
[Continued]
___________________________________________________________________
| | | | |
| Geometric | Lower | Upper | |
Study | mean | 95th | 95th | |
No. | (+/- GSD) | percentile | percentile | Reference |
| (c) | of | of | |
| | distribution | distribution | |
| | (f) | (f) | |
_______|_____________|______________|______________|_______________|
| | | | |
1 .... | 0.98+/-1.71 | 0.4 | 2.4 | Kowal et al. |
| 0.91+/-1.63 | 0.4 | 2.0 | (1979). |
| 0.85+/-1.59 | 0.4 | 1.8 | |
| 1.37+/-1.65 | 0.6 | 3.2 | |
2 .... | | (g)(0) | (g)(5.8) | Ellis et al. |
| | | | (1983) |
3 .... | 0.6+/-1.87 | 0.2 | 1.8 | Frieberg and |
| | | | Vahter |
| | | | (1983). |
| 1.2+/-2.13 | 0.3 | 4.4 | |
| 0.5+/-1.85 | 0.2 | 1.4 | |
| 0.8+/-2.22 | 0.2 | 3.1 | |
4 .... | 1.2+/-2.0 | 0.4 | 3.9 | Thun et al. |
| | | | (1989). |
5 .... | | (g)(0) | (g)(5.6) | Mueller et al.|
| | | | (1989) |
_______|_____________|______________|______________|_______________|
Arithmetic and/or geometric means and standard deviations are provided in Table 4 for measurements among the populations defined in each study listed. The range of reported measurements and/or the 95% upper and lower confidence intervals for the means are presented when this information was reported in a study. For studies reporting either an arithmetic or geometric standard
deviation along with a mean, the lower and upper 95th percentile for the distribution also were derived and reported in the table.
The data provided in Table 4 from Kowal et al. (1979) are from studies conducted between 1974 and 1976 evaluating CDB levels for the general population in Chicago, and are considered to be representative of the U.S. population. These studies indicate that the average CDB concentration among those not occupationally exposed to cadmium is approximately 1 m g/l.
In several other studies presented in Table 4, measurements are reported separately for males and females, and for smokers and nonsmokers. The data in this table indicate that similar CDB levels are observed among males and females in the general population, but that smokers tend to exhibit higher CDB levels than nonsmokers. Based on the Kowal et al. (1979) study, smokers not occupationally exposed to cadmium exhibit an average CDB level of 1.4 m g/l.
In general, nonsmokers tend to exhibit levels ranging to 2 m g/l, while levels observed among smokers range to 5aeg/l. Based on the data presented in Table 4, 95% of those not occupationally exposed to cadmium exhibit CDB levels less than 5 m g/l.
5.1.6.2 CDB concentrations among exposed workers.
Table 5 is a summary of results from studies reporting CDB levels among workers exposed to cadmium in the work place. As in Table 4, arithmetic and/or geometric means and standard deviations are provided if reported in the listed studies. The absolute range, or the 95% confidence interval around the mean, of the data in each study are provided when reported. In addition, the lower and upper 95th percentile of the distribution are presented for each study i which a mean and corresponding standard deviation were reported. Table 5 also provides estimates of the duration, and level, of exposure to cadmium in the work place if these data were reported in the listed studies. The data presented in Table 5 suggest that CDB levels are dose related. Sukuri et al. (1983) show that higher CDB levels are observed among workers experiencing higher work place exposure. This trend appears to be true of every the studies listed in the table.
TABLE 5. - BLOOD CADMIUM IN WORKERS EXPOSED TO CADMIUM
IN THE WORKPLACE
________________________________________________________________
| | | | |
| Work | | | Mean |
| environment | | Employment | concentration |
Study | (worker | Number | in | of cadmium |
number | population | in | years | in air |
| monitored) | study | (mean) | (ug/m(3)) |
| | | | |
________|________________|________|____________|________________|
| | | | |
1 ..... | Ni-Cd battery | | 3-40 | < than = to 90 |
| plant and Cd | | | |
| production | | | |
| plant: | | | |
| (Workers | | | |
| without | | | |
| kidney | | | |
| lesions).....| 96 | .......... | .............. |
| (Workers with | | | |
| kidney | | | |
| lesions).....| 25 | .......... | .............. |
| | | | |
2 ..... | Ni-Cd battery | | | |
| plant: | | | |
| | | | |
| (Smokers) .....| 7 | (5) | 10.1 |
| (Nonsmokers)...| 8 | (9) | 7.0 |
| | | | |
3 ..... | Cadmium alloy | | | |
| plant: | | | |
| | | | |
| (High exposure | | | |
| group)........| 7 | (10.6) | [1,000-5 yrs; |
| (Low exposure | | | 40-5 yrs] |
| group)........| 9 | (7.3) | |
| | | | |
4 ..... | Retrospective | 19 | 15-41 | |
| study of | | | |
| wokers with | | | |
| renal | | | |
| problems: | | | |
| | | | |
| (Before | | | |
| removal).....| ...... | (27.2) | .............. |
| (After | | | |
| removal).....| ...... | (g)(4.2) | .............. |
| | | | |
5 ..... | Cadmium | | | |
| production | | | |
| plant: | | | |
| | | | |
| (Workers | | | |
| without renal| | | |
| dysfunction).| 33 | 1-34 | .............. |
| (Workers with | | | |
| renal | 18 | 10-34 | .............. |
| dysfunction).| | | |
| | | | |
6 ..... | Cd-Cu alloy | | | |
| plant.........| 75 | Up to 39 | .............. |
| | | | |
| | | | |
7 ..... | Cadmium | | | |
| recovery | | | |
| operation - | | | |
| Current (19) | | | |
| and former | | | |
| (26) workers. | 45 | (19.0) | .............. |
| | | | |
8 ..... | Cadmium | | | |
| recovery | | | |
| operation | 40 | .......... | .............. |
________|________________|________|____________|________________|
TABLE 5. - BLOOD CADMIUM IN WORKERS EXPOSED TO CADMIUM
IN THE WORKPLACE
[Continued]
____________________________________________________________________
| |
| Work | Concentrations of Cadmium in Blood(a)
| environment |__________________________________________
Study | (worker | | |
number | population | Arithmetic | Absolute | Geometric
| monitored) | mean | range or | mean
| | (+/- S.D.)(b) |(95% C.I.)(c)| (GSD)(d)
________|________________|_______________|_____________|__________
| | | |
1 ..... | Ni-Cd battery | | |
| plant and Cd | | |
| production | | |
| plant: | | |
| | | |
| (Workers | | |
| without | | |
| kidney | | |
| lesions).....| 21.4 +/- 1.9 | ........... | .........
| (Workers with | | |
| kidney | | |
| lesions).....| 38.8 +/- 3.8 | ........... | .........
| | | |
2 ..... | Ni-Cd battery | | |
| plant: | | |
| | | |
| (Smokers) .....| 22.7 | 7.3 - 67.2 |
| (Nonsmokers)...| 7.0 | 4.9 - 10.5 |
| | | |
3 ..... | Cadmium alloy | | |
| plant: | | |
| | | |
| (High exposure | | |
| group)........| 20.8 +/- 7.1 | ........... | ........
| (Low exposure | | |
| group)........| 7.1 +/- 1.1 | ........... | ........
| | | |
4 ..... | Retrospective | | |
| study of | | |
| wokers with | | |
| renal | | |
| problems: | | |
| | | |
| (Before | | |
| removal).....| 39.9 +/- 3.7 | 11 - 179 | .........
| (After | | |
| removal).....| 14.1 +/- 5.6 | 5.7 - 27.4 | .........
| | | |
5 ..... | Cadmium | | |
| production | | |
| plant: | | |
| | | |
| (Workers | | |
| without renal| | |
| dysfunction).| 15 +/- 5.7 | 7 - 31 | .........
| (Workers with | | |
| renal | | |
| dysfunction).| 24 +/- 8.5 | 10 - 34 | .........
| | | |
6 ..... | Cd-Cu alloy | | |
| plant.........| ............. | ........... | 8.8 +/- 1.1
| | | |
7 ..... | Cadmium | | |
| recovery | | |
| operation - | | |
| Current (19) | | |
| and former | | |
| (26) workers. | ............. | .......... | 7.9 +/- 2.0
| | | |
8 ..... | Cadmium | | |
| recovery | 10.2 +/- 5.3 | 2.2 - 18.8 | ..........
| operation | | |
________|________________|_______________|_____________|___________
TABLE 5. - BLOOD CADMIUM IN WORKERS EXPOSED TO CADMIUM
IN THE WORKPLACE
[Continued]
____________________________________________________________________
| |
| Work | Concentrations of Cadmium in Blood(a)
| environment |__________________________________________
Study | (worker | | |
number | population | Lower 95th | Upper 95th |
| monitored) | percentile of | percentile of | Reference
| | range(c) | range(c) |
| | ( )(f) | ( )(f) |
________|________________|_______________|_______________|__________
| | | |
1 ..... | Ni-Cd battery | | | Lauwerys
| plant and Cd | | | et al.
| production | | | 1976.
| plant: | | |
| | | |
| (Workers | | |
| without | | |
| kidney | | |
| lesions).....| (18) | (25) |
| (Workers with | | |
| kidney | | |
| lesions).....| (32) | (45) |
| | | |
2 ..... | Ni-Cd battery | | | Adamsson
| plant: | | | et al.
| | | | (1979).
| (Smokers) .....| | |
| (Nonsmokers)...| | |
| | | |
3 ..... | Cadmium alloy | | | Sukuri
| plant: | | | et al.
| | | | 1982.
| (High exposure | | |
| group)........| (7.3) | (34) |
| (Low exposure | | |
| group)........| (5.1) | (9.1) |
| | | |
4 ..... | Retrospective | | | Roels
| study of | | | et al.
| wokers with | | | 1982.
| renal | | |
| problems: | | ............. |
| | | |
| (Before | | |
| removal).....| (34) | (46) |
| (After | | |
| removal).....| (4.4) | (24) |
| | | |
5 ..... | Cadmium | | | Ellis
| production | | | et al.
| plant: | | | 1983.
| | | |
| (Workers | | |
| without renal| | |
| dysfunction).| (5.4) | (25) |
| (Workers with | | |
| renal | | |
| dysfunction).| (9.3) | (39) |
| | | |
6 ..... | Cd-Cu alloy | | | Mason
| plant.........| 7.5 | 10 | et al.
| | | | 1988.
