Levels of zinc, copper, cadmium, and lead in fruits ...

Environ Monit Assess (2015) 187:676 DOI 10.1007/s10661-015-4905-8

Levels of zinc, copper, cadmium, and lead in fruits and vegetables grown and consumed in Aseer Region, Saudi Arabia Mohammed D. Y. Oteef & Khaled F. Fawy & Hisham S. M. Abd-Rabboh & Abubakr M. Idris

Received: 19 July 2015 / Accepted: 30 September 2015 # Springer International Publishing Switzerland 2015

Abstract The levels of four metals (Zn, Cu, Cd, and Pb) were evaluated in two fruit types (apricot and fig), a fruity vegetable (tomato), and three leafy vegetables (arugula, spinach, and lettuce) that are commonly grown and consumed in Aseer Region, Saudi Arabia. Flame atomic absorption spectrophotometry was employed for quantification. The quality of results was checked by a certified reference material (NIST SRM 1570a). Good recovery values in the range of 87–104 % were achieved. Metals were quantified in washed and unwashed samples to evaluate the effect of washing. Statistically, no significant difference was noticed (p>0.05), except for Zn in arugula and Cu in apricot and spinach. The levels of metals found in the analyzed fruits and vegetables were in their normal ranges in crops and not posing any serious

M. D. Y. Oteef : K. F. Fawy : H. S. M. Abd-Rabboh : A. M. Idris Department of Chemistry, College of Science, King Khalid University, Abha 62529, Saudi Arabia M. D. Y. Oteef (*) Department of Chemistry, College of Science, Jazan University, Jazan 82817, Saudi Arabia e-mail: [email protected] M. D. Y. Oteef e-mail: [email protected] H. S. M. Abd-Rabboh Department of Chemistry, Faculty of Science, Ain Shams University, Cairo 11566, Egypt

risks to the consumers in Aseer Region. The toxic elements Pb and Cd were well below the maximum levels set in the Saudi and international food standards. Zn and Cu levels were comparable to the ranges reported in worldwide previous studies. Keywords Metal levels . Fruits . Vegetables . Atomic absorption spectrometry . Heavy metals . Aseer . Saudi Arabia

Introduction Due to the high rate of diabetes, hypertension, and obesity in Saudi Arabia, healthy diet, including fruits and vegetables, is highly recommended by the Ministry of Health. Internationally, the World Health Organization also promotes the consumption of fruits and vegetables, a matter that is considered as a global priority (FAO 2003; FAO/WHO 2004). Furthermore, a diet rich in fruit and vegetables has been reported to reduce the risks of ischemic heart disease, stroke, and colorectal as well as gastric, lung, and esophageal cancers, which should save the lives of almost three million a year (WHO 2002). Despite their valuable benefits for humans, fruits and vegetables may pose serious risks to human health when contaminated with toxic substances such as pesticides and metals. The sources of metals in fruits and vegetables are from natural or anthropogenic origin. Metals from both origins take the way to plants through

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water and soil. Metals of anthropogenic origin are commonly from agricultural activities, i.e., the use of fertilizers and pesticides, in addition to the emissions from metal plating, mining operations, fertilizer industries, tanneries, paint industry, battery manufacturing, paper industries, pesticides industry, transportation, and sewage treatment plants (Bradl 2005). Metals have been classified as essential (e.g., Zn, Cu, Fe, Mn, and Se), probably essential (e.g., V and Co), and potentially toxic (As, Cd, Pb, Hg, and Ni) (Ebdon 2001). However, all these metals have toxic effects when there is excessive exposure (Woimant & Trocello 2014). Exposure to excessive levels of zinc, for example, may appear with symptoms including the called metal fume fever, stomach cramps, nausea, and vomiting. Prolonged ingestion of high zinc levels for several months may cause anemia, damage the pancreas, and decrease the levels of high-density lipoprotein (HDL) cholesterol. In contrast, zinc deficiency may cause loss of appetite, decreased sense of taste and smell, decreased immune function, slow wound healing, and skin sores. It may also cause poorly developed sex organs and retarded growth in young men. In addition, zinc deficiency in pregnant women may cause birth defects (ATSDR 2005). Regarding toxic metals, it has been reported that Pb, even at low levels, causes harms in the hematological and neurological systems (Jooste et al. 2015; USEPA 2004). Pb can also affect the cardiovascular system and kidneys (ATSDR 2007; Jooste et al. 2015). Lung and prostate cancers (Fraser et al. 2013; Nawrot et al. 2006; Vinceti et al. 2007), in addition to kidney and bone diseases (Alfvén et al. 2002; Fraser et al. 2013; Jarup & Akesson 2009), were reported to be linked to Cd exposure. Because of the benefits of consuming fruits and vegetables as a healthy diet, and the probability of their exposure to metal contamination, the investigation of metal levels is of great concern. Accordingly, this work was dedicated to the quantification and assessment of Zn, Cu, Cd, and Pb in two fruit types (apricot and fig), a fruity vegetable (tomato), and three leafy vegetables (arugula, spinach, and lettuce) that are commonly grown and consumed in Aseer Region, Saudi Arabia.

