Biological Trace Element Research

, Volume 142, Issue 3, pp 350–361 | Cite as

Evaluation of Status of Cadmium, Lead, and Nickel Levels in Biological Samples of Normal and Night Blindness Children of Age Groups 3–7 and 8–12 Years

  • Hassan Imran Afridi
  • Tasneem Gul Kazi
  • Naveed Kazi
  • Sirajuddin
  • Ghulam Abbas Kandhro
  • Jameel Ahmed Baig
  • Abdul Qadir Shah
  • Sham Kumar Wadhwa
  • Sumaira Khan
  • Nida Fatima Kolachi
  • Faheem Shah
  • Mohammad Khan Jamali
  • Mohammad Balal Arain
Article

Abstract

The causes of night blindness in children are multifactorial, and particular consideration has been given to childhood trace metals toxicity, which is the most common problem found in underdeveloped countries. This study was designed to compare the levels of cadmium (Cd), lead (Pb), and nickel (Ni) in scalp hair, blood, and urine of night blindness children age ranged 3–7 and 8–12 years of both genders, comparing them to sex- and age-matched controls. A microwave-assisted wet acid digestion procedure was developed as a sample pretreatment, for the determination of Cd, Pb, and Ni in biological samples of night blindness children. The proposed method was validated by using conventional wet digestion and certified reference samples of hair, blood, and urine. The digests of all biological samples were analyzed for Cd, Pb, and Ni by electrothermal atomic absorption spectrometry. The results indicated significantly higher levels of Cd, Pb, and Ni in the biological samples (blood, scalp hair, and urine) of male and female night blindness children, compared with control subjects of both genders. These data present guidance to clinicians and other professional investigating toxicity of trace metals in biological samples of night blindness children.

Keywords

Night blindness Cadmium Lead Nickel Biological samples Wet acid digestion methods Children Age groups (3–12) Atomic absorption spectrophotometer 

Introduction

Night blindness may be an early sign of vitamin A deficiency. Such a deficiency may result from a diet low in animal foods (the main source of vitamin A and essential minerals) such as eggs, dairy products, organ meats, and fish. Christian et al. and Brody [1, 2] showed that vitamin A deficiency is widespread in developing countries.

Human cells employ metals, such as copper, zinc, and iron, to control significant metabolic and signaling functions, making them essential for life. Other metals can be potentially toxic such as the heavy metals lead (Pb), nickel (Ni), cadmium (Cd), mercury, and thallium. Pb, in particular, is a neurotoxin that has been linked to visual deterioration [3], central and peripheral nervous system disorders [4], renal dysfunction [5], and hypertensive cardiovascular disease [6]. Recently, Eichenbaum and Zheng [7] showed that the retina and choroid can accumulate the heavy metal Pb.

Pb-induced toxicities continue to be a major public health concern. In addition to its toxic effect on the central nervous system, Pb toxicity has been linked to renal dysfunction, hypertensive cardiovascular disease, and auditory disturbances and visual disability [8]. Recent studies from a rat retinal model indicate that low-level exposure to Pb produces scotopic visual deterioration, characterized by diminished rod-mediated electroretinogram, rod and bipolar apoptotic cell death, and inhibited cyclic guanine monophosphate phosphodiesterase (cGMP PDE) [3]. While Pb toxicity has been linked to visual dysfunction in mammals, few studies have been performed to quantitate Pb distribution in human eyes. Even less has been learned about molecular and cellular mechanisms underlying Pb-induced visual disturbance. The retinal pigment epithelium serves as a defensive barrier to the retina in the same way as the choroid plexus functions to the brain [9]. The previous studies have shown that the choroid plexus sequesters Pb, resulting in reduced secretion of transthyretin (TTR) to the cerebrospinal fluid [10]. TTR is a 55,000-Da protein and functions as a carrier protein for thyroid hormones, retinal, and retinol-binding protein. In the eyes, TTR plays a critical role in transporting retinol to the photoreceptor for the phototransduction process [9]. However, little quantitative information is available concerning the distribution of TTR in human eyes.

Cd toxicity has been associated with renal disease, hypertension, and an increased prevalence of cardiovascular disease [11]. Heavy metals like Ni and Cd are ubiquitous pollutants that have permanently contaminated air, water, and soil. The toxic effect of heavy metals usually involves an interaction between the heavy metal ion and the specific target protein, resulting in a change in protein structure and function [12]. Cells involved in the transport of trace metals are particularly susceptible to toxicity. The retinal pigment epithelium is a metal-chelating tissue that is capable of binding essential and toxic heavy metals because of the high affinity of metals to melanin in retinal pigment epithelium melanosomes [13, 14].