7 ..... | Cadmium | | | Thun
| recovery | | | et al.
| operation - | | | 1989.
| Current (19) | | |
| and former | | |
| (26) workers. | 2.5 | 25 |
| | | |
8 ..... | Cadmium | | | Mueller
| recovery | | | et al.
| operation | (1.3) | (19) | 1989.
________|________________|_______________|_______________|__________
CDB levels reported in Table 5 are higher among those showing signs of cadmium-related kidney damage than those showing no such damage. Lauwerys et al. (1976)
report CDB levels among workers with kidney lesions that generally are above the levels reported for workers without kidney lesions. Ellis et al. (1983) report a similar observation comparing workers with and without renal dysfunction, although they found more overlap between the 2 groups than Lauwerys et al.
The data in Table 5 also indicate that CDB levels are higher among those experiencing current occupational exposure than those who have been removed from such exposure. Roels et al. (1982) indicate that CDB levels observed among workers experiencing ongoing exposure in the work place are almost entirely above levels observed among workers removed from such exposure. This finding suggests that CDB levels decrease once cadmium exposure has ceased.
| | A comparison of the data presented in Tables 4 and 5 indicates that CDB levels observed among cadmium-exposed workers is significantly higher than levels observed among the unexposed groups. With the exception of 2 studies presented in Table 5 (1 of which includes former workers in the sample group tested), the lower 95th percentile for CDB levels among exposed workers are greater than 5 m g/l, which is the value of the upper 95th percentile for CDB levels observed among those who are not occupationally exposed. Therefore, a CDB level of 5 m g/l represents a threshold above which significant work place exposure to
cadmium may be occurring. | |
5.1.7 Conclusions and Recommendations for CDB
Based on the above evaluation, the following recommendations are made for a CDB proficiency program.
5.1.7.1 Recommended method
The method of Stoeppler and Brandt (1980) should be adopted for analyzing CDB. This method was selected over other methods for its straightforward sample-preparation procedures, and because limitations of the method were described adequately. It also is the method used by a plurality of laboratories currently participating in the CTQ proficiency program. In a recent CTQ interlaboratory comparison report (CTQ 1991), analysis of the methods used by laboratories to measure CDB indicates that 46% (11 of 24) of the participating laboratories used the Stoeppler and Brandt methodology (HNO 3 deproteinization of blood followed by analysis of the supernatant by GF-AAS). Other CDB methods employed by participating laboratories identified in the CTQ report include dilution of blood (29%), acid digestion (12%) and miscellaneous methods (12%).
Laboratories may adopt alternate methods, but it is the responsibility of the laboratory to demonstrate that the alternate methods meet the data quality objectives defined for the Stoeppler and Brandt method (see Section 5.1.7.2 below).
5.1.7.2 Data quality objectives
Based on the above evaluation, the following data quality objectives (DQOs) should facilitate interpretation of analytical results.
Limit of Detection. 0.5 m g/l should be achievable using the Stoeppler and Brandt method. Stoeppler and Brandt (1980) report a limit of detection equivalent to <0.2 m g/l in whole blood using 2 m l aliquots of deproteinized, diluted blood samples.
Accuracy. Initially, some of the laboratories performing CDB measurements may be expected to satisfy criteria similar to the less severe criteria specified by the CTQ program, i.e., measurements within 2 m g/l or 15% (whichever is greater) of the target value. About 60% of the laboratories enrolled in the CTQ program could meet this criterion on the first proficiency test (Weber 1988).
Currently, approximately 12 laboratories in the CTQ program are achieving an accuracy for CDB analysis within the more severe constraints of + 1 m g/l or 15% (whichever is greater). Later, as laboratories gain experience, they should achieve the level of accuracy exhibited by these 12 laboratories. The experience in the CTQ program has shown that, even without incentives, laboratories benefit from the feedback of the program; after they have analyzed 40-50 control samples from the program, performance improves to thepoint where about 60% of the laboratories can meet the stricter criterion of + 1 m g/l or 15% (Weber 1988). Thus, this stricter target accuracy is a reasonable DQO.
Precision. Although Stoeppler and Brandt (1980) suggest that a coefficient of variation (CV) near 1.3% (for a 10 m g/l concentration) is achievable for within-run reproducibility, it is recognized that other factors affecting within- and between- run comparability will increase the achievable CV. Stoeppler and Brandt (1980) observed CVs that were as high as 30% for low concentrations (0.4 m g/l), and CVs of less than 5% for higher concentrations.
For internal QC samples (see Section 3.3.1), laboratories should to attain an overall precision near 25%. For CDB samples with concentrations less than 2 m g/l, a target precision of 40% is reasonable, while precisions of 20% should be achievable for concentrations greater than 2 m g/l. Although these values are
more strict than values observed in the CTQ interlaboratory program reported by Webber (1988), they are within the achievable limits reported by Stoeppler and Brandt (1980).
5.1.7.3 Quality assurance/quality control
Commercial laboratories providing measurement of CDB should adopt an internal QA/QC program that incorporates the following components: Strict adherence to the selected method, including all calibration requirements; regular incorporation of QC samples during actual runs; a protocol for corrective actions, and documentation of these actions; and, participation in an interlaboratory proficiency program. Note that the nonmandatory QA/QC program presented in Attachment 3 is based on the Stoeppler and Brandt method for CDB analysis. Should an alternate method be adopted, the laboratory should develop a QA/QC program satisfying the provisions of Section 3.3.1.
5.2 Measuring Cadmium in Urine (CDU)
As in the case of CDB measurement, proper determination of CDU requires strict analytical discipline regarding collection and handling of samples. Because cadmium is both ubiquitous in the environment and employed widely in coloring
agents for industrial products that may be used during sample collection, preparation and analysis, care should be exercised to ensure that samples are not contaminated during the sampling procedure.
Methods for CDU determination share many of the same features as those employed for the determination of CDB. Thus, changes and improvements to methods for measuring CDU over the past 40 years parallel those used to monitor CDB. The direction of development has largely been toward the simplification of sample preparation techniques made possible because of improvements in analytic techniques.
5.2.1 Units of CDU Measurement
Procedures adopted for reporting CDU concentrations are not uniform. In fact, the situation for reporting CDU is more complicated than for CDB, where concentrations are normalized against a unit volume of whole blood.
Concentrations of solutes in urine vary with several biological factors (including the time since last voiding and the volume of liquid consumed over the last few hours); as a result, solute concentrations should be normalized against another characteristic of urine that represents changes in solute
concentrations. The 2 most common techniques are either to standardize solute concentrations against the concentration of creatinine, or to standardize solute concentrations against the specific gravity of the urine. Thus, CDU concentrations have been reported in the literature as "uncorrected" concentrations of cadmium per volume of urine (i.e., m g Cd/l urine), "corrected" concentrations of cadmium per volume of urine at a standard specific gravity (i.e., m g Cd/l urine at a specific gravity of 1.020), or "corrected" mass concentration per unit mass of creatinine (i.e., m g Cd/g creatinine). (CDU concentrations [whether uncorrected or corrected for specific gravity, or normalized to creatinine] occasionally are reported in nanomoles [i.e., nmoles] of cadmium per unit mass or volume. In this protocol, these values are converted to m g of cadmium per unit mass or volume using 89 nmoles of cadmium=10 m g.)
While it is agreed generally that urine values of analytes should be normalized for reporting purposes, some debate exists over what correction method should be used. The medical community has long favored normalization based on creatinine concentration, a common urinary constituent. Creatinine is a normal product of tissue catabolism, is excreted at a uniform rate, and the total amount excreted per day is constant on a day-to-day basis (NIOSH 1984b). While this correction method is accepted widely in Europe, and within some
occupational health circles, Kowals (1983) argues that the use of specific gravity (i.e., total solids per unit volume) is more straightforward and practical (than creatinine) in adjusting CDU values for populations that vary by age or gender.
Kowals (1983) found that urinary creatinine (CRTU) is lower in females than males, and also varies with age. Creatinine excretion is highest in younger males (20-30 years old), decreases at middle age (50-60 years), and may rise slightly in later years. Thus, cadmium concentrations may be underestimated for some workers with high CRTU levels.
Within a single void urine collection, urine concentration of any analyte will be affected by recent consumption of large volumes of liquids, and by heavy physical labor in hot environments. The absolute amount of analyte excreted may be identical, but concentrations will vary widely so that urine must be corrected for specific gravity (i.e., to normalize concentrations to the quantity of total solute) using a fixed value (e.g., 1.020 or 1.024). However, since heavy-metal exposure may increase urinary protein excretion, there is a tendency to underestimate cadmium concentrations in samples with high specific gravities when specific-gravity corrections are applied.
Despite some shortcomings, reporting solute concentrations as a function of creatinine concentration is accepted generally; OSHA therefore recommends that CDU levels be reported as the mass of cadmium per unit mass of creatinine ( m g/g CTRU).
Reporting CDU as m g/g CRTU requires an additional analytical process beyond the analysis of cadmium: Samples must be analyzed independently for creatinine so that results may be reported as the ratio of cadmium to creatinine concentrations found in the urine sample. Consequently, the overall quality of the analysis depends on the combined performance by a laboratory on these 2 determinations. The analysis used for CDU determinations is addressed below in terms of m g Cd/l, with analysis of creatinine addressed separately. Techniques for assessing creatinine are discussed in Section 5.4.
Techniques for deriving cadmium as a ratio of CRTU, and the confidence limits for independent measurements of cadmium and CRTU, are provided in Section 3.3.3.
5.2.2 Analytical Techniques Used to Monitor CDU
Analytical techniques used for CDU determinations are similar to those employed
for CDB determinations; these techniques are summarized in Table 3. As with CDB monitoring, the technique most suitable for CDU determinations is atomic absorption spectroscopy (AAS). AAS methods used for CDU determinations typically employ a graphite furnace, with background correction made using either the deuterium-lamp or Zeeman techniques; Section 5.1.1 provides a detailed description of AAS methods.
5.2.3 Methods Developed for CDU Determinations
Princi (1947), Smith et al. (1955), Smith and Kench (1957), and Tsuchiya (1967) used colorimetric procedures similar to those described in the CDB section above to estimate CDU concentrations. In these methods, urine (50 ml) is reduced to dryness by heating in a sand bath and digested (wet ashed) with mineral acids. Cadmium then is complexed with dithiazone, extracted with chloroform and quantified by spectrophotometry. These early studies typically report reagent blank values equivalent to 0.3 m g Cd/l, and CDU concentrations among nonexposed control groups at maximum levels of 10 m g Cd/l -erroneously high values when compared to more recent surveys of cadmium concentrations in the general population.
By the mid-1970s, most analytical procedures for CDU analysis used either wet
ashing (mineral acid) or high temperatures (>400 ° C) to digest the organic matrix of urine, followed by cadmium chelation with APDC or DDTC solutions and extraction with MIBK. The resulting aliquots were analyzed by flame or graphite-furnace AAS (Kjellstrom 1979).
Improvements in control over temperature parameters with electrothermal heating devices used in conjunction with flameless AAS techniques, and optimization of temperature programs for controlling the drying, charring, and atomization processes in sample analyses, led to improved analytical detection of diluted urine samples without the need for sample digestion or ashing. Roels et al. (1978) successfully used a simple sample preparation, dilution of 1.0 ml aliquots of urine with 0.1 N HNO 3, to achieve accurate low-level determinations of CDU.
In the method described by Pruszkowska et al. (1983), which has become the preferred method for CDU analysis, urine samples were diluted at a ratio of 1:5 with water; diammonium hydrogenphosphate in dilute HNO 3 was used as a matrix modifier. The matrix modifier allows for a higher charring temperature without loss of cadmium through volatilization during pre-atomization. This procedure also employs a stabilized temperature platform in a graphite furnace, while nonspecific background absorbtion is corrected using the Zeeman
technique. This method allows for an absolute detection limit of approximately 0.04 m g Cd/l urine.
5.2.4 Sample Collection and Handling
Sample collection procedures for CDU may contribute to variability observed among CDU measurements. Sources of variation attendant to sampling include time-of-day, the interval since ingestion of liquids, and the introduction of external contamination during the collection process. Therefore, to minimize contributionsfrom these variables, strict adherence to a sample-collection protocol is recommended. This a protocol should include provisions for normalizing the conditions under which urine is collected. Every effort also should be made to collect samples during the same time of day.
Collection of urine samples from an industrial work force for biological monitoring purposes usually is performed using "spot" (i.e., single-void) urine with the pH of the sample determined immediately. Logistic and sample-integrity problems arise when efforts are made to collect urine over long periods (e.g., 24 hrs). Unless single-void urines are used, there are numerous opportunities for measurement error because of poor control over sample collection, storage and environmental contamination.
To minimize the interval during which sample urine resides in the bladder, the following adaption to the "spot" collection procedure is recommended: The bladder should first be emptied, and then a large glass of water should be consumed; the sample may be collected within an hour after the water is consumed.
5.2.5 Best Achievable Performance
Performance using a particular method for CDU determinations is assumed to be equivalent to the performance reported by the research laboratories in which the method was developed. Pruszkowska et al. (1983) report a detection limit of 0.04 m g/l CDU, with a CV of <4not=tween0-5 m g/l.The CDC reports a minimum CDU detection limit of 0.07 m g/l using a modified method based on Pruszkowska et al. (1983). No CV is stated in this protocol; the protocol contains only rejection criteria for internal QC parameters used during accuracy determinations with known standards (Attachment 8 of exhibit 106 of OSHA docket H057A). Stoeppler and Brandt (1980) report a CDU detection limit of 0.2 m g/l for their methodology.
5.2.6 General Method Performance
For any particular method, the expected initial performance from commercial laboratories may be somewhat lower than that reported by the research laboratory in which the method was developed. With participation in appropriate proficiency programs, and use of a proper in-house QA/QC program incorporating provisions for regular corrective actions, the performance of commercial laboratories may be expected to improve and approach that reported by a research laboratories. The results reported for existing proficiency programs serve to specify the initial level of performance that likely can be expected from commercial laboratories offering analysis using a particular method.
Weber (1988) reports on the results of the CTQ proficiency program, which includes CDU results for laboratories participating in the program. Results indicate that after receiving 60 samples (i.e., after participating in the program for approximately 3 years), approximately 80% of the participating laboratories report CDU results ranging between + 2 m g/l or 15% of the consensus mean, whichever is greater. On any single sample of the last 15 samples, the proportion of laboratories falling within the specified range is between 75 and 95%, except for a single test for which only 60% of the laboratories reported acceptable results. For each of the last 15 samples, approximately 60% of the laboratories reported results within + 1 m g or 15% of
the mean, whichever is greater. The range of concentrations included in this set of samples was not reported.
Another report from the CTQ (1991) summarizes preliminary CDU results from their 1991 interlaboratory program. According to the report, for 3 CDU samples with values of 9.0, 16.8, 31.5 m g/l, acceptable results (target + 2 m g/l) were achieved by only 44-52% of the 34 laboratories participating in the CDU program. The overall CVs for these 3 CDU samples among the 34 participating laboratories were 31%, 25%, and 49%, respectively. The reason for this poor performance has not been determined.
| | A more recent report from the CTQ (Weber, private communication) indicates that 36% of the laboratories in the program have been able to achieve the target of + 1 m g/l or 15% for more than 75% of the samples analyzed over the last 5 years, while 45% of participating laboratories achieved a target of + 2 m g/l or 15% for more than 75% of the samples analyzed over the same period. | |
| | Note that results reported in the interlaboratory programs are in terms of m g Cd/l of urine, unadjusted for creatinine. The performance indicated, therefore, is a measure of the performance of the cadmium portion of the analyses, and does not include variationthat may be introduced during the analysis of CRTU. | |
5.2.7 Observed CDU Concentrations
Prior to the onset of renal dysfunction, CDU concentrations provide a general indication of the exposure history (i.e., body burden) (see Section 4.3). Once renal dysfunction occurs, CDU levels appear to increase and are no longer indicative solely of cadmium body burden (Friberg and Elinder 1988).
5.2.7.1 Range of CDU concentrations observed among unexposed samples
Surveys of CDU concentrations in the general population were first reported from cooperative studies among industrial countries (i.e., Japan, U.S. and Sweden) conducted in the mid-1970s. In summarizing these data, Kjellstrom (1979) reported that CDU concentrations among Dallas, Texas men (age range: 9- 59 years; smokers and nonsmokers) varied from 0.11-1.12 m g/l (uncorrected for creatinine or specific gravity). These CDU concentrations are intermediate between population values found in Sweden (range: 0.11-0.80 m g/l) and Japan (range: 0.14-2.32 m g/l).
Kowal and Zirkes (1983) reported CDU concentrations for almost 1,000 samples collected during 1978-79 from the general U.S. adult population (i.e., nine
states; both genders; ages 20-74 years). They report that CDU concentrations are lognormally distributed; low levels predominated, but a small proportion of the population exhibited high levels. These investigators transformed the CDU concentrations values, and reported the same data 3 different ways: m g/l urine (unadjusted), m g/l (specific gravity adjusted to 1.020), and m g/g CRTU. These data are summarized in Tables 6 and 7.
Based on further statistical examination of these data, including the lifestyle characteristics of this group, Kowal (1988) suggested increased cadmium absorption (i.e., body burden) was correlated with low dietary intakes of calcium and iron, as well as cigarette smoking.
CDU levels presented in Table 6 are adjusted for age and gender. Results suggest that CDU levels may be slightly different among men and women (i.e., higher among men when values are unadjusted, but lower among men when the values are adjusted, for specific gravity or CRTU). Mean differences among men and women are small compared to the standard deviations, and therefore may not be significant. Levels of CDU also appear increase with age. The data in Table 6 suggest as well that reporting CDU levels adjusted for specific gravity or as a function of CRTU results in reduced variability.
