COMPARATIVE ASSESSMENT OF MINERAL ELEMENTS AND HEAVY METALS ACCUMULATION IN VEGETABLE SPECIES

The heavy metal (cadmium, lead, nickel, chromium) and mineral element (potassium, phosphorus, calcium, magnesium) levels in edible parts of tomato, potato, spinach, beetroot, parsley, parsnip, carrot, cauliflower, pepper and broccoli were determined by atomic absorption spectrophotometry. Six samples for each species originating from different localities were collected from green markets. The heavy metal concentrations ranged from <0.01 to 2.37, <0.01 to 7.72, <0.01 to 5.37 and <0.01 to 9.31 g/g for cadmium, chromium, nickel and lead, respectively. The order of the macroelement concentrations in dry matter for all vegetable species was as follows: potassium > calcium > phosphorus > magnesium. The highest mean levels of the heavy metals, as well as of potassium, calcium and magnesium, were found in spinach. A large number of samples containing high levels of toxic heavy metals, especially of cadmium and lead, impose the necessity for strict regulative guidelines concerning individual vegetable crops production, harvest, handling and storing, in order to diminish possibility of contamination.


INTRODUCTION
Production of healthy and high-quality plants for human consumption should be considered as the prime goal of successful food production, beyond economic benefit.Vegetables and fruits are essential source of vitamins, mineral elements, fibres and also beneficial for human health (Radwan and Salama, 2006).Their everyday intake is highly recommended in the human diet, and their role in maintenance of health, and in prophylaxis and curing of numerous diseases is evident (D 'Mello, 2003).Contents of both nutritive components important for human nutrition and other ineligible substances, such as heavy metals, determine the foodstuff quality and safety (Demirezen and Aksoy, 2006; Khoshgoftar-manesh et al., 2009;Mastilović et al., 2010).Plants acquire heavy metals primarily from the soil, in concomitance with uptake and accumulation of elements essential for both plant metabolism and consumers organism, as well as from particle deposition on plant parts exposed to contaminated air (Zurera et al., 1997).Some of the heavy metals are trace elements (iron, copper, manganese, and zinc), commonly found in foodstuffs, fruits and vegetables, and are essential for a healthy life in small amounts.Also, plants easily uptake and accumulate toxic elements.Lead, cadmium, and mercury are nonessential heavy metals widely distributed throughout the environment due to soil erosion, intensive industrial and agricultural processes (Fowler, 1973;Svete et al., 2001).These elements are potent metabolic poisons to plants, animals, and humans, due to the poor potential of natural detoxification and their tendency to accumulate within the organism (Radwan and Salama, 2006).Higher doses of these metals may cause metabolic disorders, growth inhibition for most plants, thus reducing food quality (Majid et al., 2011).Therefore, the monitoring of heavy metals accumulation in plant foods has gained attention in recent years due to either their regulative role in metabolic processes or toxic effects on living organisms.The heavy metal contents in vegetables depend on the species and genotype, environmental conditions, manner of cultivation (field or glass house production), plant part, leaf arrangement (outer or inner), and methods of harvesting, processing and cooking (Quarterman, 1973;Stevanović et al., 2001;Jomová et al., 2004).
Growing demands for food production due to increase of human population encourage individual production of crop species at small farms.These products are available for consumers at green markets, but the process of their cultivation, harvest, transport and eventual storage conditions, as well as chemical composition analyses are far beyond any legislative control.
Considering the lack of information on mineral and heavy metal contents in vegetables originating from individual production on small farms, the aims of the present work were the following: a) to analyse the concentration of cadmium, nickel, lead and chromium in edible parts of vegetables, b) to identify good sources of essential mineral elements (potassium, phosphorus, magnesium and calcium) among analysed plants, c) to compare the heavy metals concentration with maximum permitted levels for food, d) to identify species efficient in heavy metal accumulation in edible parts of plants, e) to explore the relation between the origin of the vegetables and the heavy metal and mineral contents.