Materials and methods Study area Aseer Region is located in the southwest of Saudi Arabia between latitude 17° 25′ and 19° 50′ to the north and longitude 50.00° to 41° 50′ to the east with an area of 81,000 km2 (Arshad 2015). The estimated population of this region for the year 2015 is 2,194,463 (SCDSI 2010). Abha is the capital city for this region, and Khamis Mushait is the second main city and the commercial city of this area. Aseer Region is one of the main national parks and tourism destinations in Saudi Arabia. Assoda is a summer resort located to the northwest of Abha City. Figure 1 shows the study area and sampling locations in Aseer Region.

Sampling In this study, most of the fruit and vegetable samples were collected from the farms and markets in Abha and Khamis Mushait. Markets included wholesale markets, roadside retailers, small grocery shops, and large supermarkets. Some fruit samples were collected from Assoda summer resort as many fruit farms are located there. All samples were collected over the period March 2013 to March 2014. Samples were collected and processed directly for the analysis within 1–3 days. Edible parts only were used for the analysis. Samples included two fruit types (apricot and fig), a fruity vegetable (tomato), and three leafy vegetables (arugula, spinach, and lettuce), which are among the most grown and consumed fruits and vegetables in Aseer Region. To assure good representation of markets and farms as possible, each type was represented by 11 samples, except lettuce that was represented by five samples. As a result, a total of 60 samples were collected. About 3 kg of each sample was collected in a labeled polyethylene bag. At the laboratory, samples were divided into two portions, “washed” and “unwashed.” The “washed” portion of each sample was carefully washed with distilled water before further preparations, while the “unwashed” portion was prepared without rinsing. Both portions were prepared directly for the analysis or if necessary stored in the fridge at 4 °C for no more than 3 days. Each portion was prepared in triplicates for the analysis.


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Fig. 1 Study area and sampling locations in Aseer Region

Reagents and materials

Sample preparation

Water used in the current study was double distilled and then deionized (resistivity 18.2 MΩ cm at 25 °C). Hydrogen peroxide (35 %, Puriss p.a.) and nitric acid (70 %, Puriss p.a.) were purchased from SigmaAldrich (Steinheim, Germany). Standard solutions (1,000 mg/L) of Zn, Cd, and Pb were purchased from Acros (Geel, Belgium). Standard solution (1,000 mg/L) of Cu was purchased from Panreac (Barcelona, Spain). Standard reference material for trace elements in spinach leaves (SRM 1570a) was obtained from the US National Institute of Standards and Technology (Gaithersburg, USA).

Only edible parts of each fruit and vegetable type were used for the analysis. Any rotted or damaged parts were removed before preparation as a simulation to the real food preparation for human consumption. Fresh sample was weighed and then cut into small portions for drying in an oven at 105 °C. Completely dry sample was achieved within 72–96 h. Complete dryness was checked visually. Dry sample was ground in a household coffee grinder and sieved in a 0.8-mm mesh. It was then stored in a pre-labeled polyethylene bag for digestion. A wet digestion method, which was described by Altundag and Tuzen (2011), was modified and applied for all fruits and vegetables with the exception of spinach. A dry ashing digestion method described by Karla (1998) was used for spinach samples. In the wet digestion method, about 5 g of the dried and ground sample was placed in a glass digestion tube. A volume of 9 mL of nitric acid (70 %) was added to the sample and left for about 4 h at room temperature. Further 9 mL of nitric acid was added, and sample was left to digest overnight at room temperature. A third portion of 9 mL of nitric acid was added, and the heating program was started in the digestion block with a temperature of 45 °C for 45 min then raised to 90 °C for more 45 min. After which, the sample was left to cool

Instrumentation An atomic absorption spectrometer (AAS, SpectrAA 220, Varian, Mulgrave, Australia) equipped with sample introduction system (SPS 5, Varian, Mulgrave, Australia) was used for metal analysis. Air/acetylene flame and deuterium background corrector were employed. Wet digestion was conducted using an Automatic Kjeldahl Digestion Unit (model DKL, Velp Scientifica, Usmate, Italy). Dry ashing was achieved using a programmable muffle furnace (Lenton, Hope Valley, UK).

