Health Risk and Metal Contamination Assessment in Orange (Citrus aurantium) of Pakistan: a Multivariate Study.

Toxic metals (TMs) are serious health hazard and are known to accumulate in fruits, which are consumed by people, owning to their distinct sweet flavor and potential health benefits. In the present study, arsenic (As), lead (Pb), chromium (Cr), copper (Cu), cadmium (Cd), nickel (Ni), and mercury (Hg) were measured by atomic absorption spectrometry (AAS) techniques in orange samples from 11 farmhouses of Sargodha, Pakistan. Principal component analysis (PCA) is a very useful analysis and data-reducing tool. This technique was used to explore the relationship between different metal levels in the oranges. The conventional wet acid digestion method was validated and assured with two (02) certified reference materials NIST-SRM 1515-Apple and (CRM) NIST-1570a Spinach Leaves by recovery test coupled with standard addition procedure. The results showed that Cd and Cr average daily intakes are 0.084 and 0.15 mg slightly higher than World Health Organization/Food and Agriculture Organization of the United Nations (WHO/FAO) recommended tolerable daily intake respectively. The assessment of health risk was also carried out in form of total hazard quotient (THQ) and hazard index (HI), indicating three heavy metals (Pb, Cd, and Cr) have significant human health risk potential to consumers while Cu, As, Hg, and Ni have non-significant (THQ < 1) health risks.

Detection of Human Adenovirus, Rotavirus, and Enterovirus in Tap Water and Their Association with the Overall Quality of Water in Karachi, Pakistan.

Drinking water supplies in the developing world often serve as a biosphere for various organisms. Viral gastroenteritis is a neglected area of research in Pakistan, there are no data for the prevalence of enteric viruses in drinking water of the largest city of Karachi. The present study aimed to provide a survey of the existence of enteric viruses: human adenovirus (HAdV), human enteroviruses (hEV), and genotype A rotavirus (GARV) in tap water. Using a simple PCR approach, we detected 20%, 43%, and 23% of HAdV, hEV, and GARV in tap water samples, respectively. We have also shown an overall quality deficit of tap water at the pumping station and consumer tap. We have found no sample free from bacterial contaminations. The ranges for a total number of the heterotrophic plate count and coliform were found 8.7 × 102-4.5 × 106 CFU/mL and 210 to uncountable coliforms/100 mL, respectively. Moreover, we assessed the efficiency of small-scale water treatment methods for the removal of viruses.

Separation-preconcentration of nickel and lead in food samples by a combination of solid-liquid-solid dispersive extraction using SiO2 nanoparticles, ionic liquid-based dispersive liquid-liquid micro-extraction.

A microextraction method for the determination of nickel and lead using solid-liquid-solid dispersive extraction followed by ionic liquid-based dispersive liquid-liquid microextraction (SLSDE-ILDLLME) was presented. It was applied to the extraction of nickel and lead from food samples. Ammonium pyrrolidine dithiocarbamate (APDC) as complexing agent, [C4MIM][PF6] as ionic liquid, SiO2 as nanoparticles and 2 mol L(-1) HNO3 as eluent were used. Several important parameters such as amount of IL, extraction time, pH and volume of the complexing agent were investigated. The quantitative recoveries were obtained at pH 7.0 for analytes. Under the optimum conditions, the limits of detection (LODs) calculated using 3(Sd)blank/m were 0.17 for Ni(II) and 0.79 µg L(-1) for Pb(II) for aqueous solutions with 125 enrichment factor (EF). The limit of detections of the analyte ions (3(Sd)blank/m) for solid samples were 0.09 µg g(-1) (Ni) and 0.40 µg g(-1) (Pb). The accuracy of the proposed method was confirmed by the analysis of standard reference material (1577c bovine liver) and spiked recovery test. The proposed method was applied to determine nickel and lead levels in chicken, fish and meat samples.

Determination of essential elements (Cu, Fe and Zn) in juices of commercially available in Pakistan.

