Pumpkin (Cucurbita moscbata Duch. ex Poir.) is one of

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1 THE ASIAN JOURNAL OF HORTICULTURE Volume 7 Issue 1 June, Research Paper Article history : Received : Revised : