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Determination of Heavy Metals in Urine of Patients and Tissue of Corpses by Atomic Absorption Spectroscopy

  • A. O. OgunfowokanEmail author
  • A. S. AdekunleEmail author
  • B. A. Oyebode
  • J. A. O. Oyekunle
  • A. O. Komolafe
  • G. O. Omoniyi-Esan
Original Article
  • 59 Downloads

Abstract

This study determined the concentrations of some heavy metals (Cd, Pb, Mn, Cu and Zn) in urine samples of patients with kidney, liver and lung related diseases (age 15–70 years); and tissue samples (kidney, liver and lung) that were pathologically abnormal from corpses (age 21–50 years) during autopsies at the Obafemi Awolowo University Teaching Hospitals Complex, Ile-Ife, Nigeria. It also determined the effects of age distribution, gender and life styles on the urine samples of patients with the aforementioned diseases. A total of 35 urine samples of the patients were analyzed out of which 15 had kidney related diseases, 10 had liver related diseases and 10 had lung related diseases. Four urine samples were used as controls. The urine samples were collected before meal and the age, sex, occupation and personal habit of patients were considered when taking samples. The samples were digested using micro-wave assisted digestion method and analyzed with Atomic Absorption Spectrophotometer. The results showed that the concentrations of Cd (0.052–0.093 µg mL−1), Pb (0.150–0.376 µg mL−1), Mn (0.014–0.278 µg mL−1), Cu (0.738–2.475 µg mL−1) and Zn (0.476–0.975 µg mL−1) in urine samples (male and female) were higher than those of the control samples (Cd: 0.035 µg mL−1, Pb: 0.253 µg mL−1, Mn: 0.045 µg mL−1, Cu: 0.040 µg mL−1 and Zn: 0.716 µg mL−1) and also higher than the standard human urine levels of metals recommended by World Health Organisation. Also, Mn had the highest concentrations of all the metals determined in kidney, liver and lung tissues analysed. The study concluded that the high concentrations of heavy metals obtained confirmed the associated health complications noticed in the patients.

Keywords

Heavy metals Urine Tissues Corpses Atomic absorption spectroscopy Lung diseases 

1 Introduction

The continuous rise in the level of environmental pollutions by toxic heavy metals from the manufacturing industries, continuous use of pesticides, herbicides and inorganic fertilisers in modern farming activities, industrial wastes discharge, use of leaded gasoline, mining activities and many more calls for great concern, due to its effects on human health [1]. Heavy metals are used widely in industries for various production purposes, and living organisms require certain reduced amounts for metabolic activities. On the other hand, high levels can be harmful to the organisms causing nervous dysfunction, lower energy levels, and damage to blood composition, lungs and kidneys. In terms of the possible antagonistic effects on human health, lead, cadmium, and arsenic have caused most concern [2], because they are readily transported via food-chains and are not known to serve any vital biological function [3]. Additionally, Cr and its compounds can enter the human body via direct contact and ingestion and can cause great harm to the respiratory and digestive systems [4]. The distribution of heavy metals is greatly influenced by activities of organisms, climate, topography, parent’s materials and time [5].

Human exposure to heavy metals especially in the developing countries has continuously been on the increase. This is due to the increase in the domestic and industrial activities of man. Generally, human beings are exposed to these metals by ingestion, that is; eating and drinking, inhalation through breathing and dermal routes [6, 7]. Working in or living around an industrial site which uses these heavy metals and their compounds increases the risk of exposure routes. Lifestyles such as smoking, hunting and gathering activities also contribute to human exposure to heavy metals [7, 8]. Once the heavy metals are in the body, they are absorbed and distributed into the tissues and organs. The amount that is actually absorbed from the digestive tract can vary widely, depending on the chemical form of the metal and the age and nutritional status of the individual. Excretion primarily occurs through the kidneys and digestive tract, but heavy metals somehow persist in some storage sites, like the liver, bones, and kidneys, for many years.

Heavy metals get into the environment through natural and anthropogenic means (human activities). These include natural weathering of the earth’s crust, soil erosion, mining, urban runoff, industrial discharge, sewage effluents, application of pesticides on crop, air pollution fallout, and many others [9]. The contamination chain of heavy metals is always in form of a cyclic order: industry, atmosphere, soil, water, foods and human. Heavy metals accumulate in the body system and disrupt function in vital organs and glands such as the heart, brain, kidneys, bone, liver, lung and also displace the vital nutritional minerals from their original place, thereby, affecting their biological function. It is, however, not possible to live in an environment totally free of heavy metals.