Table 6 - URINE CADMIUM CONCENTRATIONS IN THE U.S. ADULT POPULATION:
NORMAL AND CONCENTRATION-ADJUSTED VALUES BY AGE AND SEX
_______________________________________________________________________
|
| Geometric means (and geometric standard
| deviations)
|________________________________________________
| | |
| Unadjusted | SG-adjusted(2) | Creatine-
| (ug/l) |(ug/l at 1.020) | adjusted (ug/g)
______________________|_____________|________________|_________________
| | |
SEX: | | |
Male (n=484) ...... | 0.55 (2.9) | 0.73 (2.6) | 0.55 (2.7)
Female (n=498) .... | 0.49 (3.0) | 0.86 (2.7) | 0.78 (2.7)
Age: | | |
20-29 (n=222) ..... | 0.32 (3.0) | 0.43 (2.7) | 0.32 (2.7)
30-39 (n=141) ..... | 0.46 (3.2) | 0.70 (2.8) | 0.54 (2.7)
40-49 (n=142) ..... | 0.50 (3.0) | 0.81 (2.6) | 0.70 (2.7)
50-59 (n=117) ..... | 0.61 (2.9) | 0.99 (2.4) | 0.90 (2.3)
60-69 (n=272) ..... | 0.76 (2.6) | 1.16 (2.3) | 1.03 (2.3)
______________________|_____________|________________|_________________
Table 7 - URINE CADMIUM CONCENTRATIONS IN THE U.S. ADULT POPULATION:
CUMULATIVE FREQUENCY DISTRIBUTION OF URINARY CADMIUM
_________________________________________________________________________
| | |
Range of | Unadjusted | SG-adjusted | Creatine-adjusted
Concentrations | (ug/l) | (ug/l at 1.020) | (ug/g)
| percent | percent | percent
______________________|____________|_________________|__________________
| | |
<0.5 ................ | 43.9 | 28.0 | 35.8
0.6 - 1.0 ........... | 71.7 | 56.4 | 65.6
1.1 - 1.5 ........... | 84.4 | 74.9 | 81.4
1.6 - 2.0 ........... | 91.3 | 84.7 | 88.9
2.1 - 3.0 ........... | 97.3 | 94.4 | 95.8
3.1 - 4.0 ........... | 98.8 | 97.4 | 97.2
4.1 - 5.0 ........... | 99.4 | 98.2 | 97.9
5.1 - 10.0 .......... | 99.6 | 99.4 | 99.3
10.0 - 20.0 ......... | 99.8 | 99.6 | 99.6
______________________|____________|_________________|_________________
The data in the Table 6 indicate the geometric mean of CDU levels observed among the general population is 0.52 m g Cd/l urine (unadjusted), with a geometric standard deviation of 3.0. Normalized for creatinine, the geometric mean for the population is 0.66 m g/g CRTU, with a geometric standard deviation of 2.7. Table 7 provides the distributions of CDU concentrations for the general population studied by Kowal and Zirkes. The data in this table indicate that 95% of the CDU levels observed among those not occupationally exposed to cadmium are below 3 m g/g CRTU.
5.2.7.2 Range of CDU concentrations observed among exposed workers
Table 8 is a summary of results from available studies of CDU concentrations observed among cadmium-exposed workers. In this table, arithmetic and/or geometric means and standard deviations are provided if reported in these studies. The absolute range for the data in each study, or the 95% confidence interval around the mean of each study, also are provided when reported. The lower and upper 95th percentile of the distribution are presented for each study in which a mean and corresponding standard deviation were reported. Table
8 also provides estimates of the years of exposure, and the levels of exposure, to cadmium in the work place if reported in these studies. Concentrations reported in this table are in m g/g CRTU, unless otherwise stated.
Data in Table 8 from Lauwerys et al. (1976) and Ellis et al. (1983) indicate that CDU concentrations are higher among those exhibiting kidney lesions or dysfunction than among those lacking these symptoms. Data from the study by Roels et al. (1982) indicate that CDU levels decrease among workers removed from occupational exposure to cadmium in comparison to workers experiencing ongoing exposure. In both cases, however, the distinction between the 2 groups is not as clear as with CDB; there is more overlap in CDU levels observed among each of the paired populations than is true for corresponding CDB levels. As with CDB levels, the data in Table 8 suggest increased CDU concentrations among workers who experienced increased overall exposure.
Table 8 - URINE CADMIUM CONCENTRATIONS IN WORKERS EXPOSED
TO CADMIUM IN THE WORKPLACE
______________________________________________________________
| | | | |
| Work | | | Mean |
| environment | | Employment | concentration |
Study | (worker | Number | in | of cadmium |
number | population | in | years | in air |
| monitored) | study | (mean) | (ug/m(3)) |
| | (n) | | |
________|________________|________|____________|________________|
| | | | |
1 ..... | Ni-Cd battery | ...... | 3-40 | < than = to 90 |
| plant and Cd | | | |
| production | | | |
| plant: | | | |
| (Workers | | | |
| without | | | |
| kidney | | | |
| lesions).....| 96 | .......... | .............. |
| (Workers with | | | |
| kidney | | | |
| lesions).....| 25 | .......... | .............. |
| | | | |
2 ..... | Ni-Cd battery | | | |
| plant.........| ...... | .......... | .............. |
| | | | |
| (Smokers) .....| 7 | (5) | 10.1 |
| (Nonsmokers)...| 8 | (9) | 7.0 |
| | | | |
3 ..... | Cadmium salts | 148 | (15.4) | .............. |
| production | | | |
| facility. | | | |
| | | | |
| | | | |
4 ..... | Retrospective | 19 | 15-41 | .............. |
| study of | | | |
| wokers with | | | |
| renal | | | |
| problems: | | | |
| | | | |
| (Before | | | |
| removal).....| ...... | (27.2) | .............. |
| (After | | | |
| removal).....| ...... | (4.2)(g) | .............. |
| | | | |
5 ..... | Cadmium | ...... | .......... | .............. |
| production | | | |
| plant: | | | |
| | | | |
| (Workers | | | |
| without renal| | | |
| dysfunction).| 33 | 1-34 | .............. |
| (Workers with | | | |
| renal | 18 | 10-34 | .............. |
| dysfunction).| | | |
| | | | |
6 ..... | Cd-Cu alloy | | | |
| plant.........| 75 | Up to 39 | Note h |
| | | | |
| | | | |
7 ..... | Cadmium | | | |
| recovery | | | |
| operation. | 45 | (19) | 87 |
| | | | |
8 ..... | Pigment | | | |
| manufacturing | | | |
| plant. | 29 | (12.8) | 0.18-3.0 |
| | | | |
9 ..... | Pigment | | | |
| manufacturing | | | |
| plant. | 26 | (12.1) |< than = to 3.0 |
| | | | |
________|________________|________|____________|________________|
TABLE 8. - URINE CADMIUM CONCENTRATIONS IN WORKERS EXPOSED
TO CADMIUM IN THE WORKPLACE
[Continued]
____________________________________________________________________
| |
| Work | Concentrations of Cadmium in Urine(a)
| environment |__________________________________________
Study | (worker | | |
number | population | Arithmetic | Absolute | Geometric
| monitored) | mean | range or | mean
| | (+/- S.D.)(b) |(95% C.I.)(c)| (GSD)(d)
________|________________|_______________|_____________|____________
| | | |
1 ..... | Ni-Cd battery | ............. | ........... | .........
| plant and Cd | | |
| production | | |
| plant: | | |
| | | |
| (Workers | | |
| without | | |
| kidney | | |
| lesions).....| 16.3 +/- 16.7 | ........... | .........
| (Workers with | | |
| kidney | | |
| lesions).....| 48.2 +/- 42.6 | ........... | .........
| | | |
2 ..... | Ni-Cd battery | ............. | ........... | .........
| plant.........| | |
| | | |
| (Smokers) .....| 5.5 | 1.0 - 14.7 | .........
| (Nonsmokers)...| 3.6 | 0 .5 - 9.3 | .........
| | | |
3 ..... | Cadmium salts | 15.8 | 2 - 150 | .........
| production | | |
| facility. | | |
| | | |
4 ..... | Retrospective | ............. | ........... | .........
| study of | | |
| wokers with | | |
| renal | | |
| problems: | | |
| | | |
| (Before | | |
| removal).....| 39.4 +/- 28.1 | 10.8 - 117 | .........
| (After | | |
| removal).....| 16.4 +/- 9.0 | 80 - 42.3 | .........
| | | |
5 ..... | Cadmium | ............. | ........... | .........
| production | | |
| plant: | | |
| | | |
| (Workers | | |
| without renal| | |
| dysfunction).| 9.4 +/- 6.9 | 2 - 27 | .........
| (Workers with | | |
| renal | | |
| dysfunction).| 22.8 +/- 12.7 | 8 - 55 | .........
| | | |
6 ..... | Cd-Cu alloy | | |
| plant.........| 6.9 +/- 9.4 | ........... | .........
| | | |
7 ..... | Cadmium | | |
| recovery | | |
| operation. | 9.3 +/- 6.9 | ........... | .........
| | | |
8 ..... | Pigment | | |
| manufacturing | | |
| plant. | ............. | 0.2 - 9.5 | 1.1
| | | |
9 ..... | Pigment | | |
| manufacturing | | |
| plant. | ............. | ........... | 1.25 +/-
| | | | 2.45
________|________________|_______________|_____________|___________
TABLE 8. - URINE CADMIUM CONCENTRATIONS IN WORKERS EXPOSED
TO CADMIUM IN THE WORKPLACE
[Continued]
____________________________________________________________________
| |
| Work | Concentrations of Cadmium in Urine(a)
| environment |__________________________________________
Study | (worker | | |
number | population | Lower 95th | Upper 95th |
| monitored) | percentile of | percentile of | Reference
| | range(c) | range(c) |
| | ( )(f) | ( )(f) |
________|________________|_______________|_______________|__________
| | | |
1 ..... | Ni-Cd battery | ............. | ............. | Lauwerys
| plant and Cd | | | et al.
| production | | | 1976.
| plant: | | |
| | | |
| (Workers | | |
| without | | |
| kidney | | |
| lesions).....| (0) | (44) |
| (Workers with | | |
| kidney | | |
| lesions).....| (0) | (120) |
| | | |
2 ..... | Ni-Cd battery | ............. | ............. | Adamsson
| plant.........| | | et al.
| | | | (1979).
| (Smokers) .....| ............. | ............. |
| (Nonsmokers)...| ............. | ............. |
| | | |
3 ..... | Cadmium salts | ............. | ............. | Butchet
| production | | | et al.
| facility. | | | 1980.
| | | |
4 ..... | Retrospective | ............. | ............. | Roels
| study of | | | et al.
| wokers with | | | 1982.
| renal | | |
| problems: | | |
| | | |
| (Before | | |
| removal).....| (0) | (88) |
| (After | | |
| removal).....| (1.0) | (32) |
| | | |
5 ..... | Cadmium | ............. | ............. | Ellis
| production | | | et al.
| plant: | | | 1983.
| | | |
| (Workers | | |
| without renal| | |
| dysfunction).| (0) | (21) |
| (Workers with | | |
| renal | | |
| dysfunction).| (1) | (45) |
| | | |
6 ..... | Cd-Cu alloy | (0) | (23) | Mason
| plant.........| | | et al.
| | | | 1988.
| | | |
7 ..... | Cadmium | (0) | (21) | Thun
| recovery | | | et al.
| operation. | | | 1989.
| | | |
8 ..... | Pigment | ............. | ............. | Mueller
| manufacturing | | | et al.
| plant. | | | 1989.
| | | |
9 ..... | Pigment | 0.3 | 6 | Kawada
| manufacturing | | | et al.
| plant. | | | 1990.
| | | |
________|________________|_______________|_______________|__________
Although a few occupationally-exposed workers in the studies presented in Table 8 exhibit CDU levels below 3 m g/g CRTU, most of those workers exposed to cadmium levels in excess of the PEL defined in the final cadmium rule exhibit
CDU levels above 3 m g/g CRTU; this level represents the upper 95th percentile of the CDU distribution observed among those who are not occupationally exposed to cadmium (Table 7).