In present work, the concentration of essential elements, copper (Cu), iron (Fe) and zinc (Zn) were measured in ten different juices, to evaluate whether the intakes of under study nutrients accomplished within recommended permissible levels. Flame atomic absorption spectrometry (FAAS) was used to estimate and evaluate the levels of essential elements such as Cu, Fe and Zn in ten juice samples. The elements were analyzed prior to microwave assisted acid digestion (MAD). For comparison purpose, wet acid digestion (CAD) method was used for determination of essential elements. The accuracy of proposed method was evaluated by performing addition-recovery experiments to prove the reliability of the method. All obtained analytical results by MAD and CAD showed a good agreement at a 95% confidence level. The detection limits of Cu, Fe and Zn were 22.7 microg/L, 3.85 microg/L and 3.05 microg/L. The dietary intakes of all three studied elements have also been estimated.

Speciation and evaluation of Arsenic in surface water and groundwater samples: a multivariate case study.

The principal object of the current study was to estimate total arsenic and its inorganic speciation in different origins of surface water (n=480) and groundwater (n=240) of Sindh, Pakistan. This study provided a description based on the evaluation of physico-chemical parameters of collected water samples and possible distribution of As with respect to its speciation. The concentration of total inorganic As (iAs) and its species (As(3+) and As(5+)) for the surface and underground water was reported in terms of basic statistical parameters, principal component analysis, cluster analysis, metal-to-metal correlations and linear regression analyses. The chemical correlations were observed by PCA, which were used to classify the samples by CA, based on the PCA scores. Standard addition method confirmed the accuracy; the recoveries of As(3+) and iAs were found to be >98%. The concentration of As(5+) in the water samples was calculated by the difference of the total inorganic arsenic and As(3+). The results revealed that the groundwater of the understudied area was more contaminated as compared to the surface water samples. The mean concentration of As(3+) and As(5+) in the surface water and groundwater samples were in the range 3.0 to 18.3 and 8.74-352 microg/L, respectively.

Optimization of ultrasonic-assisted acid extraction of mercury in muscle tissues of fishes using multivariate strategy.

A simple and rapid ultrasound-assisted extraction procedure was developed for the determination of total mercury (Hg) in muscle tissues of freshwater fish species. A Plackett-Burman experimental design was used as a multivariate strategy for the evaluation of the effects of variables, such as presonication time (without ultrasonic stirring), sonication time, ultrasonic bath temperature, nitric acid concentration, hydrochloric acid concentration, and sample mass of muscle tissues. Some variables showed a significant effect on recovery, and they were further optimized by a 2(3) + star central composite design that involved 16 experiments. The validation was carried out by analysis of certified reference material DORM-2 (dog fish muscles); for comparative purposes, an acid digestion induced by microwave energy was used. Cold vapor atomic absorption spectrometry was used for the determination of total Hg. No significant differences were established between the analytical results and the certified values (paired t-test at P > 0.05). The LOD and LOQ of Hg were 0.133 and 0.445 microg/kg, respectively, which demonstrated the high sensitivity of the proposed procedure for the determination of Hg at trace levels. The Hg concentrations in the muscle tissues of 10 freshwater fish species were found in the range of 35.3-67.8 microg/kg on a dried basis, which were within the permissible limit of the World Health Organization.

Respiratory effects in people exposed to arsenic via the drinking water and tobacco smoking in southern part of Pakistan.

In this study, a survey has been conducted during 2005-2007 on surface and groundwater arsenic (As) contamination and its impact on the health of local population, of villages located on the banks of Manchar lake, southern part of Sindh, Pakistan. We have also assessed the relationship between arsenic exposure through respiratory disorders in male subjects with drinking water and smoking cigarettes made from tobacco grown in agricultural land irrigated with As contaminated lake water. The biological samples (blood and scalp hair) were collected from As exposed subjects (100% smokers) and age matched healthy male subjects (40.2% smoker and 59.8% non smokers) belong to unexposed areas for comparison purposes. The As concentration in drinking water (surface and underground water), agricultural soil, cigarette tobacco and biological samples were determined by electrothermal atomic absorption spectrometry. The range of As concentrations in lake water was 35.2-158 microg/L (average 97.5 microg/L), which is 3-15 folds higher than permissible limit of World Health Organization (WHO, 2004). While the As level in local cigarette tobacco was found to be 3-6 folds higher than branded cigarettes (0.37-0.79 microg/g). Arsenic exposed subjects (with and without RD) had significantly elevated levels of As in their biological samples as compared to referent male subject of unexposed area. These respiratory effects were more pronounced in individuals who had also As induced skin lesions. The linear regressions showed good correlations between As concentrations in water versus hair and blood samples of exposed subjects with and without respiratory problems.