In view of the above facts, it is important to determine the essential trace elemental concentrations in biological samples of humans having physiological disorders, such as myocardial infarction. Various biopsy materials such as serum, scalp hair, urine, and other body fluids may be used as bio-indicators for these purposes [15]. Blood elemental analysis provides information about what the body has recently (hours to days, in some cases, weeks) absorbed. Blood levels are largely independent of tissue deposition [16]. Whole blood analysis measures total element levels that circulate extracellularly (serum/plasma) as well as intracellularly (function within blood cells). Intracellular elements are those that have specific functions within circulating blood cells [17]. This homeostatic response illustrates the effective clearance mechanisms in the blood and largely explains the short-term utility of blood analysis. The importance of exploring the depot-storage capacities of various elements, particularly the toxic ones, remains a vital aspect in elemental analysis largely met by urine and hair testing [18]. Urine analysis can provide important information to the clinician that may not be readily available with blood analysis. Levels of nutrient elements in the blood and the excreted urine are tightly controlled via metabolic, reabsorptive, and excretory mechanisms. Consequently, most urine testing is not helpful in nutritional element assessment [19, 20]. Hair analysis is a simple diagnostic technique, based on the idea that hair provides vital clues about nutritional imbalances elsewhere in the body [21, 22].

Compared with blood or urine analysis, a couple of factors such as simplicity of matrix, relatively high concentration of trace elements, easy sampling, transfer, and storage should be considered. It is also used to detect environmental toxins before overt symptoms appear because metal concentrations in hair are usually tenfold higher than in other biological material [23]. Atomic absorption spectrometric methods are frequently used for the specific determination of very low elemental concentrations in biological samples [22]. At present, the mineralization methods frequently employed for the analysis of biological samples are wet digestion with concentrated acids using either convective systems or microwave ovens [15]. The main advantage of microwave-assisted pretreatment of samples is the small amount of mineral acids required and reduced production of nitrous vapors.

The main objectives of our study were (1) to assess the concentrations of Cd, Pb, and Ni in the biological samples (scalp hair, blood, and urine) of night blindness and normal children of both genders and with ages ranging between 3–7 and 8–12 years old; (2) to evaluate the variation of these elements in night blindness children was related with age and sex factors; and (3) to investigate about the etiologies of the detected night blindness and to evaluate the role of nutrition, based on dietary history to screen high risk children for night blindness in Pakistan.

Materials and Methods

Apparatus

The analysis of elements was carried out by means of a double beam Perkin Elmer atomic absorption spectrometer model 700 (Perkin Elmer, Norwalk, CT, USA) equipped with a flame burner and graphite furnace HGA-400 (Perkin Elmer), a pyrocoated graphite tube with an integrated platform, and an autosampler AS-800 (Perkin Elmer). The instrumental parameters are shown in Table 1. Cd, chromium, and Ni were measured under optimized operating conditions using electrothermal atomic absorption spectrometry. Signals were measured as absorbance peaks in the flame absorption mode, whereas integrated absorbance values (peak area) were determined in the graphite furnace. A Pel (PMO23, Japan) domestic microwave oven (maximum heating power of 900 W) was used for digestion of the biological samples. Acid-washed polytetrafluoroethylene (PTFE) vessels and flasks were used for preparing and storing solutions.
Table 1

The number of subjects as control and night blindness patients

Age groups

Control groups

Night blindness patients

Male

Female

Male

Female

3–7

46

49

39

31

8–12

58

57

43

37

Total

104

106

82

68

Reagents and Standard Solutions

Ultrapure water obtained from ELGA lab water system (Bucks, UK) was used throughout the work. Concentrated nitric acid (65%) and hydrogen peroxide (30%) from Merck (Darmstadt, Germany) were checked for possible trace metal contamination. Working standard solutions of Cd, Pb, and Ni were prepared immediately prior to their use by stepwise dilution of certified standard solutions (1,000 ppm) Fluka Kamica (Bush, Switzerland), with 0.2 M HNO3. All solutions were stored in polyethylene bottles at 4°C. For the accuracy of methodology, certified samples (certified reference materials) of human hair (BCR 397, Brussels®, Belgium), Clincheck control-lyophilized® human urine (® Recipe, Munich, Germany), and Clincheck control-lyophilized® human whole blood (Recipe, Munich, Germany) were used. All glassware and plastic materials used were previously soaked for 24 h in 2 M nitric acid, washed with distilled water, finally rinsed with Milli-Q water, dried, and stored in class 100 laminar flow hoods.

Sample Collection and Pretreatment

The study was conducted on 150 children aged ranged between 3 and 12 years of both genders, registered in the hospital as patients with ocular problems. A total of 82 children reported poor vision in the day and in the night, and 68 children have good visions in the day but poor vision in dim light or in the night. The healthy children of the same city and residential area of Hyderabad City were not registered in the hospital but checked by the ophthalmologist for eyesight. A control group of 210 healthy children with the same age and with normal eyesight was chosen; detail was reported in Table 1. The samples from both groups were collected three times in a year to evaluate any variation in the concentration of trace metals in patients and normal subjects during 1 year. The parents of night blindness and normal subjects were interviewed and asked to complete a questionnaire in order to collect details concerning physical data, ethnic origin, health, medical reports, and dietary habits. The biochemical tests were described in Table 2. Both vitamin A and carotene were analyzed by the method of Kimble [24]. The factors used to convert the L values obtained with the Evelyn colorimeter to International Units of vitamin A and picogram of carotene. All patients and control subjects underwent a routine eye examination including visual acuity, slit lamp examination, and ophthalmoscopic test; all tests were performed by well-trained and standardized specialists in the eye hospital. A file of complete information and all the demographical data was compiled. The consent of each contributor to use the data was asked. This research project was evaluated and approved by the Higher Education Commission, Pakistan.
Table 2