MATERIAL AND METHODS
Plant samples were collected from three green markets in the city of Novi Sad, during the period September-October 2011.Two samples of the same species from each market were collected, originating from different localities in the vicinity of Novi Sad.The samples included the most represented plants in human diet of the region: tomato (Lycopersicon esculentum Mill.), potato (Solanum tuberosum L.), spinach (Spinacia oleracea L.), beetroot (Beta vulgaris L.), parsley (Petroselinum crispum (Mill.)Fuss), parsnip (Pastinaca sativa L.), carrot (Daucus carota L.), cauliflower (Brassica oleracea L. var.botrytis L.), pepper (Capsicum annuum L.) and broccoli (Brassica oleracea L. var.sylvestris L.).
Only edible parts of the collected plants were subjected to further analysis.The samples were arranged on white paper and allowed to dry at room temperature for two days, following oven-drying at 80 °C t for 24 h.Dry samples were milled and stored in clean and dry glass containers.Concentration of heavy metals and macroelements (potassium, calcium, phosphorus, magnesium) was determined by flame atomic absorption spectrophotometry (Varian, AAS240FS).The plant material was digested using closed vessel high pressure microwave digestion (Milestone D series Microwave Digestion System).The analysis of each sample was performed in three independent replicates.The data were statistically processed using analysis of variance (ANOVA).Comparison of localities was estimated using Duncan's multiple range test, at a level of significance of p<0.05.

RESULTS AND DISCUSSION
The macroelement concentrations in selected vegetable species are given in Tables 1 and 2. The values varied considerably between localities and plant species.Minerals are important and essential ingredients of diet required for optimal metabolic activities of the human body.The concentration of phosphorus ranged Data presented as mean values ± SD.Means for each species within the column followed by different letters are significantly different (p<0.05) from 0.18% (beet root, locality II) to 0.77% (parsnip, locality V).According to average phosphorus concentration, cauliflower and broccoli are good sources of this mineral in the human diet.The majority of phosphorus found in the human body is bound up with calcium in the bones as calcium phosphate, and the remainder is distributed as inorganic phosphate and in all cells as ATP, as well as in the genetic materials of DNA and RNA (Haas and Levin, 2006).It is involved in most biochemical reactions, energy production and exchange.Phosphorus and calcium combine in equal amounts to form the calcium phosphate in bones, so the ideal ratio of calcium to phosphorus in the diet is 1:1 (Haas and Levin, 2006).Of all vegetable species studied in the present work, the closest to ideal calcium/phosphorus ratio was found in parsnip (1.15) and pepper (1.17).
Calcium is the most abundant mineral in the human body, serving important functions in the formation of bones and teeth, regular behaviour of heart muscles, nerves and the blood clotting processes.Magnesium is important for proper muscle contraction, blood clotting, the regulation of blood pressure and lung function (Menchaca, 2009).It is also essential part of many enzymes responsible for transfer of energy (Danilcenko et al., 2011).In the vegetable species analysed, calcium ranged from 0.23% (parsnip, locality I) to 2.1% (beet root, locality VI), and magnesium from 0.07% (potato, locality VI) to 0.86% (beet root, locality II).The dietary calcium/magnesium ratio is responsible for the frequency of heart diseases (Kikunaga et al., 1999), and the recommended ratio that lowers this risk is 2:1.In vegetable species surveyed in the present work,   Pb-enriched manure, chemical fertilizers and pesticides (Khoshgoftarmanesh et al., 2009).Chromium is essential mineral important for carbohydrate metabolism in humans and animals (Parveen et al., 2003).The concentration of chromium in analysed vegetable species varied from 0.01 to 7.30 g/g (Table 4).Significant differences were recorded in chromium concentration between localities of origin in all sampled species.Average values obtained for each species, taking into account all six localities; fell between 1.00 and 2.00 g/g, except for spinach with 3.67 g/g of chromium.Beside chromium, spinach contained the highest average amounts of cadmium, nickel and lead as well (0.89, 3.35 and 5.81 g/g, respectively).Nickel is a nutritionally essential trace metal for several animal species, micro-organisms and plants.Therefore, either deficiency or toxicity symptoms can occur when, too little or too much Ni is taken up, respectively (Cempel and Nikel, 2006).Essentiality of nickel is considered possible for human organism due to its role as a cofactor or structural component in an enzyme (Nielsen, 2003).Most nickel in the human body originates from drinking water and food, however, the gastrointestinal route is of lesser importance, due to its limited intestinal absorption (Cempel and Nikel, 2006).The nickel concentrations in selected vegetable samples were in the range from <0.01 g/g (potato, parsnip, pepper) to 5.37 g/g (spinach) (Table 4).Nickel levels in foodstuffs generally range from less than 0.1 to 0.5 g/g (Cempel and Nikel, 2006).Range of Ni concentrations obtained in the present work was lower than that reported for vegetables grown in either urban or rural areas in Turkey (Demirezen and Aksoy, 2006).
Accumulation of heavy metals in plants may be the consequence of irrigation with contaminated water, the addition of fertilizers and metal-based pesticides, Industrial emissions, transportation, harvesting process, activities during storage and/or sale (Radwan and Sharma, 2006).High contents of these elements in soil are supposed to increase the risk of the uptake by plants, considering positive correlation between the metal content in soils and vegetables (Naser et al., 2009).Therefore, soil quality standards are important instrument for risk assessment of polluted soils and their impact on human health, water resources and other environmental impacts (Atanassov, 2007).Previous studies have reported variable relationships between the metal concentrations in certain plant parts and their total concentrations in soil.For example, De Pieri et al. (1997) reported inverse relationship between carrot root lead and total soil lead concentrations, and relatively high concentration of cadmium in potato tubers at the site with low cadmium concentration in comparison to other soils studied.According to their findings, the heavy metal concentration in the soil should not be the only criterion for estimating vegetable quality, as processes related to plant metal uptake and translocation must also be taken into account.Considering significant variability among localities concerning the heavy metal concentration in vegetable species surveyed in the present work, beside the analysis of soil quality at small farms, special attention should be paid to genotype specificity of toxic metals accumulation in certain organs.
Considering indisputable correlation between the heavy metal content in foods and aetiology of numerous diseases (WHO 1992;1995), national and international regulations on food quality have lowered the maximum permissible levels (MPC) of toxic metals in human food (Radwan and Salama, 2006).Various permissible limits for certain metals could be found in the literature.For example, Australian Government prescribed maxi-mum cadmium level in all foods at 0.05 mg kg -1 (Walker, 1988), whereas the Australia New Zealand Food Authority (ANZFA, 1997) tolerates 0.1 mg cadmium/kg fresh weight, for root, tuber and leafy vegetables.According to Codex Alimentarius Commission, the permissible limit for lead in cereal and legumes is 0.2 mg kg -1 (CAC 2003).
In the Republic of Serbia, permissible levels of cadmium were set between 0.05 and 0.20 mg/kg, while the permissible limits for lead amount 0.10 or 0.30 mg/kg, in different vegetable species (Official Gazette of Republic of Serbia, 2014).However, MPC for nickel and chromium levels in vegetables have not been defined.The acceptable levels of cadmium and lead are comparable with limits defined by Joint Codex Alimentarius Comission (FAO/WHO 2001), which set maximum values in vegetables at 0.2, 2.3 and 0.3 g/g for cadmium, chromium and lead, respectively.Unlike above cited regulative acts, which prescribe concentration of heavy metals in the vegetable fresh weight, previous Serbian regulative act set allowed levels of lead and cadmium in both fresh and dry vegetables at 1.00 and 3.00, i.e. 0.05 and 0.30 mg/kg, respectively (Official Gazette of Federal Republic of Yugoslavia, 2002).Cadmium content in edible portions of vegetable species surveyed in this work was above 0.30 g/g DM in thirteen samples (Table 3).Vegetables containing above 3.00 g/g DM of lead comprised 30% of all analysed samples (Table 3).Chromium contents were above 2.3 µg/g in six samples: potato (2.54 µg/g), cauliflower (2.69 µg/g), parsnip (4.52 µg/g), and spinach (7.73, 7.13 and 2.81 µg/g).The lowest cadmium and lead contamination was recorded in beet root and potato (cadmium: 0.03 and 0.04 g/g, lead: 0.4 and 0.65 g/g, respectively).

CONCLUSIONS
The results obtained in present work implicate that contamination of vegetables considerably depends on the point of origin.A large number of samples containing high levels of toxic heavy metals, especially of cadmium and lead, impose the ne-cessity for strict regulative guidelines concerning individual vegetable crops production, harvest, handling and storing, in order to diminish the possibility of contamination with respect to the heavy metals.This would increase food quality and safety, and also contribute to preservation of human health.

Table 2 .
Calcium and magnesium contents in edible portions of vegetable species (% DM)

Table 3 .
Cadmium and lead contents in edible portions of vegetable species ( g/g DM)