Assessment of toxic metals in raw and processed milk samples using electrothermal atomic absorption spectrophotometer.

Milk and dairy products have been recognized all over the world for their beneficial influence on human health. The levels of toxic metals (TMs) are an important component of safety and quality of milk. A simple and efficient microwave assisted extraction (MAE) method has been developed for the determination of TMs (Al, Cd, Ni and Pb), in raw and processed milk samples. A Plackett-Burman experimental design and 2(3)+star central composite design, were applied in order to determine the optimum conditions for MAE. Concentrations of TMs were measured by electrothermal atomic absorption spectrometry. The accuracy of the optimized procedure was evaluated by standard addition method and conventional wet acid digestion method (CDM), for comparative purpose. No significant differences were observed (P>0.05), when comparing the values obtained by the proposed MAE method and CDM (paired t-test). The average relative standard deviation of the MAE method varied between 4.3% and 7.6% based on analyte (n=6). The proposed method was successfully applied for the determination of understudy TMs in milk samples. The results of raw and processed milk indicated that environmental conditions and manufacturing processes play a key role in the distribution of toxic metals in raw and processed milk.

Determination of toxic elements in infant formulae by using electrothermal atomic absorption spectrometer.

In present work concentrations of toxic elements (TEs), aluminum (Al), cadmium (Cd), and lead (Pb) in seventeen imported samples of infant milk-based (IMF) and infant soy-based formulae (ISF), were measured, to evaluate whether the intakes of understudy TEs accomplished within recommended permissible levels. The TEs were analyzed by electrothermal atomic absorption spectrometer, prior to microwave induced acid digestion. The validity of methodology was tested by simultaneously analyzing certified reference material and standard addition method. It was observed that ISF contains higher concentration of understudy toxic analytes as compared to IMF. The all three TEs, Al, Cd and Pb were detected in different branded infant formulae, in the range of (1070-2170), (10.5-34.4), and (28.7-119) microg/kg, respectively. The estimated intakes of TEs as microg/kg/week for infants (>1year) through milk formulae are well below the recommended tolerable levels of these elements.

Heavy metal accumulation in different varieties of wheat (Triticum aestivum L.) grown in soil amended with domestic sewage sludge.

The concentrations of heavy metals (HMs) in plants served to indicate the metal contamination status of the site, and also revealed the abilities of various plant species to take up and accumulate them from the soil dressed with sewage sludge. A study to comprehend the mobility and transport of HMs from soil and soil amended with untreated sewage sludge to different newly breaded varieties of wheat (Anmol, TJ-83, Abadgar and Mehran-89) in Pakistan. A pot-culture experiment was conducted to study the transfer of HMs to wheat grains, grown in soil (control) and soil amended with sewage sludge (test samples). The total and ethylenediaminetetraaceticacid (EDTA)-extractable HMs in agricultural soil and soil amended with domestic sewage sludge (SDWS) and wheat grains were analysed by flame atomic absorption spectrometer/electrothermal atomic absorption spectrometer, prior to microwave-assisted wet acid digestion method. The edible part of wheat plants (grains) from test samples presented high concentration of all HMs understudy (mgkg(-1)). Significant correlations were found between metals in exchangeable fractions of soil and SDWS, with total metals in control and test samples of wheat grains. The bio-concentration factors of all HMs were high in grains of two wheat varieties, TJ-83 and Mehran-89, as compared to other varieties, Anmol and Abadgar grown in the same agricultural plots.

Determination of copper and iron in biological samples of viral hepatitis (A-E) female patients.