Clinical and biochemical characteristics of normal and night blindness subjects

Parameters

Male

Female

Normal range

Normal

Night blindness

Normal

Night blindness

3–7 years

 Height (cm)

106.4 ± 6.42

95.2 ± 5.61

105.7 ± 5.39

92.9 ± 6.33

 

 Weight (kg)

19.3 ± 1.56

15.8 ± 2.33

18.6 ± 1.12

13.9 ± 1.24

 

 Serum ferritin (μg/l)

31.9 ± 1.4

19.7 ± 2.32

33.7 ± 2.7

17.8 ± 1.8

<30

 Hemoglobin (mg/dl)

11.5 ± 0.6

9.9 ± 1.4

11.8 ± 0.6

9.6 ± 0.5

11.5–16.5

 Hematocrit (%)

43.7 ± 3.7

34.4 ± 1.7

40.2 ± 2.7

33.9 ± 2.3

35–55

 Vitamin A (IU)

136 ± 5.45

52.4 ± 4.98

137 ± 5.68

53.8 ± 4.67

>136–149

 Carotene (μg/100 ml plasma)

148.7 ± 13.8

59.8 ± 9.56

149.3 ± 11.3

56.4 ± 8.25

>140

 Red blood count (RBC (mm3))

4.4 ± 0.6

3.21 ± 0.6

4.5 ± 0.4

3.4 ± 0.8

3.5–5.5

 MCV (μm)

83.4 ± 3.24

74.3 ± 4.5

86.2 ± 1.9

73.6 ± 3.4

75–100

 MCH (pg)

31.4 ± 1.5

24.5 ± 1.3

33.6 ± 1.1

26.3 ± 0.7

25–35

 MCHC (g/dl)

34.3 ± 3.1

29.9 ± 2.3

36.8 ± 2.5

31.2 ± 2.3

31–38

 WBC (mm3)

6.9 ± 0.82

7.4 ± 0.5

6.67 ± 2.28

7.51 ± 0.42

3.5–10

 Platelets (mm3)

165.5 ± 16.5

179.2 ± 19.4

172 ± 8.9

193 ± 14.8

100–400

8–12 years

 Height (cm)

137.7 ± 1.32

125.4 ± 3.28

136.9 ± 1.68

124.7 ± 2.07

 

 Weight (kg)

31.9 ± 1.54

24.3 ± 1.87

32.2 ± 1.64

24.9 ± 1.53

 

 Serum ferritin (μg/l)

36.5 ± 2.7

28.9 ± 2.34

37.3 ± 4.4

29.4 ± 3.52

<30

 Hemoglobin (mg/dl)

12.2 ± 1.3

11.3 ± 1.4

12.5 ± 1.23

11.8 ± 2.65

11.5–16.5

 Hematocrit (%)

45.3 ± 3.21

36.9 ± 2.42

46.5 ± 1.76

34.5 ± 4.6

35–55

 Vitamin A (IU)

145 ± 3.66

56.9 ± 3.65

142 ± 3.97

58.9 ± 5.36

>136–149

 Carotene (μg/100 ml plasma)

159 ± 14.9

57.2 ± 5.43

153 ± 9.87

54.3 ± 5.62

>140

 Red blood count (RBC (mm3))

4.62 ± 0.35

3.79 ± 0.32

4.76 ± 0.62

3.89 ± 0.54

3.5–5.5

 MCV (μm)

85.4 ± 3.4

72.2 ± 5.1

88.2 ± 4.32

75.8 ± 3.5

75–100

 MCH (pg)

32.9 ± 2.6

27.8 ± 2.15

35.5 ± 2.67

29.8 ± 2.56

25–35

 MCHC (g/dl)

37.2 ± 2.46

29.8 ± 3.6

37.9 ± 1.8

29.9 ± 3.62

31–38

 WBC (mm3)

7.33 ± 1.52

7.13 ± 1.23

7.24 ± 0.36

7.02 ± 0.48

3.5–10

 Platelets (mm3)

236 ± 19.7

247 ± 15.6

229 ± 20.2

245 ± 22.6

100–400

Venous blood samples (3–5 ml) were collected using metal-free Safety Vacutainer blood collecting tubes (Becton Dickinson, Rutherford®, USA) containing >1.5 μg K2EDTA/mL and were stored at −20°C until analysis. Urine was voided directly into an acid-washed 100 ml polyethylene tubes (Kartell1, Milan, Italy), which had been decontaminated before handling. Between sampling sessions, the container was wrapped in a clean polyethylene bag. Urine samples were acidified with 1% ultrapure HNO3. Prior to sub-sampling for analysis, the sample was shaken vigorously for 1 min to ensure a homogeneous suspension. Hair samples of children were collected from the children in an area of the cranium 2–3 cm above the nape of the neck. The scalp hair samples were washed as in our previous studies [25]. After washing, hair samples were dried at 80°C for 6 h. Hair samples were put into separate plastic envelopes with an identification number for each participant.

Conventional Digestion Method

Duplicate 0.5 mL of each certified urine and blood samples and 0.2 g of human hair samples BCR 397 were placed individually into 50 mL Pyrex flasks. Five milliliters of a freshly prepared mixture of concentrated HNO3–H2O2 (2:1, v/v) were added to each flask, which were heated on a electric hot plate at 80°C, for 2–3 h, till clear transparent digests were obtained. Final solutions were made up to 10 mL with 2 M HNO3. The final solutions were collected in polyethylene flask and stored at 4°C till analysis. Duplicate biological samples (scalp hair, blood, and urine) of each night blindness and control human subjects were treated as described above for reference samples.

Microwave-Assisted Acid Digestion Method

A microwave-assisted digestion procedure was carried out in order to achieve a shorter digestion time. Duplicate samples of scalp hair (200 mg) and 0.5 mL of blood and urine samples of each night blindness and control individuals were directly placed into Teflon PFA flasks. Two milliliters of a freshly prepared mixture of concentrated HNO3–H2O2 (2:1, v/v) were added to each flask, left for 10 min. After this period, the flasks were placed in a covered PTFE container. This was then heated following a one-stage digestion program at 80% of total power (900 W), during 2–3 min for blood and urine and 4–5 min for hair samples. After the digestion, the flasks were left to cool, and the resulting solution was evaporated almost to dryness to remove excess acid, and then diluted to 10.0 ml in volumetric flasks with 0.1 M nitric acid. Blank extractions (without sample) were carried out performing the complete procedure of both methods.

To establish the validity described for our methodology, five replica samples of each certified sample (human hair, blood, and urine) were digested as reported above. All experiments were conducted at room temperature (30°C) following the well-established laboratory protocols. All digests obtained from both methods were analyzed for Cd, Pb, and Ni by electrothermal atomic absorption spectrophotometer. The concentrations were obtained directly from calibration graphs after correction of the absorbance for the signal from an appropriate reagent blank. The validity and efficiency of the microwave-assisted digestion method was checked with those obtained from conventional wet acid digestion method [26].

Analytical results of the certified samples, Clincheck control-lyophilized® human whole blood, Clincheck control-lyophilized® human urine, and Certified Human hair® BCR 397, certified reference materials, obtained by both digestion methods were close to that of the certified values, which confirmed the reliability of the methods. The percentage recovery of all elements in certified reference material samples obtained by conventional digestion method varied between 97.8% and 99.3%, but it was time consuming. The microwave-assisted digestion method was efficient and takes less than 10 min to complete the digestion of the three certified biological samples. Mean values for all the elements differed less than 1–2% from the certified values. The coefficient of variation differed less than 2% for the different elements. Non-significant differences were observed (p > 0.05) when comparing the values obtained by both methods (paired t test; Table 3).
Table 3

Determination of Cd, Pb, and Ni in certified samples by conventional (CDM) and microwave digestion method (MWD; N = 10)

Elements

Conventional digestion method (CDM)

Microwave digestion method (MWD)

T valuea

Percent recoveryb

Certified values

Certified sample of whole blood (μg/l)

 Cd

1.21 ± 0.07 (5.78)

1.187 ± 0.09 (7.58)

0.0912

98.10

1.2 ± 0.4

 Pb

105.67 ± 8.2 (7.76)

104.8 ± 7.3 (6.96)

0.0012

99.18

105 ± 24

 Ni

7.52 ± 0.62 (8.24)

7.39 ± 0.49 (6.63)

0.041

98.27

7.5 ± 1.8

Certified sample of urine (μg/l)

 Cd

11.83 ± 0.91 (7.69)

11.72 ± 0.85 (7.25)

0.44027

99.07

11.8 ± 2.5

 Pb

41.2 ± 1.96 (4.75)

40.77 ± 2.86 (7.01)

0.0038

98.9

41.0 ± 9.9

 Ni

11.88 ± 0.87 (7.32)

11.72 ± 0.69 (5.89)

0.0173

98.6

11.9 ± 3.0

Certified sample of human hair (μg/g)

 Cd

0.53 ± 0.025 (4.72)

0.524 ± 0.024 (4.58)

0.2256

98.87

0.52 ± 0.024

 Pb

33.29 ± 1.21 (3.63)

32.56 ± 1.18 (3.62)

0.096

97.8

33 ± 1.2

 Ni

46.07 ± 1.41 (3.06)

45.75 ± 1.38 (3.02)

0.9242

99.3

46.0 ± 1.4c

Values in parenthesis are RSD

aPaired t test between CDM and MWD; df = 9, T (critical) at 95% CL = 2.262, p < 0.50

bPercent recovery was calculated according to \( \frac{{\left[ {\text{MDM}} \right]}}{{\left[ {\hbox{CDM}} \right]}} \times {1}00 \)

cIndicative value

Results

The results reported in Table 4 show that the concentrations of trace toxic metals (Cd, Ni, and Pb) were altered in all three biological samples of night blindness children as related to controls. Trace mineral patterns in biological samples are providing fruitful data not only as diagnostic procedure but also in providing answers pertaining to treatment.
Table 4

Trace element concentrations in biological samples (scalp hair, blood, and urine samples) of normal and night blindness subjects

Age groups

Male, n = 186

Female, n = 174

Normal

Night blindness

p value

Normal

Night blindness

p value

Cadmium

 Scalp hair (μg/g)

