A comparative assessment of some biochemical markers

in occupational lead exposure.

Bhagwat V R1, Patil A J2 and Bhagwat K V3

1. Prof & HOD (Biochemistry), SBH Govt. Medical College, DHULE.2. Associate Professor (Biochemistry) KIMSU,KARAD. 3. Asstt Professor, Humanity Dept, SSVP’s College of Engineering, DHULE (Maharashtra)

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Introduction

Lead, an industrially useful metal with wide applications.

Industrialization use ………… Pollution.

Biologically Pb has no role, useless, unwanted mineral for living organisms.

Industrial use of LEAD

Occupations such as Lead battery manufacturing, silver jewelry making, painting have inherent dangers of lead toxicity.

Occupational exposure at small scale units in developing countries unregulated.

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LEA

D

Silv

er Je

welry

Lead is used as alloy in making all types of Silver Jewelry. And for marking designs on silver rings.

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Lead

(

Pb)

Metarubricytosis and slight basophilic stippling of erythrocytes in lead toxicosis. The remaining erythrocytes appear hypochromic because lead inhibits the synthesis of hemoglobin (Wright-Leishman stain).

Lead toxicity most organ systems. Major Hematological, Nervous etc

Moderate to clinically significant toxicity is very common in small scale industrial units.

Lead lines

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Lead interference

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Enzyme inhibition by LeadLead inhibits 3 enzymes in Hb

biosynthesis1. δ-Amino Levulinic acid

dehydratase (ALAD) (Step 2)

2. Coproporpyrinogen oxidase. (Step 6)

3. Ferrochelatase (step 8)

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Erythropiesis and lead

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Purpose of the study:To assess the lead toxicity in SSI workers and to evaluate biochemical markers suitable for screening occupational exposure in developing country like India.

Aims & Objecti

ves

Mate

rial a

nd

Meth

od

s

Setting & Design: Industrial, Rural, Case control study.

Subjects: Employees of SSI units of,› Battery manufacturing & recycling.› Silver jewelry making.› Spray painting. Size & selection: 30 from each category,

Random, non-stratified.

Controls: 30 healthy subjects from rural areas,

clinically normal, age matched (non-exposed) Samples:

Random blood, urine samples with proper preservatives. Processing (Pre analytical treatment)

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Methods of sample analysis

Lead (Pb) by Atomic absorption spectrophotometry (Perkin Elmer A300; Abs λ 283.3 nm).

Zinc Protoporphyrin (ZPP): by Hematoflurometry (Wayne NA NSSLS – C42A; 1996). Ex 415 nm, Em 595 nm)

δ-amino levulinic acid (ALA) by colorimetric method (Wada et al Br J Industr Med 1969; 26: 240-43).

Porphobilinogen (PBG) by colorimetric method, Ehrlich’s reaction (Mauzeroall & Granick, JBC 1956; 435-46).

Cost-Effective index: Cost-Benefit analysisStatistical methods: Mean, SD, Student t test.

Scheme of ALA analysis

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Marker(units)

BMWn=30

SJMn=30

SPn=30

NECn =35

1 BPb(μg/dl)

53.65 ± 16.98(25.8 – 78.0)

48.56 7.39*(30.2 - 64.7)

22.32 8.87*(7.5 - 45.7)

12.52 ± 4.08(2.8 – 22.0)

2 UPb(μg/dl)

20.04 ± 15.20(5.2 – 62.8)

9.39 6.52

(2.0 - 30.7)11.22 7.15**

(1.5 - 30.2)6.97 ± 3.59(1 – 13.2)

3 ZPP(μg/dl)

122.6 ± 201.5(10 – 972)

9.55 3.21

(1.0 - 11)15.67 4.24*(8.12 - 23.12)

8.25 ± 7.79(1 – 35)

4 ALA(mg/dl)

38.10 ± 17.52(9.3 – 74.9)

25.13 9.52*(9.38- 50)

11.82 3.53***

(7.05 - 21.17)9.62 ± 5.45(2.5 – 17.5)

5 PBG(mg/dl)

13.95 ± 5.23(7.0 - 24.7)

12.06 4.81***

(7.05 - 24.7)10.45 3.33

(1.0 - 57)10.10 ± 2.87(3.5 – 15.8)

Table I Biochemical markers in the occupation groups.