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down to room temperature. Five milliliters of hydrogen peroxide was then added, and temperature was raised to 120 °C for 2 h. The steps of cooling, adding hydrogen peroxide, and heating at 120 °C for 2 h were repeated two times to completely digest samples. The digest was then filtered through Whatman® no. 42 filter paper into a 50-mL volumetric flask, and the volume was completed to the mark with deionized water. This solution was used directly for AAS measurement or in some cases further dilution was conducted. In the dry ashing method, 5 g of dried and ground spinach leaves was placed in a 50-mL porcelain crucible and ashed in a programmable muffle furnace. Temperature was raised gradually in a rate of 275 °C/h to reach 550 °C. Sample was kept ashing at this temperature for 16 h before cooling down. When cool, drops of deionized water followed by 10 mL of 50 % (v/v) nitric acid were added slowly. The crucible was then placed on a hot plate to dryness and then placed again in the muffle furnace for ashing at 550 °C for 2 h. This process was repeated from three to four times till getting a white or gray ash. The ash was then dissolved in 10 mL of 20 % (v/v) nitric acid on a hot plate at about 100 °C. The solution was left to cool down and then filtered through Whatman® no. 42 filter paper into a 50-mL volumetric flask. The volume was then completed to the mark with deionized water. This solution was used directly for AAS measurement or in some cases further dilution was conducted. Atomic absorption spectrophotometry measurements The digested samples were analyzed for Zn, Cu, Cd, and Pb using flame AAS. Prepared reagent blanks and digests of the reference material were analyzed in parallel with each type of fruits and vegetables. The AAS system was calibrated using at least five standard solutions for external calibration of each metal. The instrument software was used to generate the calibration curves and to calculate all concentrations in milligram per liter.


analyzed in replicates, in parallel to each batch of fruit and vegetable samples. The mean and standard deviation of the measured values, in addition to certified values and recovery values, are compiled in Table 1. Good recovery values were obtained (87–104 %) for Zn, Cu, and Cd, which are comparable to recovery values reported in the literature for different analytical methods used for the analysis of metals in fruits and vegetables (Akinyele & Shokunbi, 2015; Altundag & Tuzen 2011; Guerra et al. 2012; Radwan & Salama 2006; Rahman et al. 2014). The level of Pb in the certified material SRM 1570a (0.20 mg/kg) was found lower than the limit of detection of the flame AAS method used. The limit of detection (LOD) and the limit of quantification (LOQ) of the wet digestion method were estimated for the examined metals based on the instrumental responses for replicate analyses of reagent blanks. The values of LOD and LOQ for the four metals are shown in Table 2. Seven replicates of reagent blank were prepared using the relevant procedure. Replicates were analyzed three times by the AAS after switching off and restarting the instrument. LOD was calculated as three times the standard deviation of the resulted 21 determinations, while LOQ was calculated as ten times the standard deviation (Gebrekidan et al. 2013; Rahmalan et al. 1996). Values were expressed as milligram per liter for solutions and were converted to the corresponding milligram per kilogram dry weight using a weight of 5 g of dry sample mass and an extract solution volume of 50 mL. However, the milligram per liter values were the ones used in evaluating the results. The experimental values of LOD and LOQ obtained in the current study are comparable to other values obtained in previous studies using flame AAS or even better for some metals (Gebrekidan et al. 2013; Guerra et al. 2012; Sharma et al. 2008). In the case of Pb, lower LODs are achieved using more sensitive techniques such as graphite furnace AAS (Bagdatlioglu et al. 2010; Corguinha et al. 2015) and ICP-MS (Rahman et al. 2014).

Results and discussion Quality of measurements

Assessment of levels of metals in fruits and vegetables

The quality of the two sample treatment methods applied in the current study was checked by the standard reference material 1570a. The reference material was

The levels of zinc, copper, cadmium, and lead in the different fruits and vegetables analyzed in the current study are presented in Table 3.