  3–7

1.14 ± 0.37

2.78 ± 0.54

0.001

1.03 ± 0.25

2.56 ± 0.41

 

  Rangea

0.75–1.53

2.21–3.35

0.75–1.32

2.13–2.99

0.001

  8–12

1.83 ± 0.24

3.4 ± 0.6

0.001

1.63 ± 0.31

3.25 ± 0.46

0.001

  Rangea

1.52–2.11

2.75–4.05

1.30–1.97

2.87–3.72

 Blood (μg/l)

  3–7

2.56 ± 0.43

4.48 ± 0.32

0.001

2.47 ± 0.36

4.18 ± 0.25

0.001

  Rangea

2.11–3.04

4.04–4.84

2.09–2.86

3.81–4.47

  8–12

3.38 ± 0.36

5.67 ± 0.43

0.001

3.14 ± 0.19

5.45 ± 0.37

0.001

  Rangea

3.02–3.79

5.12–6.14

2.92–3.35

5.02–5.93

 Urine (μg/l)

  3–7

1.2 ± 0.32

2.45 ± 0.43

0.001

0.8 ± 0.35

2.28 ± 0.16

0.001

  Rangea

0.71–1.58

2.02–2.93

0.52–1.17

2.14–2.47

  8–12

1.42 ± 0.36

2.9 ± 0.53

0.001

1.3 ± 0.47

2.8 ± 0.41

0.001

  Rangea

1.02–1.83

2.24–3.45

0.82–1.79

2.37–3.25

Nickel

 Scalp hair (μg/g)

  3–7

4.3 ± 0.56

7.64 ± 1.62

0.001

4.24 ± 1.18

7.34 ± 1.36

0.001

  Rangea

3.67–4.93

6.00–9.26

3.01–5.50

5.94–8.79

  8–12

5.38 ± 1.21

8.4 ± 1.08

0.001

4.92 ± 1.34

8.1 ± 0.98

0.001

  Rangea

4.41–6.69

7.30–9.56

3.66–5.94

7.05–9.97

 Blood (μg/l)

  3–7

0.74 ± 0.24

2.35 ± 0.21

0.001

0.68 ± 0.19

2.21 ± 0.16

0.001

  Rangea

0.48–0.99

2.12–2.63

0.47–0.89

2.14–2.41

  8–12

1.14 ± 0.26

2.74 ± 0.49

0.001

1.03 ± 0.23

2.63 ± 0.54

0.001

  Rangea

0.86–1.42

2.29–3.15

0.75–1.29

2.11–3.19

 Urine (μg/l)

  3–7

1.35 ± 0.29

4.78 ± 0.39

0.001

1.18 ± 0.22

4.36 ± 0.26

0.001

  Rangea

0.92–1.66

4.36–5.19

0.94–1.42

4.05–4.63

  8–12

2.54 ± 0.42

5.4 ± 0.67

0.001

2.39 ± 0.41

5.12 ± 0.37

0.001

  Rangea

1.81–2.89

4.81–6.11

1.92–2.83

4.69–5.52

Lead

 Scalp hair (μg/g)

  3–7

4.3 ± 0.54

12.5 ± 1.3

0.001

4.2 ± 0.45

11.5 ± 0.37

0.001

  Rangea

3.68–4.87

11.1–13.9

3.69–4.69

10.7–11.9

  8–12

6.9 ± 1.5

16.9 ± 2.6

0.001

6.43 ± 1.36

15.6 ± 1.26

0.001

  Rangea

5.33–8.32

14.2–19.2

4.93–7.82

14.3–16.9

 Blood (μg/l)

  3–7

144 ± 20.2

276 ± 23.6

0.002

137 ± 20.8

265 ± 27.4

0.001

  Rangea

121–167

247–302

115–160

240–286

  8–12

165 ± 13.9

322 ± 36.2

0.003

149 ± 14.9

306 ± 28.2

0.001

  Rangea

149–181

293–359

132–165

274–335

 Urine (μg/l)

  3–7

43.5 ± 4.3

65.8 ± 3.64

0.001

41.2 ± 4.6

63.9 ± 5.6

0.001

  Rangea

38.6–49.2

61.5–69.2

36.8–46.5

57.9–69.8

  8–12

47.6 ± 6.4

73.2 ± 9.6

0.001

46.6 ± 5.2

71.6 ± 7.32

0.001

  Rangea

41.4–55.3

62.2–81.9

40.8–52.0

63.7–79.2

aMinimum–maximum values

The concentrations of Cd in the scalp hair samples of control children of both age groups, 3–7 and 8–12 years, were found in the range of 0.75–1.53 and 1.52–2.11 μg/g, while the levels of zinc in male night blindness children of both age groups were in the range of 2.21–3.35 and 2.75–4.05 μg/g; the same trend was observed in females. Range of Cd levels in blood samples of referent subjects of both genders (2.11–3.79 mg/l and 2.09–3.35 μg/l) was significantly lower as compared to the range of Cd values observed in blood samples of night blindness subjects (4.04–6.14 and 3.81–5.93 μg/l) in male and females, respectively. The excretion of Cd was high in night blindness patients of both genders.