Workers such as Mahugija et al. have reported high contamination levels of heavy metals in urine samples of school children from selected industrial and non-industrial areas in Tanzania [10]. Similarly, Sá et al. investigated the presence and distribution of heavy metals in samples of renal cell carcinoma, as well as adjacent renal tissue (control samples) in patients submitted to radical or partial nephrectomy for renal cell carcinoma [11]. Sani and Abdullahi determined heavy metals concentration in body fluids (blood and urine) of metal workers in Kano metropolis, Nigeria to monitor occupational activities and concluded that the workers are at health risks due to the high concentration of the metals [12]. Other recent reports on levels of heavy metals in urine, blood and water samples have also been documented [13, 14, 15]. However, heavy metals determination in human kidney, liver and lungs are very scarce as compared to that of animal’s specimen. For example, there were reports on heavy metals in: the livers and kidneys of slaughtered cattle, sheep and goats [16, 17]; liver, kidney and meat of beef (cow), mutton (sheep), caprine (goat) and chicken [18]; blood and various organs (lung, liver, kidney and cerebral cortex) of rats [19]; muscle, liver, and spleen tissues of a large commercially valuable catfish species from Brazil [20]; and a one-time high dose oral administration of heavy metals mixture (HMMs) induced systemic toxicity in rats [21] have also been reported. However, few cases of studies of heavy metals in human kidney [22]; 10 autopsied human organs (liver, kidney, cerebrum, heart, spleen, lung, bone, blood, hair and nail) of Koreans [23] and the liver tissues of autopsy cases in Ankara, Turkey [24] have been investigated.

Different analytical techniques have been employed for heavy metals determination in biological samples and these include Atomic Absorption Spectrophotometry (AAS) [13, 16, 17, 18, 20, 24] and Inductively coupled plasma atomic emission spectrometry (ICP-AES) [19, 23]. Atomic absorption spectroscopy (AAS) is an analytical technique widely used for elemental determination because of its simplicity, sensitivity, low limit of detection, cost effectiveness and ability to determine over 70 elements in solution and in different matrices including biological fluids, water, air particulates and pharmaceutical products to mention a few. Apart from the free access to AAS, other characteristic and advantages of the technique had led to the choice of the equipment for heavy metals analysis in urine, kidney, liver and lung tissues in the present study. Also, microwave assisted digestion (MAD) was employed for sample digestion because it is cost effective, provides high pressure/temperature aggressive digestion of samples with excellent recovery for multi-elemental analysis, at much lesser time compared to the hot plate digestion method.

There has been increasing concern, mainly in the developed world, about exposures, intakes and absorption of heavy metals by humans. Due to increase in population, there is a need for a cleaner environment and reduction in the amount of contaminants reaching people as a result of anthropogenic activities. The practical implication of this trend in the developed world has been the imposition of new and more restrictive regulations. Unfortunately, this is not the case in the developing world. In Nigeria, for example, there are paucity of data and no enough statistical data to show the levels of these heavy metals present in diseased human organs, hence, this study. Therefore, the study determined the levels of some heavy metals (Pb, Cd, Mn, Zn, and Cu) in urine samples of patients diagnosed with kidney, liver and lung diseases; and lung, liver, kidney tissues of human corpses. It also explored the effect of age distribution, lifestyles (e.g. occupation, eating and drinking habits, smoking and drug usage) and gender on the levels of heavy metals in the urine samples of patients living with liver, lung and kidney related diseases. It is expected that results obtained from the study will provide data on human exposure to heavy metals, how these metals are distributed in tissues and organs of Nigerians, plausible causes of the high concentrations in human body (if found higher) and ways of reducing human exposure to the metals.

2 Materials and Methods

2.1 Reagents Used

All reagents used for this work were of analytical grade and they include: nitric acid (HNO3), hydrogen peroxide (H2O2), lead nitrate, Pb(NO3)2, copper sulphate (CuSO4·5H2O) and cadmium acetate (Cd(CH3COO)2). Solutions were prepared using doubly-distilled deionised water.