There is accumulative evidence that the metabolism of iron and copper is altered in viral hepatic diseases, and these nutrients might have specific roles in their pathogenesis and progress. The aim of present study was to compare the level of copper (Cu) and iron (Fe) in biological samples (serum, urine, and scalp hair) of female patients suffering from different viral hepatitis (A, B, C, D, and E; n = 253) of age range 31-45 years. For comparative study, 95 healthy females of the same age group residing in the same city were selected. The elements in the biological samples were analyzed by flame atomic absorption spectrophotometry, prior to microwave-assisted acid digestion. The validity and accuracy of the methodology was checked by using certified reference materials (CRMs) and with those values obtained by conventional wet acid digestion method on same CRMs. The results of this study showed that the mean values of Cu and Fe were higher in sera and scalp hair samples of hepatitis patients than age-matched control subjects, while the difference was significant (p < 0.001), in the cases of viral hepatitis B and viral hepatitis C as compared to viral hepatitis A, D, and E. The urinary levels of these elements were found higher in the hepatitis patients than in the age-matched healthy controls (p < 0.05). These results are consistent with literature-reported data, confirming that hepatic iron and copper overload can directly cause lipid peroxidation and eventually hepatic damage.

The correlation of arsenic levels in drinking water with the biological samples of skin disorders.

Arsenic (As) poisoning has become a worldwide public health concern. The skin is quite sensitive to As and skin lesions are the most common and earliest nonmalignant effects associated to chronic As exposure. In 2005-2007, a survey was carried out on surface and groundwater arsenic contamination and relationships between As exposure via the drinking water and related adverse health effects (melanosis and keratosis) on villagers resides on the banks of Manchar lake, southern part of Sindh, Pakistan. We screened the population from arsenic-affected villages, 61 to 73% population were identified patients suffering from chronic arsenic toxicity. The effects of As toxicity via drinking water were estimated by biological samples (scalp hair and blood) of adults (males and females), have or have not skin problem (n=187). The referent samples of both genders were also collected from the areas having low level of As (<10 microg/L) in drinking water (n=121). Arsenic concentration in drinking water and biological samples were analyzed using electrothermal atomic absorption spectrometry. The range of arsenic concentrations in lake surface water was 35.2-158 microg/L, which is 3-15 folds higher than World Health Organization [WHO, 2004. Guidelines for drinking-water quality third ed., WHO Geneva Switzerland.]. It was observed that As concentration in the scalp hair and blood samples were above the range of permissible values 0.034-0.319 microg As/g for hair and <0.5-4.2 microg/L for blood. The linear regressions showed good correlations between arsenic concentrations in water versus hair and blood samples of exposed skin diseased subjects (R2=0.852 and 0.718) as compared to non-diseased subjects (R2=0.573 and 0.351), respectively.

Evaluation of an ultrasonic acid digestion procedure for total heavy metals determination in environmental and biological samples.

In this study, a sample preparation method based on ultrasonic assisted acid digestion (UAD) has been evaluated for total heavy metals (Cd, Cr, Ni and Pb) determination in different environmental (soil, sediment and sewage sludge), and biological (fish muscles, vegetables and grains) samples, using electrothermal atomic absorption spectrometry (ETAAS). The investigated parameters influencing UAD such as presonication time, sonication time, temperature of ultrasonic bath, and different acid mixtures were fully optimized, whereas power was maintained constant at 100% of nominal power of ultrasonic bath. Six different sets of above parameters were applied on six certified reference materials (CRMs) having different matrices. The accuracy of the method was also tested by comparing the results with those obtained from conventional hot plate assisted acid digestion method on same CRMs. Analytical results for HMs by both methods showed no significant difference at 95% confidence limit (p<0.05). Recoveries of HMs ranging from 96.2% to 102% and 96.3% to 98.6% were obtained from biological and environmental samples, respectively. The average relative standard deviation of UAD method varied between 3.5% and 8.2%, depending on the analyte.

Estimation of toxic metals in scalp hair samples of chronic kidney patients.

The determination of toxic metals (TMs) in the biological samples of human beings is an important clinical screening procedure. The aim of this work is to determine total content of TMs, aluminum (Al), cadmium (Cd), nickel (Ni), and lead (Pb) in scalp hair samples of chronic kidney male patients (CKPs) on maintenance hemodialysis, during the period of 2005-2007. The study included 115 CKPs (all smokers) and 150 controls or referents [82 (nonsmokers) and 68 (smokers)]. Both controls and patients (males) were of the same age group (ranged 25-55 years), socioeconomic status, localities, and dietary habits. The scalp hair samples were analyzed by electrothermal atomic absorption spectrometer, prior to microwave-induced acid digestion. The accuracy of the total Al, Cd, Ni, and Pb measurements was tested by simultaneously analyzing certified reference material (human hair NCS ZC81002). No significant differences were observed between the analytical results and the certified values (paired t test at p > 0.05). The levels of TMs in scalp hair samples of patients were found to be higher as compared to control nonsmoker and smokers. Moreover, the study shows that levels of Al, Cd, Ni, and Pb in scalp hair samples may be useful to evaluate the impact of cigarette smoking in kidney failure patients.