The elevated level of Ni was observed in scalp hair of night blindness children of both genders (Table 4). The level of Ni in blood samples of night blindness children is lower (2.12–3.15 and 2.14–3.19 μg/l) as compared to values of Ni found in blood samples of referents subjects (0.48–1.42 and 0.47–1.29 μg/l) of male and females, respectively, of both age groups. The excretion of Ni was high in night blindness children of both genders.

The Pb levels in hair revealed significant difference (p < 0.001) between controls and patients; levels for healthy male and female of both age groups were found in the range of 3.68–8.32 and 3.69–7.82 μg/g, while range for patients was observed as 11.1–19.2 and 10.7–16.9 μg/g for both male and female, respectively. In blood samples, the range of Pb values in healthy subjects of both age groups was found to be lower (121–181 mg/l and 115–165 μg/l) than those values obtained for Pb in blood samples of night blindness children (247–359 and 240–335 μg/l) for male and females, respectively (Table 4). The excretion of Ni was high in night blindness children of both genders.

Discussion

Minerals play an important role in the subtle biochemistry of the body as do vitamins. Virtually all enzymatic reactions in the body require minerals as cofactors. Metal ions are integral functional components of many enzymes and transcriptional regulatory proteins.

Night blindness is considered as an early sign of vitamin A deficiency. Low intake of fruit and vegetables containing beta-carotene, which the body converts into vitamin A, may also contribute to a vitamin A deficiency. The relationship of micronutrient deficiency with the risk of developing certain major visual disorders and the therapeutic role of these micronutrients are now being recognized. Night blindness may be an early sign of vitamin A deficiency. Such a deficiency may result from diets low in animal foods, the main source of vitamin A, zinc, iron, and copper [27].

Analyses of Ni, Pb, and Cd in scalp hair, blood, and urine samples of both age groups of control and night blindness persons indicate significant difference (Table 4). The children with a history of night blindness had a significantly higher concentration of Cd, Pb, and Ni in scalp hair, blood, and urine samples than the normal persons of both age groups (p = 0.001–0.007). Our results are also consistent with other studies, which have shown that significant levels of Cd accumulate in the blood and lenses of macular degeneration patients compared with controls.

Pb, Cd, and Ni were found in all of the pigmented ocular tissues (e.g., retinal pigment epithelium/choroid, and ciliary body) that we studied. The importance of melanin binding of heavy metals in pigmented ocular tissues is unclear. Melanin may confer tissue protection by acting as a filter or detoxicant for heavy metals from the adjacent neural retina and photoreceptor cells [28, 29]. Similar to the retinal pigment epithelium, the choroid plexus of the brain sequesters Pb, acting as a defensive barrier to prevent entry of toxic elements into the brain [9]. Conversely, melanin binding of heavy metals throughout the life of an individual produces a local reservoir of potentially toxic elements that ultimately could reach a concentration that is destructive to the retinal pigment epithelium and adjacent retina.

Low-level Pb exposure produces scotopic vision deterioration and rod and bipolar apoptotic cell death [30, 31]. In rabbits, Pb poisoning causes swelling of the retinal pigment epithelium [32], which leads to degeneration of the photoreceptors [33]. Visual loss in humans after systemic Pb poisoning is usually related to encephalopathy and optic neuropathy, because of the toxic effect of Pb on the brain and optic nerve. Recent evidence, however, indicates that Pb, Cd, and Ni can exert oxidative stress by producing reactive oxygen species that result in lipid peroxidation, DNA damage, and depletion of cell antioxidant defense systems [29]. Oxidative stress and free radical damage is thought to play a significant role in age-related macular degeneration [34].

The pigmented tissues of the eye, such as the retinal pigment epithelium, choroid, iris, and ciliary body, have a high affinity for metal ions [35]. Melanin within the pigment granules binds metal ions [36]. Metal ions are bound by melanosomes according to atomic weight and volume (e.g., the percentage binding of calcium 30%, zinc 37%, Pb 62%, iron 65%, and mercury 72%). Metals such as zinc, copper, calcium, manganese, molybdenum, and iron are found in ocular melanosomes, particularly within the retinal pigment epithelium [37, 38]. Heavy metals can effectively compete for the same binding sites as other metal ions [39] and have the capacity to replace previously bound metals and alter ocular metal concentrations [40, 41]. Once bound, heavy metals are not easily amenable to displacement [42].

Conclusion

It was also concluded that levels of toxic elements were higher in children having ocular problems. Among other things, regular monitoring of metals in biological/clinical specimens therefore plays an important part in, among other things, identifying possible sources of contamination/intoxication, as well as in preventing, through early detection, the onset of metal-related illness in individuals. Children are particularly susceptible to essential elemental deficiency due to their increased mineral needs for rapid growth and the relatively low iron and zinc content of their diets due to poverty. It is necessary to add these minerals via food supplements. The results of this study provided guidance to clinicians and other professionals investigating deficiency of essential trace metals and excessive levels of toxic metals in biological samples of healthy and anemic children.

Notes

Acknowledgement

The authors would like to thank the Higher Education Commission, Islamabad, Pakistan, for sponsoring this project.