2.2 Sample Collection

Ethical clearance certificate was obtained from the Ethics and Research Committee of the Obafemi Awolowo Teaching Hospitals Complex, Ile-Ife. Prior to sample collection. Informed consents were obtained from the patients or their relations by signing the informed consent forms. The samples were collected from May 2016 to February 2017 (10 months). Midstream early morning urine samples were collected in sterilized polyethylene sample bottles from patients diagnosed with various liver, lung and kidney related diseases. The samples were collected before meal. The age range of the patients were between 15 and 70 years of age. The urine collections were supervised by health professionals. Age, sex, occupation and personal habit of patients were also considered when taking samples. A total of 35 patients consented and their urine samples were collected out of which 15 (8 males and 7 females) had kidney related cases, 10 (6 males and 4 females) had lung related cases and 10 (6 males and 4 females) had liver related cases. Four urine samples were collected from volunteered individuals (2 males and 2 female) with no known ailment as control for this research.

Tissue samples (kidney, liver and lungs) were also collected at Post-Mortem Investigation Unit of the Department of Morbid Anatomy and Forensic Medicine, Obafemi Awolowo University Teaching Hospitals Complex, Ile-Ife. A total of 18 pathologically abnormal tissue samples from corpses with varying diseases of kidney, liver and lungs were collected, from which 6 (2 males and 4 females) samples had diseased kidney; 6 (2 males and 4 females) had diseased liver and 6 (1 male and 5 females) had diseased lung conditions. The ages ranged from 21 to 50 years. The autopsies were performed few hours after death. The liver samples were taken from the superior anterior surface of the right lobe, the kidney cortex samples were obtained from the lower pole and the lung samples were taken from the lower part by trained medical pathologists. All the samples were taken to the laboratory and stored in the refrigerator at 4 °C before analysis.

2.3 Sample Preparation

Urine samples were digested using the procedure described by Memon et al. [25] while the tissue samples were digested using the method described by Cerulli et al. [26]. All samples were done in duplicates. The urine samples were allowed to be thawed thoroughly at room temperature. 4 mL freshly prepared HNO3/H2O2 (2:1 v/v) was added to 1 mL of each urine sample and left for 10 min as a pre- digestion time, then placed in a microwave oven. The samples were then heated following a one-stage digestion, programmed at 80% of the total power in a Samsung domestic microwave oven with maximum heating capacity of 950 W for 2–3 min. The digestion flasks were then allowed to cool. For the tissue samples, 0.5 g of each tissue sample was treated overnight in 5 mL conc. HNO3 and then subjected to Microwave Assisted Digestion (MAD). Two (2) mL H2O2 was then added and further digested to obtain clear solution, evaporated to near dryness to remove excess acid. The digested solutions (urine and tissue) were evaporated to semi-dried mass to remove excess acid and diluted with 0.5 mL of 0.1 M HNO3. Each sample solution was transferred to a 25 mL flask and diluted to mark with doubly-distilled water and finally transferred into a sterilized polyethylene bottle for instrumental analysis. The digested samples were analysed for Cu, Pb, Cd, Mn and Zn using PG990 Atomic Absorption Spectrophotometer (AAS) at the Centre for Energy Research and Development (CERD), Obafemi Awolowo University, Ile-Ife (Scheme 1).
Scheme 1

Samples preparation procedures for urine and the tissues (kidney, liver and lung) samples

2.4 Quality Control Measure

2.4.1 Blank Determination

Blank determination was carried out following the same procedure described above for urine and tissue samples. The presence of 0.5 mL of 0.1 M HNO3 in the final solution was necessary to maintain acidic environment and avoid formation of insoluble hydroxides before analysis. The digested blank samples were analysed for Cu, Pb, Cd, Mn and Zn.

2.4.2 Recovery Analysis

The recovery experiment was done by adding 4 mL of freshly prepared mixture of nitric acid and hydrogen peroxide in ratio 2:1 vol./vol. to 1 mL of urine sample. The solutions were then spiked with 5 mL of 10 ppm of Cu2+, Cd2+ and Pb2+ ions respectively. Also, 5 mL of concentrated nitric acid was added to 0.5 g of lung and 0.5 g kidney tissues in 2 separate Teflon beakers and were allowed to dissolve overnight. The dissolved samples were spiked with 5 mL of 10 ppm Pb2+ and Cd2+ ions and digested using the method described by Memon et al. [25]. The digested spiked samples were made up to the mark in a 25 mL volumetric flask with doubly-distilled water. The digested samples were stored in a polyethylene sample bottles at 4 °C prior to AAS analysis. Unspiked samples were also digested and analysed in order to calculate the percentage recovery using the formula below [27].
$$ \% \;{\text{Recovery}} = \frac{{{\text{C}}_{1} - {\text{C}}_{2} }}{\text{C}} \times 100, $$
where, C1 is the concentration of metal in spiked sample, C2 is the concentration of metal in unspiked sample, C is the concentration of metal in standard used for spiking