The mean CDU levels reported in Table 8 among occupationally-exposed groups studied (except 2) exceed 3 m g/g CRTU. Correspondingly, the level of exposure reported in these studies (with 1 exception) are significantly higher than what workers will experience under the final cadmium rule. The 2 exceptions are from the studies by Mueller et al. (1989) and Kawada et al. (1990); these studies indicate that workers exposed to cadmium during pigment manufacture do not exhibit CDU levels as high as those levels observed among workers exposed to cadmium in other occupations. Exposure levels, however, were lower in the pigment manufacturing plants studied. Significantly, workers removed from occupational cadmium exposure for an average of 4 years still exhibited CDU levels in excess of 3 m g/g CRTU (Roels et al. 1982). In the single-exception study with a reported level of cadmium exposure lower than levels proposed in the final rule (i.e., the study of a pigment manufacturing plant by Kawada et al. 1990), most of the workers exhibited CDU levels less than 3 m g/g CRTU (i.e., the mean value was only 1.3 m g/g CRTU). CDU levels among workers with such limited cadmium exposure are expected to be significantly lower than levels reported on Table 8.
Based on the above data, a CDU level of 3 m g/g CRTU appear to represent a threshold above which significant work place exposure to cadmium occurs over the work span of those being monitored. Note that this threshold is not as distinct as the corresponding threshold described for CDB. In general, the variability associated with CDU measurements among exposed workers appears to be higher than the variability associated with CDB measurements among similar workers.
5.2.8 Conclusions and Recommendations for CDU
The above evaluation supports the following recommendations for a CDU proficiency program. These recommendations address only sampling and analysis procedures for CDU determinations specifically, which are to be reported as an unadjusted m /g Cd/l urine. Normalizing this result to creatinine requires a second analysis for CRTU so that the ratio of the 2 measurements can be obtained. Creatinine analysis is addressed in Section 5.4. Formal procedures for combining the 2 measurements to derive a value and a confidence limit for CDU in m g/g CRTU are provided in Section 3.3.3.
5.2.8.1 Recommended method
The method of Pruszkowska et al. (1983) should be adopted for CDU analysis. This method is recommended because it is simple, straightforward and reliable (i.e., small variations in experimental conditions do not affect the analytical results).
| | A synopsis of the methods used by laboratories to determine CDU under the interlaboratory program administered by the CTQ (1991) indicates that more than 78% (24 of 31) of the participating laboratories use a dilution method to prepare urine samples for CDU analysis. Laboratories may adopt alternate methods, but it is the responsibility of the laboratory to demonstrate that the alternate methods provide results of comparable quality to the Pruszkowska method. | |
5.2.8.2 Data quality objectives
The following data quality objectives should facilitate interpretation of analytical results, and are achievable based on the above evaluation.
Limit of Detection. A level of 0.5 m g/l (i.e., corresponding to a detection limit of 0.5 m g/g CRTU, assuming 1 g CRT/l urine) should be achievable.
Pruszkowska et al. (1983) achieved a limit of detection of 0.04 m g/l for CDU based on the slope the the curve for their working standards (0.35 pg Cd/0.0044, A signal=1% absorbance using GF-AAS).
The CDC reports a minimum detection limit for CDU of 0.07 m g/l using a modified Pruszkowska method. This limit of detection was defined as 3 times the standard deviation calculated from 10 repeated measurements of a "low level" CDU test sample (Attachment 8 of exhibit 106 of OSHA docket H057A).
Stoeppler and Brandt (1980) report a limit of detection for CDU of 0.2 m g/l using an aqueous dilution (1:2) of the urine samples.
Accuracy. A recent report from the CTQ (Weber, private communication) indicates that 36% of the laboratories in the program achieve the target of + 1 m g/l or 15% for more than 75% of the samples analyzed over the last 5 years, while 45% of participating laboratories achieve a target of + 2 m g/l or 15% for more than 75% of the samples analyzed over the same period. With time and a strong incentive for improvement, it is expected that the proportion of laboratories successfully achieving the stricter level of accuracy should increase. It should be noted, however, these indices of performance do not include variations resulting from the ancillary measurement of CRTU (which is
recommended for the proper recording of results). The low cadmium levels expected to be measured indicate that the analysis of creatinine will contribute relatively little to the overall variability observed among creatinine-normalized CDU levels (see Section 5.4). The initial target value for reporting CDU under this program, therefore, is set at + 1 m g/g CRTU or 15% (whichever is greater).
Precision. For internal QC samples (which are recommended as part of an internal QA/QC program, Section 3.3.1), laboratories should attain an overall precision of 25%. For CDB samples with concentrations less than 2 m g/l, a target precision of 40% is, while precisions of 20% should be achievable for CDU concentrations greater than 2 m g/l. Although these values are more stringent than those observed in the CTQ interlaboratory program reported by Webber (1988), they are well within limits expected to be achievable for the method as reported by Stoeppler and Brandt (1980).
5.2.8.3 Quality assurance/quality control
Commercial laboratories providing CDU determinations should adopt an internal QA/QC program that incorporates the following components: Strict adherence to the selected method, including calibration requirements; regular incorporation
of QC samples during actual runs; a protocol for corrective actions, and documentation of such actions; and, participation in an interlaboratory proficiency program. Note that the nonmandatory program presented in Attachment 1 as an example of an acceptable QA/QC program, is based on using the Pruszkowska method for CDU analysis. Should an alternate method be adopted by a laboratory, the laboratory should develop a QA/QC program equivalent to the nonmandatory program, and which satisfies the provisions of Section 3.3.1.
5.3 Monitoring b 2-Microglobulin in Urine (B2MU)
As indicated in Section 4.3, B2MU appears to be the best of several small proteins that may be monitored as early indicators of cadmium-induced renal damage. Several analytic techniques are available for measuring B2M.
5.3.1 Units of B2MU Measurement
Procedures adopted for reporting B2MU levels are not uniform. In these guidelines, OSHA recommends that B2MU levels be reported as m g/g CRTU, similar to reporting CDU concentrations. Reporting B2MU normalized to the concentration of CRTU requires an additional analytical process beyond the analysis of B2M: Independent analysisfor creatinine so that results may be reported as a ratio
of the B2M and creatinine concentrations found in the urine sample. Consequently, the overall quality of the analysis depends on the combined performance on these 2 analyses. The analysis used for B2MU determinations is described in terms of m g B2M/l urine, with analysis of creatinine addressed separately. Techniques used to measure creatinine are provided in Section 5.4. Note that Section 3.3.3 provides techniques for deriving the value of B2M as function of CRTU, and the confidence limits for independent measurements of B2M and CRTU.
5.3.2 Analytical Techniques Used to Monitor B2MU
One of the earliest tests used to measure B2MU was the radial immunodiffusion technique. This technique is a simple and specific method for identification and quantitation of a number of proteins found in human serum and other body fluids when the protein is not readily differentiated by standard electrophoretic procedures. A quantitative relationship exists between the concentration of a protein deposited in a well that is cut into a thin agarose layer containing the corresponding monospecific antiserum, and the distance that the resultant complex diffuses. The wells are filled with an unknown serum and the standard (or control), and incubated in a moist environment at room temperature. After the optimal point of diffusion has been reached, the
diameters of the resulting precipition rings are measured. The diameter of a ring is related to the concentration of the constituent substance. For B2MU determinations required in the medical monitoring program, this method requires a process that may be insufficient to concentrate the protein to levels that are required for detection.
Radioimmunoassay (RIA) techniques are used widely in immunologic assays to measure the concentration of antigen or antibody in body-fluid samples. RIA procedures are based on competitive-binding techniques. If antigen concentration is being measured, the principle underlying the procedure is that radioactive-labeled antigen competes with the sample's unlabeled antigen for binding sites on a known amount of immobile antibody. When these 3 components are present in the system, an equilibrium exists. This equilibrium is followed by a separation of the free and bound forms of the antigen. Either free or bound radioactive- labeled antigen can be assessed to determine the amount of antigen in the sample. The analysis is performed by measuring the level of radiation emitted either by the bound complex following removal of the solution containing the free antigen, or by the isolated solution containing the residual-free antigen. The main advantage of the RIA method is the extreme sensitivity of detection for emitted radiation and the corresponding ability to detect trace amounts of antigen. Additionally, large numbers of tests can be
performed rapidly.