References

  1. 1.
    Christian P, West KP, Khatry SK (2001) Maternal night blindness increases risk of mortality in the first 6 months of life among infants in Nepal. J Nutr 131:1510–1512PubMedGoogle Scholar
  2. 2.
    Brody T (1999) Nutritional Biochemistry, 2nd edn. Academic, San DiegoGoogle Scholar
  3. 3.
    Fox DA, Campbell ML, Blocker YS (1997) Functional alterations and apoptotic cell death in the retina following developmen-tal or adult lead exposure. Neurotoxicology 18:645–664PubMedGoogle Scholar
  4. 4.
    Bressler J, Kim KA, Chakraborti C, Goldstein G (1999) Mechanism of lead neurotoxicity. Neurochem Res 24:595–600PubMedCrossRefGoogle Scholar
  5. 5.
    Humphreys DJ (1991) Effects of exposure to excessive quantities of lead on animals. Br Vet J 147:18–30PubMedGoogle Scholar
  6. 6.
    Khalil-Manesh F, Gonik HC, Weiler EJ et al (1993) Lead-induced hypertension: possible role of endothelial factors. Am J Hypertens 6:723–729PubMedGoogle Scholar
  7. 7.
    Eichenbaum JW, Zheng W (2000) Distribution of lead and transthyretin in human eyes. Clin Toxicol 38:377–381CrossRefGoogle Scholar
  8. 8.
    Hu H, Rabinowitz M, Smith D (1998) Bone lead as a biological marker in epidmiologic studies of chronic toxicity: conceptual paradigms. Env Health Persp 105:1–8CrossRefGoogle Scholar
  9. 9.
    Cavallaro T, Martone RL, Dwork AJ et al (1990) The retinal pigment epithelium is the unique site of transthyretin synthesis in the rat eye. Invest Ophthal Vis Sci 31:497–501PubMedGoogle Scholar
  10. 10.
    Zheng W, Shen H, Blaner WS et al (1996) Chronic lead exposure alters transthyretin concentration in rat cerebrospinal fluid: the role of the choroid plexus. Toxicol Appl Pharmacol 139:445–450PubMedCrossRefGoogle Scholar
  11. 11.
    Satarug S, Baker JR, Urbenjapol S et al (2003) A global perspective on cadmium pollution and toxicity in non-occupationally exposed population. Toxicol Lett 137:65–83PubMedCrossRefGoogle Scholar
  12. 12.
    Zheng W, Blaner WS, Zhao Q (1999) Inhibition by Pb of production and secretion of transthyretin in the choroid plexus: its relationship to thyroxine transport at the blood-CSF barrier. Toxicol Appl Pharmacol 155:24–31PubMedCrossRefGoogle Scholar
  13. 13.
    Stillman MJ, Presta A (2000) Characterizing metal ion interactions with biological molecules—the spectroscopy of metallothionein. In: Zalups RZ, Koropatnick J (eds) Molecular biology and toxicology of metals. Taylor & Francis, New York, pp 276–299Google Scholar
  14. 14.
    Ulshafer RJ, Allen CB, Rubin ML (1990) Distributions of elements in the human retinal pigment epithelium. Arch Ophthalmol 108:113–117PubMedGoogle Scholar
  15. 15.
    Afridi HI, Kazi TG, Kazi GH et al (2006) Essential trace and toxic element distribution in the scalp hair of Pakistani myocardial infarction patients and controls. Biol Trace Elem Res 113:19–34PubMedCrossRefGoogle Scholar
  16. 16.
    Polkowska Z, Kozlowska K, Namiesnik J, Przyjazny A (2004) Biological fluids as a source of information on the exposure of man to environmental chemical agents. Crit Rev Anal Chem 34(2):105–119CrossRefGoogle Scholar
  17. 17.
    Rodushkin I, Odman OF, Olofsson R, Axelsson MD (2000) Determination of 60 elements in whole blood by sector field inductively coupled plasma mass spectrometry. J Anal At Spectrom 15(8):937–944CrossRefGoogle Scholar
  18. 18.
    De Castro Maciel CJ, Miranda GM, De Oliveira DP et al (2003) Determination of cadmium in human urine by electrothermal atomic absorption spectrometry. Anal Chim Acta 491(2):231–237CrossRefGoogle Scholar
  19. 19.
    Khalique A, Ahmad S, Anjum T et al (2005) A comparative study based on gender and age dependence of selected metals in scalp hair. Environ Monit Assess 104(1–3):45–57PubMedCrossRefGoogle Scholar
  20. 20.
    Senofonte O, Violante N, Caroli S (2000) Assessment of reference values for elements in human hair of urban schoolboys. J TEs Med Biology 14(1):6–13Google Scholar
  21. 21.
    Kazi TG, Arain MB, Baig JA et al (2009) The correlation of arsenic levels in drinking water with the biological samples of skin disorders. Sci Total Environ 407:1019–1026PubMedGoogle Scholar
  22. 22.
    Kazi TG, Jalbani N, Kazi N et al (2009) Estimation of toxic metals in scalp hair samples of chronic kidney patient. Biol Trace Elem Res 125(3):16–27CrossRefGoogle Scholar
  23. 23.