Speciation of heavy metals in untreated sewage sludge by using microwave assisted sequential extraction procedure.

A fast microwave assisted extraction procedure was developed and optimized for their eventual exploitation in the three-stage sequential extraction procedure proposed by modified BCR protocol (the community Bureau of Reference now the European Union "Measurement and Testing Programme"). The effects of the microwave treatment on the extraction of Cd, Cr, Cu, Ni, Pb and Zn from untreated sewage sludge collected from Hyderabad city (Pakistan) were compared with those obtained from sequential BCR extraction procedure. In sequential BCR method, each extraction step takes 16 h, where as with the use of compromised microwave conditions, extraction steps could be completed in about 120 s, for each step, respectively. Extractable Cd, Cr, Pb and Ni obtained by both comparable methodologies were measured by electrothermal atomic absorption spectrometry (ETAAS), while for Cu and Zn flame atomic absorption spectrometry (FAAS) was used. The validations of both extraction techniques were compared by the analysis of certified reference material of soil amended with sewage sludge (BCR 483). The results of the partitioning study of untreated waste water sewage sludge, indicate that more easily mobilized forms (step 1) were predominant for Cd, Ni and Zn (28.3, 28.4 and 43.7%), in contrast, the largest amount of Cd and Pb (66.4 and 72.8%) was associated with the iron/manganese oxide while Cr and Ni (71.2 and 38.7%) in organic matter/sulphide fractions. The overall metal recoveries in steps 1-3 (excluding residual step) were 95.3-104% of those obtained with the sequential BCR protocol. The accuracy of the proposed microwave extraction method (expressed as %R.S.D.) was lower than 10% for all metals.

Determination of toxic elements in different brands of cigarette by atomic absorption spectrometry using ultrasonic assisted acid digestion.

In this paper, a sample preparation method based on ultrasonic assisted acid extraction (UAE) of Al, As, Cd, Ni and Pb from filler tobacco (FT) of different local branded (LBCs) and imported branded cigarettes (IBCs) consumed in Pakistan. Acid extracts obtained upon sonication were determined by atomic absorption spectrometry. Parameters influencing extraction such as pre-sonication time 10-15 min, sonication time 2-10 min, mass of sample 0.1-0.5 g, particle size <35-125 microm and, extractant (nitric acid) concentrations (2-10%) were fully optimized to get quantitative recovery of toxic elements (TEs). The proposed method was validated with a certified reference material (CRM) Virginia tobacco leaves (ICHTJ-cta-VTL-2), and also applied a conventional acid digestion method (CDM) on same CRM, for comparative purpose. The average relative standard deviation of UAE method of CRM, ranged as 1.64-8.42% for N = 6, depending on the analytes. Extraction efficiencies for CRM were close to 97.1, 97.5, 98.5, 97.8 and 96.7 for Al, As, Cd, Ni and Pb, respectively. The concentrations of Al, As, Cd, Ni and Pb in cigarettes were present in all branded cigarettes, there is no any significance difference among LBCs and IBCs, except in few cases of LBCs, have high level of Cd and Al. Our results were compared with other worldwide studies.

Evaluation of toxic metals in blood and urine samples of chronic renal failure patients, before and after dialysis.