The enzyme-linked immunosorbent assay (ELISA) techniques are similar to RIA techniques except that nonradioactive labels are employed. This technique is safe, specific and rapid, and is nearly as sensitive as RIA techniques. An enzyme-labeled antigen is used in the immunologic assay; the labeled antigen detects the presence and quantity of unlabeled antigen in the sample. In a representative ELISA test, a plastic plate is coated with antibody (e.g., antibody to B2M). The antibody reacts with antigen (B2M) in the urine and forms an antigen-antibody complex on the plate. A second anti-B2M antibody (i.e., labeled with an enzyme) is added to the mixture and forms an antibody-antigen-antibody complex. Enzyme activity is measured spectrophotometrically after the addition of a specific chromogenic substrate which is activated by the bound enzyme. The results of a typical test are calculated by comparing the spectrophotometric reading of a serum sample to that of a control or reference serum. In general, these procedures are faster and require less laboratory work than other methods.
In a fluorescent ELISA technique (such as the one employed in the Pharmacia Delphia test for B2M), the labeled enzyme is bound to a strong fluorescent dye. In the Pharmacia Delphia test, an antigen bound to a fluorescent dye competes
with unlabeled antigen in the sample for a predetermined amount of specific, immobile antibody. Once equilibrium is reached, the immobile phase is removed from the labeled antigen in the sample solution and washed; an enhancement solution then is added that liberates the fluorescent dye from the bound antigen-antibody complex. The enhancement solution also contains a chelate that complexes with the fluorescent dye in solution; this complex increases the fluorescent properties of the dye so that it is easier to detect.
To determine the quantity of B2M in a sample using the Pharmacia Delphia test, the intensity of the fluorescence of the enhancement solution is measured. This intensity is proportional to the concentration of labeled antigen that bound to the immobile antibody phase during the initial competition with unlabeled antigen from the sample. Consequently, the intensity of the fluorescence is an inverse function of the concentration of antigen (B2M) in the original sample. The relationship between the fluorescence level and the B2M concentration in the sample is determined using a series of graded standards, and extrapolating these standards to find the concentration of the unknown sample.
5.3.3 Methods Developed for B2MU Determinations
B2MU usually is measured by radioimmunoassay (RIA) or enzyme-linked
immunosorbent assay (ELISA); however, other methods (including gel electrophoresis, radial immunodiffusion, and nephelometric assays) also have been described (Schardun and van Epps 1987). RIA and ELISA methods are preferred because they are sensitive at concentrations as low as micrograms per liter, require no concentration processes, are highly reliable and use only a small sample volume.
Based on a survey of the literature, the ELISA technique is recommended for monitoring B2MU. While RIAs provide greater sensitivity (typically about 1 << mu>>g/l, Evrin et al. 1971), they depend on the use of radioisotopes; use of radioisotopes requires adherence to rules and regulations established by the Atomic Energy Commission, and necessitates an expensive radioactivity counter for testing. Radioisotopes also have a relatively short half-life, which corresponds to a reduced shelf life, thereby increasing the cost and complexity of testing. In contrast, ELISA testing can be performed on routine laboratory spectrophotometers, do not necessitate adherence to additional rules and regulations governing the handling of radioactive substances, and the test kits have long shelf lives. Further, the range of sensitivity commonly achieved by the recommended ELISA test (i.e., the Pharmacia Delphia test) is approximately 100 <<mu>>g/l (Pharmacia 1990), which is sufficient for monitoring B2MU levels resulting from cadmium exposure. Based on the studies listed in Table 7
(Section 5.3.7), the average range of B2M concentrations among the general, nonexposed population falls between 60 and 300 m g/g CRTU. The upper 95th percentile of distributions, derived from studies in Table 9 which reported standard deviations, range between 180 and 1,140 m g/g CRTU. Also, the Pharmacia Delphia test currently is the most widely used test for assessing B2MU.
5.3.4 Sample Collection and Handling
As with CDB or CDU, sample collection procedures are addressed primarily to identify ways to minimize the degree of variability introduced by sample collection during medical monitoring. It is unclear the extent to which sample collection contributes to B2MU variability. Sources of variation include time-of-day effects, the interval since consuming liquids and the quantity of liquids consumed, and the introduction of external contamination during the collection process. A special problem unique to B2M sampling is the sensitivity of this protein to degradation under acid conditions commonly found in the bladder. To minimize this problem, strict adherence to a sampling protocol is recommended. The protocol should include provisions for normalizing the conditions under which the urine is collected. Clearly, it is important to minimize the interval urine spends in the bladder. It also is recommended that
every effort be made to collect samples during the same time of day.
Collection of urine samples for biological monitoring usually is performed using "spot" (i.e., single-void) urine. Logistics and sample integrity become problems when efforts are made to collect urine over extended periods (e.g., 24 hrs). Unless single-void urines are used, numerous opportunities exist for measurement error because of poor control over sample collection, storage and environmental contamination.
To minimize the interval that sample urine resides in the bladder, the following adaption to the "spot" collection procedure is recommended: The bladder should be emptied and then a large glass of water should be consumed; the sample then should be collected within an hour after the the water is consumed.
5.3.5 Best Achievable Performance
The best achievable performance is assumed to be equivalent to the performance reported by the manufacturers of the Pharmacia Delphia test kits (Pharmacia 1990). According to the insert that comes with these kits, QC results should be within + 2 SDs of the mean for each control sample tested; a CV of less than
or equal to 5.2% should be maintained. The total CV reported for test kits is less than or equal to 7.2%.
5.3.6 General Method Performance
Unlike analyses for CDB and CDU, the Pharmacia Delphia test is standardized in a commercial kit that controls for many sources of variation. In the absence of data to the contrary, it is assumed that the achievable performance reported by the manufacturer of this test kit will serve as an achievable performance objective. The CTQ proficiency testing program for B2MU analysis is expected to use the performance parameters defined by the test kit manufacturer as the basis of the B2MU proficiency testing program.
Note that results reported for the test kit are expressed in terms of m g B2M/l of urine, and have not been adjusted for creatinine. The indicated performance, therefore, is a measure of the performance of the B2M portion of the analyses only, and does not include variation that may have been introduced during the analysis of creatinine.
5.3.7 Observed B2MU Concentrations
As indicated in Section 4.3, the concentration of B2MU may serve as an early indicator of the onset of kidney damage associated with cadmium exposure.
5.3.7.1 Range of B2MU concentrations among unexposed samples
Most of the studies listed in Table 9 report B2MU levels for those who were not occupationally exposed to cadmium. Studies noted in the second column of this table (which contain the footnote "d") reported B2MU concentrations among cadmium-exposed workers who, nonetheless, showed no signs of proteinuria. These latter studies are included in this table because, as indicated in Section 4.3, monitoring B2MU is intended to provide advanced warning of the onset of kidney dysfunction associated with cadmium exposure, rather than to distinguish relative exposure. This table, therefore, indicates the range of B2MU levels observed among those who had no symptoms of renal dysfunction (including workers with none of these symptoms).
Table 9 - B-2-MICROGLOBULIN CONCENTRATIONS OBSERVED IN URINE
AMONG THOSE NOT OCCUPATIONALLY EXPOSED TO CADMIUM
____________________________________________________________________
| | | | | |
Study| No. in | Geometric |Geomet- |Lower 95th|Upper 95th|
No. | study | mean | ric |percentile|percentile|Reference
| | |standard|of distri-|of distri-|
| | |devia- |bution(a) |bution(a) |
| | | tion | | |
_____|________|____________|________|__________|__________|_________
| | | | | |
1....|133 m(b)| 115 ug/g(c)| 4.03...| 12.......| 1,140 | ishizaki
| | | | | ug/g(c)..| et al.
| | | | | | 1989.
2....|161 f(b)| 146 ug/g(c)| 3.11...| 23.......| 940 | ishizaki
| | | | | ug/g(c)..| et al.
| | | | | | 1989.
3....| 10.....| 84 ug/g....|........|..........|..........| Ellis
| | | | | | et al.
| | | | | | 1983.
4....| 203....| 76 ug/l....|........|..........|..........| Stewart
| | | | | | and
| | | | | | Hughes
| | | | | | 1981.
5....| 9......| 103 ug/g...|........|..........|..........| Chia
| | | | | | et al.
| | | | | | 1989.
6....| 47(d)..| 86 ug/L....| 1.9....| 30 ug/l..| 250 ug/L.| Kjell-
| | | | | | strom
| | | | | | et al.
| | | | | | 1977.
7....|1,000(e)| 68.1 ug/gr | 3.1 |< 10 u/gr | 320 ug/gr| Kowal
| | Cr(f)....| m & f.| Cr(h)..| Cr (h)..| 1983.
8....| 87.....| 71 ug/g(i).|........| 7(h).....| 200(h)...| Buchet
| | | | | | et al.
| | | | | | 1980.
9....| 10.....|0.073 mg/24h|........|..........|..........| Evrin
| | | | | | et al.
| | | | | | 1971.
10...| 59.....| 156 ug/g...| 1.1(j).| 130......| 180......| Mason
| | | | | | et al.
| | | | | | 1988.
11...| 8......| 118 ug/g...|........|..........|..........| Iwao
| | | | | | et al.
| | | | | | 1980.
12...| 34.....| 79 ug/g....|........|..........|..........| Wibowo
| | | | | | et al.
| | | | | | 1982.
13...| 41 m...|............|........|..........| 400 ug/gr| Falck
| | | | | Cr(k) | et al.
| | | | | | 1983.
14...| 35(n)..| 67.........|........|..........|..........| Roels
| | | | | | et al.
| | | | | | 1991.
15...| 31(d)..| 63.........|........|..........|..........| Roels
| | | | | | et al.
| | | | | | 1991.
16...| 36(d)..| 77(i)......|........|..........|..........| Miksche
| | | | | | et al.
| | | | | | 1981.
17...| 18(n)..| 130........|........|..........|..........| Kawada
| | | | | | et al.
| | | | | | 1989.
18...| 32(p)..| 122........|........|..........|..........| Kawada
| | | | | | et al.
| | | | | | 1989.
19...| 18(d)..| 295........| 1.4....| 170......| 510......| Thun
| | | | | | et al.
| | | | | | 1989.