3 Results and Discussion

3.1 Recovery Analysis

The results of the recovery analysis for Cu, Pb and Cd in the urine and tissue samples are presented in Table 1. The recoveries of the metals ranged from 72 ± 3.8% to 92 ± 5.3%. The method attained good precisions with relatively low standard deviation. The percentage recoveries showed that the uncertainties in the methods of analyses used were within acceptable limit and the procedure adopted was effective.
Table 1

Percentage recovery of heavy metals in tissue and urine samples

Sample

Metals

Percentage recovered

Tissue

Pb2+

75.0 ± 3.8a

 

Cd2+

72.0 ± 2.5

Urine

Cu2+

92.0 ± 5.3

 

Cd2+

89.0 ± 6.2

 

Pb2+

86.0 ± 3.4

aStandard error of triplicate analysis

3.2 Concentration (µg mL−1) of Heavy Metals in Urine Samples

Table 2 presents the heavy metals concentration in the urine control samples for both male and female patients. The concentrations of Pb (0.253 ± 0.020 µg mL−1) and Zn (0.550 ± 0.051 µg mL−1) were higher compared with other metals analysed in male control urine samples. A similar trend was observed for the female control urine sample where higher concentrations of Pb (0.235 ± 0.017 µg mL−1) and Zn (0.716 ± 0.035 µg mL−1) were recorded. However, the concentrations of Cd (Male: 0.035 ± 0.012 µg mL−1, Female: 0.026 ± 0.018) and Pb from this study were lower compare with (Cd: 0.38 mg L−1 and Pb: 0.03 mg L−1) in urine control samples (male and female) studied in Maiduguri, Nigeria [13].
Table 2

Concentration (µg mL−1) of heavy metals in urine control samples

Age

Cd

Pb

Mn

Cu

Zn

Male

     

 33

0.050 ± 0.021a

0.200 ± 0.013

0.030 ± 0.014

0.060 ± 0.012

0.520 ± 0.076

 45

0.020 ± 0.003

0.306 ± 0.026

0.050 ± 0.013

0.020 ± 0.014

0.580 ± 0.025

 Mean ± SD

0.035 ± 0.012 

0.253 ± 0.020 

0.040 ± 0.014

0.040 ± 0.013

0.550 ± 0.051

Female

 42

0.021 ± 0.011a

0.200 ± 0.014

0.060 ± 0.026

0.020 ± 0.013

0.717 ± 0.017

 56

0.031 ± 0.025

0.270 ± 0.020

0.030 ± 0.042

0.030 ± 0.014

0.715 ± 0.053

 Mean ± SD

0.026 ± 0.018

0.235 ± 0.017

 0.045 ± 0.034

0.025 ± 0.014

0.716 ± 0.035

aStandard error of triplicate analysis

The mean concentrations of heavy metals (µg mL−1) for urine samples of male and female patients with respect to their ages are presented in Figs. 1 and 2. It was observed that Cd has the lowest concentration which ranged from 0.062 ± 0.022 to 0.084 ± 0.043 µg mL−1 for male patients and 0.052 ± 0.033 to 0.093 ± 0.041 µg mL−1 for female patients. Copper has highest concentrations ranging from 0.738 ± 0.044 to 2.475 ± 0.073 µg mL−1 and 0.738 ± 0.076 to 1.613 ± 0.151 µg mL−1 for male and female patients respectively.
Fig. 1

Concentrations (µg mL−1) of heavy metals in urine samples of male patients with respect to their ages (Yr.)

Fig. 2

Concentration (µg mL−1) of heavy metals in urine samples of female patients with respect to their ages (Yr.)

