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Research Article Assessment of Diagnostic Accuracy and Optimal Cut Points of Blood Lead Levels on Serum Proteins Among Workers E

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Research Article Assessment of Diagnostic Accuracy and Optimal Cut Points of Blood Lead Levels on Serum Proteins Among Workers E INTERNATIONAL JOURNAL OF MEDICAL BIOCHEMISTRY DOI: 10.14744/ijmb.2019.41736 Int J Med Biochem 2019;2(3):81-7 Research Article Assessment of diagnostic accuracy and optimal cut points of blood lead levels on serum proteins among workers exposed to Pb at a lead battery plant Ravibabu Kalahasthi, Tapu Barman, Ravi Prakash Jamalpur, Vinay Kumar Adepu Department of Biochemistry, Regional Occupational Health Centre (Southern), Bangalore, India Abstract Objectives: This study assessed the diagnostic accuracy and the optimal cut point of blood lead (Pb) level (BLL) in serum proteins (total protein, albumin, globulin, and albumin/globulin ratio) among workers with exposure to Pb at a Pb battery plant. Methods: An examination of the diagnostic accuracy and optimal cut point value of BLL in serum proteins among workers exposed to Pb was performed using analysis of the area under the curve (AUC) and coordinates of receiving operating characteristic (ROC) curve. The study group consisted of 176 males who worked in a Pb battery plant. A con- trol group consisted of 80 male office workers with no unusual occupational Pb exposure. The BLL of the patients was assessed using an atomic absorption spectrophotometer. The serum protein levels (total protein and albumin) were determined using diagnostic kits. The serum globulin was calculated from the total protein and albumin measures. The albumin:globulin ratio (A/G ratio) was calculated from the albumin and globulin levels. Results: The serum total protein and globulin concentration values were lower and the A/G ratio was significantly higher in the study group compared with the control group. A negative and significant association was found between the BLL and serum total protein and serum globulin values. A positive and significant association was found between the BLL and the serum albumin level and the A/G ratio. The AUC value was 0.626 (95% confidence interval [CI]: 0.540– 0.712; p=0.005) for serum total protein with a cutoff value of ≤6.9 g/dL. For serum albumin, the AUC value was 0.466 (95% CI: 0.344–0.587; p=0.574) with a cutoff value of ≤4.0 g/dL. The AUC value of serum globulin was 0.625 (95% CI: a0.541–0.708, p=0.005) with a cutoff value of ≤2.4g/dL. The A/G ratio had an AUC value of 0.606 (95% CI: 0.521–0.692; p=0.012) with a cutoff value >1.5g/dL. An optimal BLL cut point value of 25.5µg/dL was noted for serum total protein, with 56% sensitivity and 51% specificity. The BLL value of 24.5 µg/dL was determined to be the optimal cut point for serum albumin, with 44% sensitivity and 44% specificity. The use of 27.5 µg/dL as an optimal BLL cut point value for serum globulin and the A/G ratio demonstrated 55% sensitivity and 55% specificity. Conclusion: The AUC values of serum total protein, globulin, and the A/G ratio were found to be significant when compared with the serum albumin. BLL optimal cut point values for serum proteins in Pb exposure ranged from 24.5 µg/dL to 27.5 µg/dL. The change in serum total protein, globulin, and A/G ratio suggested reduced liver function due to Pb-binding interactions. Keywords: Area under the curve, blood lead level, coordinates of receiver operating characteristic curve, serum proteins ead (Pb)-binding proteins originate in the kidney, brain, associated with Pb poisoning [2]. Systemic effects of hyper- Lerythrocytes, liver, and lung [1]. Proteins related to heme tension, frank anemia, cognitive deficits, infertility, immune synthesis, calcium metabolism, and neurotransmission are imbalances, delayed skeletal and deciduous dental develop- Address for correspondence: Ravibabu Kalahasthi, M.Sc. PhD. Department of Biochemistry, Regional Occupational Health Centre (Southern), Bangalore, India Phone: 22172500 E-mail: [email protected] ORCID: 0000-0003-2728-7382 Submitted Date: April 24, 2019 Accepted Date: June 27, 2019 Available Online Date: October 09, 2019 ©Copyright 2019 by International Journal of Medical Biochemistry - Available online at www.internationalbiochemistry.com OPEN ACCESS This work is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License. 