_____|________|____________|________|__________|__________|________
To the extent possible, the studies listed in Table 9 provide geometric means and geometric standard deviations for measurements among the groups defined in
each study. For studies reporting a geometric standard deviation along with a mean, the lower and upper 95th percentile for these distributions were derived and reported in the table.
Table 10 - B-2-MICROGLOBULIN CONCENTRATIONS OBSERVED IN
URINE AMONG OCCUPATIONALLY-EXPOSED WORKERS
__________________________________________________________________
| | |
| | Concentration of B(2)-microglobulin |
| | in urine |
| |__________________________________________|
| | | | | |
Study | | Geometric | Geom. | L 95% of | U 95% of | Reference
number| N | mean | Std. | range(b) | range(b) |
| | (ug/g)(a) | Dev. | | |
______|_____|___________|_______|__________|___________|___________
| | | | | |
1.....|1,424| 160 | 6.19 | 8.1 | 3,300 | Ishizaki
| | | | | | et al.
| | | | | | 1989.
2.....|1,754| 260 | 6.50 | 12 | 5,600 | Ishizaki
| | | | | | et al.
| | | | | | 1989.
3.....| 33| 210 |.......|..........|...........| Ellis
| | | | | | et al.
| | | | | | 1983.
4.....| 65| 210 |.......|..........|...........| Chia
| | | | | | et al.
| | | | | | 1989.
5.....|(c)44| 5,700 | 6.49 | (d)300 | (d)98,000 | Kjellstrom
| | | | | | et al.
| | | | | | 1977.
6.....| 148| (e)180 |.......| (f)110 | (f)280 | Buchet
| | | | | | et al.
| | | | | | 1980.
7.....| 37| 160 | 3.90 | 17 | 1,500 | Kenzaburo
| | | | | | et al.
| | | | | | 1979.
8.....|(c)45| 3,300 | 8.70 | (d)310 | (d)89,000 | Mason
| | | | | | et al.
| | | | | | 1988.
9.....|(c)10| 6,100 | 5.99 | (f)650 | (f)57,000 | Falck
| | | | | | et al.
| | | | | | 1983.
10....|(c)11| 3,900 | 2.96 | (d)710 | (d)15,000 | Elinder
| | | | | | et al.
| | | | | | 1985.
11....|(c)12| 300 |.......|..........|...........| Roels
| | | | | | et al.
| | | | | | 1991.
12....| (g)8| 7,400 |.......|..........|...........| Roels
| | | | | | et al.
| | | | | | 1991.
13....|(c)23| (h)1,800 |.......|..........|...........| Roels
| | | | | | et al.
| | | | | | 1989.
14....| 10| 690 |.......|..........|...........| Iwao
| | | | | | et al.
| | | | | | 1980.
15....| 34| 71 |.......|..........|...........| Wibowo
| | | | | | et al.
| | | | | | 1982.
16....|(c)15| 4,700 | 6.49 | (d)590 | (d)93,000 | Thun
| | | | | | et al.
| | | | | | 1989.
______|_____|___________|_______|__________|___________|____________
The data provided from 15 of the 19 studies listed in Table 9 indicate that the geometric mean concentration of B2M observed among those who were not occupationally exposed to cadmium is 70-170 m g/g CRTU. Data from the 4 remaining studies indicate that exposed workers who exhibit no signs of proteinuria show mean B2MU levels of 60-300 m g/g CRTU. B2MU values in the study by Thun et al. (1989), however, appear high in comparison to the other 3 studies. If this study is removed, B2MU levels for those who are not occupationally exposed to cadmium are similar to B2MU levels found among cadmium-exposed workers who exhibit no signs of kidney dysfunction. Although the mean is high in the study by Thun et al., the range of measurements reported in this study is within the ranges reported for the other studies.
Determining a reasonable upper limit from the range of B2M concentrations observed among those who do not exhibit signs of proteinuria is problematic.
Elevated B2MU levels are among the signs used to define the onset of kidney dysfunction. Without access to the raw data from the studies listed in Table 9, it is necessary to rely on reported standard deviations to estimate an upper limit for normal B2MU concentrations (i.e., the upper 95th percentile for the distributions measured). For the 8 studies reporting a geometric standard deviation, the upper 95th percentiles for the distributions are 180-1140 m g/g CRTU. These values are in general agreement with the upper 95th percentile for the distribution (i.e., 631 m g/g CRTU) reported by Buchet et al. (1980). These upper limits also appear to be in general agreement with B2MU values (i.e., 100-690 m g/g CRTU) reported as the normal upper limit by Iwao et al. (1980), Kawada et al. (1989), Wibowo et al. (1982), and Schardun and van Epps (1987). These values must be compared to levels reported among those exhibiting kidney dysfunction to define a threshold level for kidney dysfunction related to cadmium exposure.
5.3.7.2 Range of B2MU concentrations among exposed workers
Table 10 presents results from studies reporting B2MU determinations among those occupationally exposed to cadmium in the work place; in some of these studies, kidney dysfunction was observed among exposed workers, while other studies did not make an effort to distinguish among exposed workers based on
kidney dysfunction. As with Table 9, this table provides geometric means and geometric standard deviations for the groups defined in each study if available. For studies reporting a geometric standard deviation along with a mean, the lower and upper 95th percentiles for the distributions are derived and reported in the table.
The data provided in Table 10 indicate that the mean B2MU concentration observed among workers experiencing occupational exposure to cadmium (but with undefined levels of proteinuria) is 160-7400 m g/g CRTU. One of these studies reports geometric means lower than this range (i.e., as low as 71 m g/g CRTU); an explanation for this wide spread in average concentrations is not available.
Seven of the studies listed in Table 10 report a range of B2MU levels among those diagnosed as having renal dysfunction. As indicated in this table, renal dysfunction (proteinuria) is defined in several of these studies by B2MU levels in excess of 300 m g/g CRTU (see footnote "c" of Table 10); therefore, the range of B2MU levels observed in these studies is a function of the operational definition used to identify those with renal dysfunction. Nevertheless, a B2MU level of 300 m g/g CRTU appears to be a meaningful threshold for identifying those having early signs of kidney damage. While levels much higher than 300 m g/g CRTU have been observed among those with renal dysfunction, the vast
majority of those not occupationally exposed to cadmium exhibit much lower B2MU concentrations (see Table 9). Similarly, the vast majority of workers not exhibiting renal dysfunction are found to have levels below 300 m g/g CRTU (Table 9).
The 300 m g/g CRTU level for B2MU proposed in the above paragraph has support among researchers as the threshold level that distinguishes between cadmium-exposed workers with and without kidney dysfunction. For example, in the guide for physicians who must evaluate cadmium-exposed workers written for the Cadmium Council by Dr. Lauwerys, levels of B2M greater than 200-300 m g/g CRTU are considered to require additional medical evaluation for kidney dysfunction (exhibit 8-447, OSHA docket H057A). The most widely used test for measuring B2M (i.e., the Pharmacia Delphia test) defines B2MU levels above 300 m g/l as abnormal (exhibit L-140-1, OSHA docket H057A).
Dr. Elinder, chairman of the Department of Nephrology at the Karolinska Institute, testified at the hearings on the proposed cadmium rule. According to Dr. Elinder (exhibit L-140-45, OSHA docket H057A), the normal concentration of B2MU has been well documented (Evrin and Wibell 1972; Kjellstrom et al. 1977a; Elinder et al. 1978, 1983; Buchet et al. 1980; Jawaid et al. 1983; Kowal and Zirkes, 1983). Elinder stated that the upper 95 or 97.5
percentiles for B2MU among those without tubular dysfunction is below 300 m g/g CRTU (Kjellstrom et al. 1977a; Buchet et al. 1980; Kowal and Zirkes, 1983). Elinder defined levels of B2M above 300aeg/g CRTU as "slight" proteinuria.
5.3.8 Conclusions and Recommendations for B2MU
Based on the above evaluation, the following recommendations are made for a B2MU proficiency testing program. Note that the following discussion addresses only sampling and analysis for B2MU determinations (i.e., to be reported as an unadjusted m g B2M/l urine). Normalizing this result to creatinine requires a second analysis for CRTU (see Section 5.4) so that the ratio of the 2 measurements can be obtained.
5.3.8.1 Recommended method
The Pharmacia Delphia method (Pharmacia 1990) should be adopted as the standard method for B2MU determinations. Laboratories may adopt alternate methods, but it is the responsibility of the laboratory to demonstrate that alternate methods provide results of comparable quality to the Pharmacia Delphia method.
5.3.8.2 Data quality objectives
The following data quality objectives should facilitate interpretation of analytical results, and should be achievable based on the above evaluation.
Limit of Detection. A limit of 100 m g/l urine should be achievable, although the insert to the test kit (Pharmacia 1990) cites a detection limit of 150 m g/l; private conversations with representatives of Pharmacia, however, indicate that the lower limit of 100 m g/l should be achievable provided an additional standard of 100 m g/l B2M is run with the other standards to derive the calibration curve (Section 3.3.1.1). The lower detection limit is desirable due to the proximity of this detection limit to B2MU values defined for the cadmium medical monitoring program.
Accuracy. Because results from an interlaboratory proficiency testing program are not available currently, it is difficult to define an achievable level of accuracy. Given the general performance parameters defined by the insert to the test kits, however, an accuracy of + 15% of the target value appears achievable.
Due to the low levels of B2MU to be measured generally, it is anticipated that the analysis of creatinine will contribute relatively little to the overall
variability observed among creatinine-normalized B2MU levels (see Section 5.4). The initial level of accuracy for reporting B2MU levels under this program should be set at + 15%.
Precision. Based on precision data reported by Pharmacia (1990), a precision value (i.e., CV) of 5% should be achievable over the defined range of the analyte. For internal QC samples (i.e., recommended as part of an internal QA/QC program, Section 3.3.1), laboratories should attain precision near 5% over the range of concentrations measured.