The determination of toxic elements in the biological samples of human beings is an important clinical screening procedure. The aim of this work was to determine total content of toxic elements-aluminum (Al), cadmium (Cd), and lead (Pb)-in whole blood and urine samples of male chronic renal failure patients (CRFPs) on maintenance hemodialysis from 2006 to 2007. The study included 100 CRFPs, plus 150 healthy volunteers in the control group. The concentration of toxic elements (TEs) were determined in blood sample before and after hemodialysis, while urine sample was determined once, before dialysis. Toxic elements were analyzed by electrothermal atomic absorption spectrometer, prior to microwave-induced acid digestion. The accuracy of the total Al, Cd, and Pb measurements was tested by simultaneously analyzing certified reference materials. No significant differences were established between the analytical results and the certified values (paired t-test at p > 0.05). The levels of TEs in blood samples of patients before dialysis were found to be higher than blood samples after dialysis session. In the control group, the blood levels of Al, Cd, and Pb were significantly lower than the chronic renal failure patients. Moreover, the study shows that analyzing levels of Al, Cd, and Pb may be useful in hemodialysis patients in evaluating TEs status.

Improved extraction method for the determination of iron, copper, and nickel in new varieties of sunflower oil by atomic absorption spectroscopy.

A simple and fast procedure is proposed for the extraction of iron (Fe), copper (Cu), and nickel (Ni) in 16 varieties of sunflower seed oil samples using an ultrasonic bath. The experimental parameters of the ultrasonic-assisted extraction (UAE) method were optimized to improve the sensitivity and detect the metals at trace levels in minimum time. Conventional wet acid digestion method was used for comparative purposes. The optimum recovery of all 3 metals was obtained by UAE for 7 min, while the separation of aqueous and organic phases after extraction using centrifugation (UAE-2) required 3 min, as compared to the conventional equilibration method (UAE-1) that required 90 min. The respective recoveries of Cu, Fe, and Ni obtained with UAE-2 were in the range of 95.8-97.5, 93.5-98.3, and 95.6-98.2%, respectively, for different varieties of sunflower oil samples. Accuracy was determined by the standard addition method. Under the optimum operating conditions, the limits of detection obtained from the standard addition curves were 21.7, 20.4, and 35.6 ng/mL for Fe, Cu, and Ni, respectively. The fact that all varieties of sunflower oil contain significant amounts of Fe, Cu, and Ni indicates the deterioration of sunflower oil quality immediately after extraction from seeds, which poses a threat to oil quality and human health.

A multivariate study: variation in uptake of trace and toxic elements by various varieties of Sorghum bicolor L.

The aim of present study was to evaluate the variation in uptake of elements (As, Ca, Cd, Co, Cr, Cu, Fe, K, Mg, Mn, Na, Ni, Pb and Zn) by different varieties of Sorghum bicolor L., plants grown in soil amended with untreated industrial waste water sewage sludge (SUIS), on same experimental plots. The power of chemometrics was also used in exploring the potential natural and/or anthropogenic sources responsible for elemental contents in different varieties of sorghum. Hierarchical cluster analysis was used to explore the different variety of sorghum grouping according to corresponding their SUIS samples as additional information to the output obtained by principal component analysis. Significant genotypic variation was detected in the fourteen elements concentrations in sorghum grains, indicating the possibility to reduce the concentration of toxic elements in grains through breeding approach. It was observed that high tolerance limit of toxic elements was observed in sorghum variety PARC-SV-1.

Evaluation of status of toxic metals in biological samples of diabetes mellitus patients.

There is accumulating evidence that the metabolism of several trace elements is altered in diabetes mellitus and that these nutrients might have specific roles in the pathogenesis and progress of this disease. The aim of present study was to compare the level of toxic elements, lead (Pb), cadmium (Cd), and arsenic (As) in biological samples (whole blood, urine, and scalp hair) of patients having diabetes mellitus type-2 age ranged (31-60) (n=238), with those of age matched non-diabetics (ND) as control subjects (n=196), of both genders. The concentrations of elements were measured by means of an atomic absorption spectrophotometer after microwave-assisted acid digestion. The validity and accuracy was checked by conventional wet acid digestion method and using certified reference materials. The overall recoveries of all elements were found in the range of 98.1-99.4% of certified values. The results of this study showed that the mean values of Pb, Cd and, As were significantly higher in scalp hair samples of smoker and non-smoker diabetic patients as compared to control subjects (p<0.001). The concentration of understudy toxic metals was also high in blood and urine samples of DM patient but difference was more significant in smoker DM patients. These results are consistent with those obtained in other studies, confirming that toxic metals may play a role in the development of diabetes mellitus.