Generally, the concentrations of metals recorded for urine samples of diseased patients (male and female) are far higher than those of the control samples. It was also observed in this study that the highest mean concentrations of Cd (0.084 ± 0.043 µg mL−1), Pb (0.375 ± 0.273 µg mL−1), Mn (0.278 ± 0.111 µg mL−1) and Cu (2.475 ± 0.073 µg mL−1) were observed in older ages (55-64 years) in male urine samples, while Zn has uniform distributions across the ages. However, reverse trend in the metal concentrations was observed for the female urine samples (Fig. 2). The mean metal concentration, Cu (1.613 ± 0.151 µg mL−1) and Zn (0.975 ± 0.043 µg mL−1) were more in the lower age grade. The concentrations of Cu in the urine samples for both sexes were higher than those of Pb, Cd, Mn and Zn and the maximum recommended levels of 0.02 µg mL−1 for Cu in human urine reported by WHO [28]. The higher values of Cu might be due to personal habits of the patients (food consumption, drinking contaminated water with Cu, intake of alcohol, swimming in pools, eating vegetarian diet, and high copper food such as nuts, seeds and avocado) [29]. Thus, the effects of high concentration of copper might lead to different health issues such as brain damages i.e. effect on nervous system, liver disease, memory loss, depression, anxiety, attention deficit, hyperactivity, autism, bipolar disorder, Wilson’s disease, and many more [29]. The concentrations of Cd in urine samples of patients already diagnosed with liver, lungs and kidney related diseases were higher than Cd levels in control samples. Also, Cd levels for both male and female urine samples were above the standard recommended level of 0.000185 µg mL−1 in human urine reported by ATSDR [29].

Pb concentrations (0.222 ± 0.121 to 0.438 ± 0.034 µg mL−1) are also above the control samples (0.235 ± 0.017 to 0.253 ± 0.020 µg mL−1) and far above the maximum required limit for Pb in human urine (0.000677 µg mL−1) [30]. Humans are exposed to Pb through inhalation of contaminated air (fumes from leaded gasoline), water, smoking and food except occupational exposed individuals [31]. Toxic effects of Pb in adults include renal disease, memory loss, abdominal pain, anemia [32, 33]. Therefore, fifteen patients from this study with renal related diseases might be as a result of the high level of Pb in their body.

ATSDR [29] gave the maximum required limit for Mn in human urine level to be 0.00119 µg mL−1, which is below the levels obtained in this study for both male (0.014 ± 0.008 to 0.278 ± 0.111 µg mL−1) and female (0.112 ± 0.041 to 0.250 ± 0.075 µg mL−1) patients urine samples across all age groups. The primary source of Mn exposure for the general populace is through food intake. The highest concentrations of Mn are found in grains, nuts, legumes, and fruit. Mn has potential to bio-concentrate, particularly in lower organisms of the food chain. Exposure to high levels of Mn dust can result in lung inflammation and impaired lung function [29].

Table 3 presents the t test while Table 4a, b showed the paired correlation statistical interpretation data obtained for the metal concentrations in male and female patients urine samples. The t-test (Table 3) gave values of: Cd (3.888), Pb (3.774), Mn (2.933), Cu (3.281) and Zn (3.800) for male samples and Cd (5.474), Pb (4.879), Mn (5.098), Cu (5.048) and Zn (5.193) for the female patients respectively. These values are higher than the t- critical value (2.20) at p ≤ 0.05 confidence level. This suggests that there is significant difference in the concentrations of heavy metals between male and female patients. The correlation factors (Table 4) show that there are positive correlations among the levels of metals present in both male and female patients.
Table 3

T-test analysis of heavy metals in urine samples

Metals

Test value = 0

T

Df

Sig. (2-tailed)

Mean difference

95% confidence interval of the difference

Lower

Higher

Male patients

 Cd

3.888

11

0.003

0.041917

0.01819

0.06565

 Pb

3.774

11

0.003

0.165833

0.06913

0.26253

 Mn

2.933

11

0.014

0.090417

0.02256

0.15827

 Cu

3.281

11

0.007

0.760750

0.25049

1.27101

 Zn

3.800

11

0.003

0.409417

0.17230

0.64653

Female patients

 Cd

5.474

11

0.000

0.056500

0.03378

0.07922

 Pb

4.879

11

0.000

0.212417

0.11660

0.30823

 Mn

5.098

11

0.000

0.130417

0.07411

0.18672

 Cu

5.048

11

0.000

0.902167

0.50882

1.29551

 Zn

5.193

11

0.000

0.535917

0.30877

0.76307

t-critical at 95% confidence level P ≤ 0.05 = 2.20

Table 4

(a) Correlation values of heavy metals in urine samples of male patients; (b) correlation table of heavy metals in urine samples of female patients

Control variables

Cd

Pb

Mn

Cu

Zn

(a)

     

 Cd

     