5.3.8.3 Quality assurance/quality control
Commercial laboratories providing measurement of B2MU should adopt an internal QA/QC program that incorporates the following components: Strict adherence to the Pharmacia Delphiad method, including calibration requirements; regular use of QC samples during routine runs; a protocol for corrective actions, and documentation of these actions; and, participation in an interlaboratory proficiency program. Procedures that may be used to address internal QC requirements are presented in Attachment 1. Due to differences between analyses for B2MU and CDB/CDU, specific values presented in Attachment 1 may have to be modified. Other components of the program (including characterization runs),
however, can be adapted to a program for B2MU.
5.4 Monitoring Creatinine in Urine (CRTU)
Because CDU and B2MU should be reported relative to concentrations of CRTU, these concentrations should be determined in addition CDU and B2MU determinations.
5.4.1 Units of CRTU Measurement
CDU should be reported as m g Cd/g CRTU, while B2MU should be reported as m g B2M/g CRTU. To derive the ratio of cadmium or B2M to creatinine, CRTU should be reported in units of g crtn/l of urine. Depending on the analytical method, it may be necessary to convert results of creatinine determinations accordingly.
5.4.2 Analytical Techniques Used to Monitor CRTU
Of the techniques available for CRTU determinations, an absorbance spectrophotometric technique and a high-performance liquid chromatography (HPLC) technique are identified as acceptable in this protocol.
5.4.3 Methods Developed for CRTU Determinations
CRTU analysis performed in support of either CDU or B2MU determinations should be performed using either of the following 2 methods:
1. The Du Pont method (i.e., Jaffe method), in which creatinine in a sample reacts with picrate under alkaline conditions, and the resulting red chromofore is monitored (at 510 nm) for a fixed interval to determine the rate of the reaction; this reaction rate is proportional to the concentration of creatinine present in the sample (a copy of this method is provided in Attachment 2 of this protocol); or,
2. the OSHA SLC Technical Center (OSLTC) method, in which creatinine in an aliquot of sample is separated using an HPLC column equipped with a UV detector; the resulting peak is quantified using an electrical integrator (a copy of this method is provided in Attachment 3 of this protocol).
5.4.4 Sample Collection and Handling
CRTU samples should be segregated from samples collected for CDU or B2MU analysis. Sample-collection techniques have been described under Section 5.2.4.
Samples should be preserved either to stabilize CDU (with HNO 3) or B2MU (with NaOH). Neither of these procedures should adversely affect CRTU analysis (see Attachment 3).
5.4.5 General Method Performance
Data from the OSLTC indicate that a CV of 5% should be achievable using the OSLTC method (Septon, L private communication). The achievable accuracy of this method has not been determined.
Results reported in surveys conducted by the CAP (CAP 1991a, 1991b and 1992) indicate that a CV of 5% is achievable. The accuracy achievable for CRTU determinations has not been reported.
Laboratories performing creatinine analysis under this protocolshould be CAP accredited and should be active participants in the CAP surveys.
5.4.6 Observed CRTU Concentrations
Published data suggest the range of CRTU concentrations is 1.0-1.6 g in 24-hour urine samples (Harrison 1987). These values are equivalent to about 1 g/l
urine.
5.4.7 Conclusions and Recommendations for CRTU
5.4.7.1 Recommended method
Use either the Jaffe method (Attachment 2) or the OSLTC method (Attachment 3). Alternate methods may be acceptable provided adequate performance is demonstrated in the CAP program.
5.4.7.2 Data quality objectives
Limit of Detection. This value has not been formally defined; however, a value of 0.1 g/l urine should be readily achievable.
Accuracy. This value has not been defined formally; accuracy should be sufficient to retain accreditation from the CAP.
Precision. A CV of 5% should be achievable using the recommended methods.
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| | ATTACHMENT 1: NONMANDATORY PROTOCOL FOR AN INTERNAL QUALITY ASSURANCE/QUALITY CONTROL PROGRAM | |
| | The following is an example of the type of internal quality assurance/quality control program that assures adequate control to satisfy mandatory OSHA requirements under this protocol. However, other approaches may also be acceptable. | |
| | As indicated in Section 3.3.1 of the protocol, the mandatory QA/QC program for CDB and CDU should address, at a minimum, the following: | |
| | <<degrees>> establishment of control limits; | |
| | <<degrees>> internal QC analyses and maintaining control; and | |
| | <<degrees>> corrective action protocols. | |
| | This illustrative program includes both initial characterization runs to establish the performance of the method and ongoing analysis of quality control samples intermixed with compliance samples to maintain control. | |
| | Before any analytical runs are conducted, the analytic instrument must be calibrated. This is to be done at the beginning of each day on which quality control samples and/or compliance samples are run. Once calibration is established, quality control samples or compliance samples may be run. Regardless of the type of samples run, every fifth sample must be a standard to assure that the calibration is holding. | |
| | Calibration is defined as holding if every standard is within plus or minus ( +) 15% of its theoretical value. If a standard is more than plus or minus 15% of its theoretical value, then the run is out of control due to calibration error and the entire set of samples must either be reanalyzed after recalibrating or results should be recalculated based on a statistical curve derived from the measurement of all standards. | |
| | It is essential that the highest standard run is higher than the highest sample run. To assure that this is the case, it may be necessary to run a high standard at the end of the run, which is selected based on the results obtained over the course of the run. | |
| | All standards should be kept fresh, and as they get old, they should be compared with new standards and replaced. | |
| | INITIAL CHARACTERIZATION RUNS AND ESTABLISHING CONTROL | |
| | A participating laboratory should establish four pools of quality control samples for each of the analytes for which it wishes to be accredited. The concentrations of quality control samples within each pool are to be centered around each of the four target levels for the particular analyte identified in Section 4.4 of the protocol. | |
| | Within each pool, at least 4 quality control samples need to be established with varying concentrations ranging between plus or minus 50% of the target value of that pool. Thus for the medium-high cadmium in blood pool, the theoretical values of the quality control samples may range from 5 to 15 m g/l, (the target value is m 10 g/l). At least 4 unique theoretical values must
be represented in this pool. | |
| | The range of theoretical values of plus or minus 50% of the target value of a pool means that there will be overlap of the pools. For example, the range of values for the medium-low pool for cadmium in blood is 3.5 to 10.5 m g/l while the range of values for the medium-high pool is 5 to 15 m g/l. Therefore, it is possible for a quality control sample from the medium-low pool to have a higher concentration of cadmium than a quality control sample from the medium-high pool. | |
| | Quality control samples may be obtained as commercially available reference materials, internally prepared, or both. Internally prepared samples should be well characterized and traced or compared to a reference material for which a consensus value for concentration is available. Levels of analyte in the quality control samples must be concealed from the analyst prior to the reporting of analytical results. Potential sources of materials that may be used to construct quality control samples are listed in Section 3.3.1 of the protocol. | |
| | Before any compliance samples are analyzed, control limits must be established. Control limits should be calculated for every pool of each analyte for which a
laboratory seeks accreditation, and control charts should be kept for each pool of each analyte. A separate set of control charts and control limits should be established for each analytical instrument in a laboratory that will be used for analysis of compliance samples. | |
| | At the beginning of this QA/QC program, control limits should be based on the results of the analysis of 20 quality control samples from each pool of each analyte. For any given pool, the 20 quality control samples should be run on 20 different days. Although no more than one sample should be run from any single pool on a particular day, a laboratory may run quality control samples from different pools on the same day. This constitutes a set of initial characterization runs. | |
| | For each quality control sample analyzed, the value F/T (defined in the glossary) should be calculated. To calculate the control limits for a pool of an analyte, it is first necessary to calculate the mean, X, of the F/T values for each quality control sample in a pool and then to calculate its standard deviation -- S. Thus, for the control limit for a pool, X is calculated as: | |
| | and -- S is calculated as | |
| | where N is the number of quality control samples run for a pool. | |
| | The control limit for a particular pool is then given by the mean plus or minus 3 standard deviations (X + 3 -- S). | |
| | The control limits may be no greater than 40% of the mean F/T value. If three standard deviations are greater than 40% of the mean F/T value, then analysis of compliance samples may not begin [FN1]. Instead, an investigation into the causes of the large standard deviation should begin, and the inadequacies must be remedied. Then, control limits must be reestablished which will mean repeating the running 20 quality control samples from each pool over 20 days. | |
| | [FN1] Note that the value, "40%" may change over time as experience is
gained with the program. | |
| | INTERNAL QUALITY CONTROL ANALYSES AND MAINTAINING CONTROL | |
| | Once control limits have been established for each pool of an analyte, analysis of compliance samples may begin. During any run of compliance samples, quality control samples are to be interspersed at a rate of no less than 5% of the compliance sample workload. When quality control samples are run, however, they should be run in sets consisting of one quality control sample from each pool. Therefore, it may be necessary, at times, to intersperse quality control samples at a rate greater than 5%. | |
| | There should be at least one set of quality control samples run with any analysis of compliance samples. At a minimum, for example, 4 quality control samples should be run even if only 1 compliance sample is run. Generally, the number of quality control samples that should be run are a multiple of four with the minimum equal to the smallest multiple of four that is greater than 5% of the total number of samples to be run. For example, if 300 compliance samples of an analyte are run, then at least 16 quality control samples should be run (16 is the smallest multiple of four that is greater than 15, which is 5% of 300). | |
| | Control charts for each pool of an analyte (and for each instrument in the laboratory to be used for analysis of compliance samples) should be established by plotting F/T versus date as the quality control sample results are reported. On the graph there should be lines representing the control limits for the pool, the mean F/T limits for the pool, and the theoretical F/T of 1.000. Lines representing plus or minus ( +) 2 S should also be represented on the charts. A theoretical example of a control chart is presented in Figure 1 | |
| | THEORETICAL EXAMPLE OF A CONTROL CHART FOR A POOL OF AN ANALYTE | |
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