    Wright RO, Amarasiriwardena C, Woolf AD et al (2006) Neuropsychological correlates of hair arsenic, manganese, and cadmium levels in school-age children residing near a hazardous waste site. Neurotoxicology 27(2):210–216PubMedCrossRefGoogle Scholar
  24. 24.
    Kimble MS (1939) The photoelectric determination of vitamin A and carotene in human plasma. J Lab Clin Med 24:1055Google Scholar
  25. 25.
    Afridi HI, Kazi TG, Kazi GH (2006) Analysis of heavy metals in scalp hair samples of hypertensive patients by conventional and microwave digestion methods. Spectrosc Lett 39:203–214CrossRefGoogle Scholar
  26. 26.
    Kazi TG, Afridi HI, Kazi GH, Jamali MK, Arain MB, Jalbani N (2006) Evaluation of essential and toxic metals by ultrasound-assisted acid leaching from scalp hair samples of children with macular degeneration patients. Clin Chim Acta 369(1):52–60PubMedCrossRefGoogle Scholar
  27. 27.
    VandenLangenberg GM (1998) Associations between antioxidant and zinc intake and the 5-year incidence of early age-related maculopathy in the Beaver Dam eye study. Am J Epidemiol 148(2):204–214PubMedGoogle Scholar
  28. 28.
    Yiin SJ, Chern CL, She JY et al (1999) Cadmium induced renal lipid peroxidation in rats and protection by selenium. J Toxicol Environ Health A 57:403–413PubMedCrossRefGoogle Scholar
  29. 29.
    Bhattacharyya MH, Wilson AK, Ragan SS, Jonch M (2000) Biochemical pathways in cadmium toxicity. In: Zalups RZ, Koropatnick J (eds) Molecular biology and toxicology of metals. Taylor & Francis, New York, pp 276–299Google Scholar
  30. 30.
    Fox DA, Sillman AJ (1979) Heavy metals affect rods, but not cone photoreceptors. Science 206:78–80PubMedCrossRefGoogle Scholar
  31. 31.
    Bushnell PJ, Bowman RE (1977) Scotopic vision deficits in young monkeys exposed to lead. Science 196:333–335PubMedCrossRefGoogle Scholar
  32. 32.
    Brown DVL (1974) Reactions of the rabbit retinal pigment epithelium to systemic lead poisoning. Trans Am Ophthamol Soc 72:404–447Google Scholar
  33. 33.
    Hughes WF, Coogan P (1974) Pathology of the retinal pigment epithelium and retina in rabbits poisoned with lead. Am J Pathol 77:237–254PubMedGoogle Scholar
  34. 34.
    Beatty S, Koh H, Phil M et al (2000) The role of oxidative stress in the pathogenesis of age-related macular degeneration. Surv Ophthalmol 45:115–134PubMedCrossRefGoogle Scholar
  35. 35.
    Potts AM, Au PC (1976) The affinity of melanin for inorganic ions. Exp Eye Res 22:487–491PubMedCrossRefGoogle Scholar
  36. 36.
    Larrson BS (1993) Interaction between chemicals and melanin. Pigment Cell Res 6:127–133CrossRefGoogle Scholar
  37. 37.
    Panessa BJ, Zadunaisky JA (1981) Pigment granules: a calcium reservoir in the vertebrate eye. Exp Eye Res 32:593–604PubMedCrossRefGoogle Scholar
  38. 38.
    Samuelson DA, Smith P, Ulshafer FJ et al (1993) X-ray microanalysis of ocular melanin in pigs maintained in normal and low zinc diets. Exp Eye Res 56:63–70PubMedCrossRefGoogle Scholar
  39. 39.
    Drager UC, Balkema GW (1987) Does melanin do more than protect from light? Neurosci Res Suppl 6:575–586Google Scholar
  40. 40.
    Sarna T, Hyde JS, Swartz HM (1976) Ion exchange in melanin, an electron spin resonance study with lanthanide probes. Science 192:1132–1134PubMedCrossRefGoogle Scholar
  41. 41.
    Jamall IS, Roque H (1989–1990) Cadmium-induced alterations of ocular trace elements. Influence of dietary selenium and copper. Biol Trace Elem Res 23:55–63CrossRefGoogle Scholar
  42. 42.
    Sarna T, Froncisz W, Hyde JC (1980) Cu2 probe of metal-ion binding sites in melanin using electron paramagnetic resonance spectroscopy. II. Natural melanin. Arch Biochem Biophys 202:304–313PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2010

Authors and Affiliations

  • Hassan Imran Afridi
    • 1
    • 2
  • Tasneem Gul Kazi
    • 1
  • Naveed Kazi
    • 3
  • Sirajuddin
    • 1
  • Ghulam Abbas Kandhro
    • 1
  • Jameel Ahmed Baig
    • 1
  • Abdul Qadir Shah
    • 1
  • Sham Kumar Wadhwa
    • 1
  • Sumaira Khan
    • 1
  • Nida Fatima Kolachi
    • 1
  • Faheem Shah
    • 1
  • Mohammad Khan Jamali
    • 1
  • Mohammad Balal Arain
    • 1
  1. 1.National Center of Excellence in Analytical ChemistryUniversity of SindhJamshoroPakistan
  2. 2.Mechanical and Manufacturing EngineeringDublin City UniversityDublinIreland
  3. 3.Liaquat University of Medical and Health SciencesJamshoroPakistan

Personalised recommendations