  Correlation

1.000

0.963

0.736

0.906

0.974

  Significance (2-tailed)

0.000

0.010

0.000

0.000

  Df

0

9

9

9

9

 Pb

     

  Correlation

0.963

1.000

0.859

0.845

0.967

  Significance (2-tailed)

0.000

0.001

0.001

0.000

  Df

9

0

9

9

9

 Mn

     

  Correlation

0.736

0.859

1.000

0.522

0.841

  Significance (2-tailed)

0.010

0.001

0.100

0.001

  Df

9

9

0

9

9

 Cu

     

  Correlation

0.906

0.845

0.522

1.000

0.821

  Significance (2-tailed)

0.000

0.001

0.100

0.002

  Df

9

9

9

0

9

 Zn

     

  Correlation

0.974

0.967

0.841

0.821

1.000

  Significance (2-tailed)

0.000

0.000

0.001

0.002

  Df

9

9

9

9

0

(b)

     

 Cd

     

  Correlation

1.000

0.872

0.829

0.841

0.935

  Significance (2-tailed)

0.000

0.002

0.001

0.000

  Df

0

9

9

9

9

 Pb

     

  Correlation

0.872

1.000

0.840

0.924

0.913

  Significance (2-tailed)

0.000

0.001

0.000

0.000

  Df

9

0

9

9

9

 Mn

     

  Correlation

0.829

0.840

1.000

0.821

0.862

  Significance (2-tailed)

0.002

0.001

0.002

0.001

  Df

9

9

0

9

9

 Cu

     

  Correlation

0.841

0.924

0.821

1.000

0.788

  Significance (2-tailed)

0.001

0.000

0.002

0.004

  Df

9

9

9

0

9

 Zn

     

  Correlation

0.935

0.913

0.862

0.788

1.000

  Significance (2-tailed)

0.000

0.000

0.001

0.004

  Df

9

9

9

9

0

Figure 3 presents the mean concentrations (µg mL−1) of Cd, Pb, Mn, Cu and Zn in urine samples of patients with respect to their professions. Cd has the lowest concentration which ranged from 0.048 ± 0.012 to 0.091 ± 0.020 µg mL−1, while Cu has the highest concentration ranging from 0.269 ± 0.025 to 1.313 ± 0.055 µg mL−1. Generally, the metals concentration followed the order Cu > Zn > Pb > Mn > Cd for all professions except in Security personnel’s urine samples where Zn concentration are higher than Cu. The values obtained for Cu are higher in concentration than the required level of Cu in human urine of 0.02 µg/mL reported by WHO [28], while students had the highest level of Cu (1.313 ± 0.055 µg mL−1). With respect to profession, the concentrations of heavy metal were higher than the recommended normal urine level in human as already discussed. The t-test values for the metals are Cd (1.3546), Pb (14.071), Mn (6.845), Cu (6.702) and Zn (9.551). These values were higher than the critical value (2.36) at p ≤ 0.05 and 95% confidence level. This implies that patient’s profession has significant influence on the concentration of heavy metals found in their urine samples. Also, the correlation factors values (not shown) showed that there are positive correlations among the levels of Cd and Pb; Mn and Cd; Cu and Zn; Zn and Cd present in the urine samples with reference to the patients’ professions except for Cd and Cu that showed negative correlation.
Fig. 3

Concentrations (µg mL−1) of heavy metals in urine samples of patients with respect to profession

Figure 4 shows the concentrations of Cd, Pb, Mn, Cu and Zn (µg mL−1) for urine of patients with reference to their habits. Copper had the highest concentration (1.540 ± 0.054 µg mL−1) and the metals followed the order Cu > Zn > Pb > Mn > Cd. Category of patients that drink and smoke had the highest concentrations of metal analysed, Pb (0.316 ± 0.023 µg mL−1), Mn (0.183 ± 0.152 µg mL−1), Cu (1.540 ± 0.054 µg mL−1) and Zn (0.792 ± 0.173 µg mL−1) and this had been reported and documented in literature [29, 30].
Fig. 4

Concentrations (µg mL−1) of heavy metals in urine of patients with respect to their habits

The t-test results of the metals in urine samples of patients with respect to their habits are 10.193, 10.943, 11.655, 7.808 and 13.779 for Cd, Pd, Mn, Cu, and Zn respectively. The values are higher than the t-critical value of 3.18 at 95% confidence level. This implies that patients’ habits have significant influence on the levels of heavy metals found in their urine samples. The paired correlation values (not shown) indicates positive correlation among the metals (Cd, Pb, Mn, Cu and Zn) considering the patient’s habits such as drinking and smoking. This suggests that patients’ habits might have contributed to the high levels of heavy metals found in the urine samples. It has been reported that life styles e.g. smoking, drinking are some of the major sources of Cd and Pb [34].