(Figures are mean + SD. Those in parenthesis are range of values).BMW = Battery Manufacture workers; SJM = Silver Jewelry makers; SP = Spray painters; NEC = Non exposed controls, BPb = Blood lead; UPb = Urine lead; ZPP = Erythrocyte zinc protoporphyrin; ALA = Urinary δ-Amino levulinic acid; PBG = Urinary porphobilinogen.Figures in red are highest values & those in blue are the lowest one.

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Biochemical markers in lead exposure

0

20

40

60

80

100

120

140

BPb UPb eZPP uALA uPBGMarkers

va

lue

s

BMW SJM SP NEC

BMW = Battery workers, SJM = Silver Jewelry makers, SP = Spray painters, NEC = Controls.BPb = Blood lead, UPb =Urinary lead, eZPP = Erythrocyte zinc protoporphyrin,

uALA = Urinary δ-amino levulinic acid, uPBG = Urinary Porphobilinogen.

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Lead levels in Subjects

BPb, 53.63

BPb, 48.56

BPb, 22.32

BPb, 12.52

UPb, 20.04

UPb, 9.39

UPb, 11.22

UPb, 6.97

0 10 20 30 40 50 60 70 80

BMW

SJM

SP

NEC

Microgram /dl

BMW = Battery workers, SJM = Silver Jewelry makers, SP = Spray painters, NEC = Controls.BPb = Blood lead, UPb =Urinary lead, eZPP = Erythrocyte zinc protoporphyrin,

uALA = Urinary δ-amino levulinic acid, uPBG = Urinary Porphobilinogen.

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Table IIComparison of biochemical markers for screening

occupational Pb exposure.

ZPP PBG ALA

Precision (%) CV Intra assay Inter assay

4.92.3

4.23.2

2.81.5

Sensitivity Non-linear Poor Linear ~40 mg/L

SpecificityPoor

( Chr Fe defy anemia, Porphyria, febrile state)

Moderate( Porphyria)

High(Amino acetone ns)

Correlation vs PbB 0.3789 0.1648 0.5798

Normal Range 1–16 μg/dl 7–13 mg/L 4–15 mg/L

95% CI 5.5 - 11.0 9.1 - 11.1 7.7 - 11.5

Level at PbB = 20 μg/dl 21.32 10.97 14.67

Level at PbB = 50 μg/dl 132.6 12.58 34.19

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Table IIIComparison of biochemical markers for screening

occupational Pb exposure.

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ZPP PBG ALAAssay design (steps) Simplest (1) Simple (4) Multi step (8)Time/test < 3 mins ~ 8 mins ~ 20 minsSamples used WB (R) Urine (R) Urine (R)Storage, Stability <4.°C, 5 w pH 8-9, dark,

<4°C, 2wpH 3-4, dark,

<4°C, 2 wInterference in assay

Bilirubin, Riboflavin,several drugs >5

Ehrlich’ +ve substances

Amino acetone(NR 0 – 1 mg/L)

Specificity Poor Moderate HighTest type Invasive Non-Invasive Non-InvasiveCost/test (2004) 35.11 16.98 35.53Cost effective index 3.3 2.4 7.1Rating for screening *** ** *****

ConclusionZPP is not a suitable marker for

screening of occupational lead exposure due to poor specificity which makes it unfit for screening low level toxicity at earlier stages. Urinary ALA is ideal & convenient marker as it has higher sensitivity & specificity apart from being an cost effective non-invasive investigation.

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