82 Int J Med Biochem ment, vitamin D deficiency, and gastrointestinal symptoms Individuals with a history of diabetes; liver or renal disorders; have been related to Pb exposure [3]. Pb intoxication in animals fever; malignancy; or autoimmune, chronic inflammatory, and workers exposed to Pb in the plastics industry resulted in hematological or rheumatological disorders were excluded significantly altered levels of serum aminotransaminases (as- from the study. The Institutional ethical committee of Re- partate aminotransferase [AST] and alanine aminotransferase gional Occupational Health Centre (Southern) approved this [ALT]), γ-glutamyl transpeptidase, and alkaline phosphatase, study on 3-12-2014 with letter no.142/6. The subjects were which was characterized as a hepatotoxic effect and was fol- informed about the objectives of the study and consent was lowed by intrahepatic cholestasis [4-7]. obtained before initiating their participation in the study. Animal models with acute and chronic Pb exposure have shown decreased total and protein-bound sulfhydryl (SH) Sample collection groups in homogenates and synaptosomes of the brain [8]. Whole blood samples of 5 mL (2 mL in heparin tubes +3 mL in Pb intoxication in animals has revealed a significant decrease plain tubes) were collected from each subject. The 2 mL hep- in serum total protein, albumin, and the A/G ratio [9, 10]. The arinized blood sample was used for BLL estimation. The 3 mL reduction of proteins was due to precipitation of soluble pro- whole blood sample collected in plain tubes was centrifuged teins, inhibition of protein biosynthesis through the specific at 3000 rpm for 10 minutes at 4oC to separate the serum from enzymes in cell processes, and a decrease in free amino acid red blood cells and used to examine serum proteins. utilization for protein synthesis. Occupational exposure to Pb has also been reported to cause decreased protein SH groups [11] and elevated positive acute phase proteins [12]. Recent Serum total protein studies have reported decreased levels of serum total protein, The serum total protein concentration was determined using albumin, globulin, and the A/G ratio in workers exposed to Pb the biuret method. In this approach, alkaline copper reacts in the automobile industry [13, 14], Pb battery manufacturing with the peptide bonds of proteins to form a characteristic [15, 16], petrol filling stations, and traffic officers [17]. pink to purple biuret complex. Sodium potassium tartrate Measurement of diagnostic test accuracy can include measures prevents copper hydroxide precipitation and potassium io- of sensitivity and specificity, positive and negative predictive dide prevents the auto reduction of copper. The color in- values, likelihood ratio, area under the receiver operating char- tensity is directly proportional to protein concentration. The acteristic (AUROC) curve, Youden’s index, and a diagnostic odds absorbance was measured at 546 nm. The concentration of ratio. The AUROC is defined as a plot of sensitivity with the Y- serum total protein was expressed as g/dL. axis plotted against the 1-specificity, or false positive rates, on the X-axis. It is an effective tool used for evaluating quality and performance of a diagnostic test. A test with an AUROC value Serum albumin of 1 is perfectly accurate. The lower limit for the AUROC of a di- The serum albumin concentration was measured using the agnostic test is 0.5. The line segment from (0.0) to (1.1) has an bromocresol green method described by Doumas et al. area of 0.5. The coordinates of the ROC curve help to identify [19]. The albumin in a buffered solution reacts with anionic the optimal cutoff values of a diagnostic test [18]. bromocresol green dye and produces a green color. The mea- Most of the animal and human occupational Pb exposure surement was made at an absorbance at 628 nm. The intensity studies have reported decreased serum protein measures (to- of the green color is proportional to the concentration of albu- tal protein, albumin, globulin, and A/G ratio). However, the di- min present in the sample and was expressed as g/dL. agnostic accuracy and optimal cut point values have not been explored. The aim of this study was to evaluate the diagnostic Serum globulin accuracy and optimal cut point values for BLL in serum pro- The serum globulin level was calculated by subtracting the teins for workers exposed to Pb through employment at a Pb albumin value from the corresponding value of total protein. battery plant using AUROC and coordinates of the ROC curve. The concentration of serum globulin was expressed as g/dL. Materials and Methods Albumin and globulin ratio The study group consisted of 176 male subjects who worked The A/G ratio was calculated based on the measurements of at a Pb battery manufacturing facility located in Tamil Nadu, serum albumin and globulin. India. A control group comprised 80 healthy male subjects who worked in administrative jobs with no exceptional occu- pational exposure to Pb. The serum protein values (total pro- Blood lead level tein, albumin, globulin, and A/G ratio) of the study and con- The estimation of BLL was used as an indicator of Pb expo- trol groups were compared. This study evaluated diagnostic sure. The BLL was measured using the method described by accuracy and optimal cutoff point values of BLL in serum pro- Barman et al. [20]. A 2-mL whole blood sample was prepared teins using analysis of AUROC and coordinates of ROC curves. using a microwave digestion system (ETHOS-D, Milestone Srl, Kalahasthi, Diagnostic accuracy of serum protein / doi: 10.14744/ijmb.2019.41736 83 Sorisole, Italy) with 2 mL of nitric acid (HNO ) and 0.2 mL of 3 Table 2.
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