Figure 5 presents the mean concentrations of Cd, Pb, Mn, Cu and Zn for urine of patients with respect to their diseases. The concentrations of metals followed the order Cu > Zn > Pb > Mn > Cd for all the diseases. The highest metal concentration was recorded for Cu (1.249 ± 0.118 µg mL−1) while Cd gave the lowest concentration (0.070 ± 0.048 µg mL−1). Patients suffering from Lupus Nephritis had the highest concentration of copper, which might be due to effect of environmental factors. Generally, the concentrations of metals obtained in this study were higher compared to the permissible levels of metals in urine. High concentrations of Pb and Cd could cause renal tubular damage, decrease in born mineralization, decreases lung functions and emphysema [29].
Fig. 5

Concentrations (µg mL−1) of heavy metals in urine of patients with respect to their diseases

The t-test values for Cd (19.127), Pb (9.548), Mn (7.671), Cu (12.591) and Zn (11.686) are greater than the t-test critical value (2.20) at p ≤ 0.05 and 95% confidence level. This suggests that patients’ disease has significant influence on the levels of heavy metals found in their urine samples. Also, the correlation values of the metals in urine samples (not shown) showed a positive correlation between the levels of Pb and Zn; and a weak correlation between Cu and Zn, Mn and Cd present in the patients with respect to their various diseases.

3.3 Concentration (µg g−1) of Heavy Metals in Tissue Samples

The concentrations (µg g−1) of metals in kidney samples analysed for both male and female corpses are presented in Fig. 6. Mn has the highest mean concentration of 1.025 ± 0.080 µg g−1 for male kidney samples while Pb gave the lowest concentration (0.144 ± 0.033 µg g−1. Female kidney samples gave the highest concentration for Mn (1.115 ± 0.064 µg g−1) and lowest concentration for Pb (0.153 ± 0.093 µg g−1). Thus, it could be inferred that for both gender, Mn has the highest concentration while Pb has the least concentration for the kidney samples. The concentrations of Pb in both male and female patient’s samples were lower compared to 1–1.5 µg g−1 reported by Bartis and Ashwood [35] for human kidney samples. The concentrations of Cd, Mn and Cu in this study were higher compared to 0.150 µg g−1, 0.120 µg g−1 and 0.570 µg g−1 reported by Yoo et al. [36] for the metals in human kidney samples while levels of Pb and Zn obtained in this study were lower than 1.5 µg g−1 and 40 µg g−1 reported for Pb and Zn respectively in human kidney samples by Yoo et al. [36].
Fig. 6

Concentrations (µg g−1) of heavy metals in male and female kidney samples

The t-test values revealed that Cd has t-value of 6.905 which is lower than the t-critical value of 12.71 at p ≤ 0.05. This implies that there is no significant difference in the concentration of Cd in the Kidney samples of male and female patients. Other metals, Pb, Mn, Cu and Zn had higher t-values 33.000, 23.778, 35.520 and 16.508 respectively, which are greater than the t-critical value suggesting significant differences in the metal’s concentration in the kidney samples of both male and female corpses.

The concentrations (µg g−1) of metals in liver tissue samples for both male and female corpses are represented in Fig. 7. Mn had the highest concentration (1.690 ± 0.257 µg g−1) while Pb had the lowest concentration (0.263 ± 0.156 µg g−1) for the male liver samples. In female liver samples, Mn has the highest concentration (1.690 ± 0.257 µg g−1) while Pb has the lowest concentration (0.251 ± 0.126 µg g−1). Ogunfowokan et al. [6] reported higher concentrations of Cd (31.88 µg g−1) and Zn (2.95 µg g−1) in liver of male human cadavers compared to the concentration of these metals in the present study. Also, the concentrations of Cd (0.380 ± 0.153 µg g−1) and Zn (0.407 ± 0.097 µg g−1) obtained in this study for female human cadaver’s liver were lower compare with 11.43 µg g−1 and 2.29 µg g−1 of Cd and Zn reported for the organ by Ogunfowokan et al. [6]. However, Mn concentration (1.690 µg g−1) for female cadaver’s liver reported by Ogunfowokan et al. [6] agreed with 1.690 µg g−1 Mn obtained in this study.
Fig. 7