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Table 1 Measurements of SRM 1570a using dry and wet digestion methods Metal

Certified value

Dry digestion methoda Measured valuec

Wet digestion methodb Mean recoveryd

Measured valuec

Mean recoveryd

Zn

82±3

82.43±0.22

100

84.47±6.58

103

Cu

12.2±0.6

11.37±0.19

93

10.57±0.21

87

Cd

2.89±0.07

2.72±0.09

94

3.00±0.13

104

Pb

0.20e

NAf

NAf

NAf

NAf

a

Dry digestion means of duplicate analyses

b

Wet digestion means of 11 replicates

c

Values are presented in milligram per kilogram dry weight (amount±SD)

d

Mean recovery are presented in percentage

e

Information concentration value

f

Not detected

Zinc In the present study, zinc was found in all fruits and vegetables with a mean concentration in the washed samples ranging from 7.86±1.69 to 60.55±36.02 mg/kg dry weight (Table 3). The levels were in the order arugula > spinach > lettuce > tomato > apricot > fig. The higher levels of zinc in this study were noticed in leafy vegetables compared to fruits and fruity vegetable samples, which is consistent with similar notes reported previously (Roba et al. 2015). Zinc is considered as an essential plant micronutrient, and it has a long biological half-life in plants (Nagajyoti et al. 2010). Therefore, it is usually present at sufficient levels in plant tissues. Zn levels in the current study are well in the range of zinc in crops, i.e., 15–200 mg/kg (Nagajyoti et al. 2010). Moreover, Zn levels in this study are comparable to the levels found in fruits and vegetables cultivated or marketed in other parts of Saudi Arabia (Al Jassir et al. 2005; Ali & Al-Qahtani 2012; Mohamed et al. 2003). The levels are also comparable to those found in fruits and vegetables analyzed from different countries around

Table 2 Limits of detection and quantification of metals for the wet digestion method Characteristic

Zn

Cu

Cd

Pb

LOD, mg/L

0.006

0.013

0.006

0.04

LOQ, mg/L

0.021

0.044

0.019

0.13

LOD, mg/kg dry weight

0.06

0.13

0.06

0.40

LOQ, mg/kg dry weight

0.21

0.44

0.19

1.30

the world, including India (Sharma et al. 2008), Indonesia (Siaka et al. 2014), Algeria (Cherfi et al. 2014), Egypt (Radwan & Salama 2006), and Nigeria (Babatunde et al. 2014). However, several other international studies reported relatively lower levels of zinc in fruits and vegetables compared to the levels found in this study (Altundag & Tuzen 2011; Bagdatlioglu et al. 2010; Gebrekidan et al. 2013; Hamurcu et al. 2010; Lente et al. 2014; Osma et al. 2012). Higher levels of Zn in fruits and vegetables were reported previously for samples grown in contaminated areas or irrigated with wastewaters (Chopra and Pathak 2015; Kim et al. 2015). Copper Copper mean concentration in the washed fruits and vegetables analyzed in the current study ranges from 3.54±0.83 to 9.74±1.83 mg/kg dry weight (Table 3). The mean concentration of copper in the washed fruits and vegetables was in the order spinach > tomato > arugula > apricot > lettuce > fig. Plants, in general, contain sufficient levels of this metal in their tissues as it is considered a micronutrient. The levels obtained in the current study are in agreement with the range of copper in agricultural crops of 4–15 mg/kg dry weight (Nagajyoti et al. 2010). In addition, these levels are comparable to the levels found in fruits and vegetables analyzed from other parts of Saudi Arabia (Al Jassir et al. 2005; Ali & Al-Qahtani 2012; Mohamed et al. 2003). They are also comparable to the levels found in fruits and vegetables from different parts of the world including India (Ghosh et al. 2013), Sri Lanka (Kananke

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Table 3 Metal concentrations in the different fruits and vegetables in milligram per kilogram dry weight presented as the mean±standard deviation (n=11 except for lettuce n=5) with the range in parenthesis Fruit/ Zn vegetables Washed

Cu

Cd

Pb

Unwashed

Washed

Unwashed

Washed

Unwashed

Washed

Unwashed

10.28±7.16 (5.17–29.04) 7.86±1.69 (5.35–10.13) 12.04±2.09 (9.24–14.90) 60.55±36.02 (25.86–126.30)

11.03±8.36 (4.70–33.32) 7.92±1.88 (4.73–10.90) 12.05±1.94 (9.46–14.75) 87.62±67.69 (23.41–234.88)

4.97±0.99 (2.71–6.40) 3.54±0.83 (2.09–4.86) 5.58±1.83 (3.17–8.56) 5.56±1.29 (3.89–8.78)

5.23±1.16 (2.60–6.96) 3.70±0.76 (2.18–4.68) 5.82±1.90 (2.92–7.97) 6.29±1.32 (4.42–8.62)