Concentrations (µg g−1) of heavy metals in male and female liver samples

Results of the t-test shows that t-values for Cd (8.067) and Pb (6.107) were lower than the t-critical value of 12.71 at 95% confidence level and p ≤ 0.05. This means that concentrations of Cd and Pb have no significant difference in the liver samples of both male and female corpses, while the t-values for Cu (143.625) and Zn (61.615) were greater than t-critical value (12.71) at 95% confidence level and p ≤ 0.05 which is an indication that there are significant differences between the concentrations of these metals for the liver samples of both male and female corpses.

The concentrations (µg g−1) of heavy metals in lung tissue samples for both male and female corpses are presented in Fig. 8. Mn had the highest mean concentration of 1.430 ± 0.126 μg g−1 and 1.198 ± 0.088 μg g−1 in male and female samples respectively, while Pb had the least concentration of 0.175 ± 0.052 μg g−1 (male) and 0.105 ± 0.022 μg g−1 (female) samples. The t-test values revealed that Cd, Pb and Mn had t-values of 2.043, 4.000 and 11.328 respectively which are lower than t-critical value of 12.71 at 95% confidence level. This also suggests that there are no significant differences between these metals in the lung tissues of both male and female corpses studied.
Fig. 8

Concentrations (µg g−1) of heavy metals in male and female lung samples

4 Conclusions

This study determined the levels of Cd, Pb, Mn, Cu, and Zn in urine samples of patients with kidney, liver and lung related diseases; and in autopsy kidney, liver and lungs that were pathologically diseased, using Atomic Absorption Spectroscopy (AAS). The concentrations of the metals in the control urine samples used in this study were found to be higher than the maximum required levels recommended by ATSDR and WHO in human urine. The higher concentrations of Cd, Pb, Mn and Cu in male urine samples were found from ages 55 to 64 while for female urine samples, while lower concentrations were found in the lower age categories (ages 24–29). The concentrations of Cu in the urine samples for both male and female patients were found to be higher than those of Cd, Pb, Mn and Zn, and also higher than the recommended standard level in human urine. Generally, the concentrations of metals obtained in this study were higher compared to the permissible levels of metals in urine and therefore may be responsible for the associated diseases noticed in the patients. In most cases, the calculated t-values are higher than t-critical value for the metals at 95% confidence level for most variables compared suggesting that patients’ profession, habits and diseases have significant influence on the levels of heavy metals found in their urine samples.

Analysis of the diseased (human corpses) samples showed that higher concentrations of Mn was recorded for kidney, liver and lung samples for both male and female corpses while Pb had the lowest concentrations. Generally, the levels of heavy metals obtained in the tissue samples from this study corpse were lower than those obtained in previous studies reported in literature.

Notes

Acknowledgements

The authors would like to appreciate the Ethics and Research Committee of Obafemi Awolowo University Teaching Hospitals Complex Ile-Ife, Nigeria, for the ethical clearance certificate and the staff of Morbid Anatomy and Forensic Medicine Department, Renal Unit and Medical ward for their assistance with samples collection.

Funding

No internal (local) or international funding. The research was solely self-sponsored.

Compliance with Ethical Standards

Conflict of interest

There is no conflict of interest to be disclosed by authors.

Ethical Standards

Prior to collection of samples, ethical clearance certificate was obtained from the Ethics and Research Committee of Obafemi Awolowo University Teaching Hospitals Complex, Ile Ife, Nigeria. An informed consent of patients was seeked before urine and blood collection which was supervised by clinical experts and medical technologists at the hospitals.

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Copyright information

© The Tunisian Chemical Society and Springer Nature Switzerland AG 2019

Authors and Affiliations

  • A. O. Ogunfowokan
    • 1
    Email author
  • A. S. Adekunle
    • 1
    Email author
  • B. A. Oyebode
    • 1
  • J. A. O. Oyekunle
    • 1
  • A. O. Komolafe
    • 1
    • 2
  • G. O. Omoniyi-Esan
    • 1
    • 2
  1. 1.Department of ChemistryObafemi Awolowo UniversityIle-IfeNigeria
  2. 2.Department of Morbid Anatomy and Forensic MedicineObafemi Awolowo UniversityIle-IfeNigeria

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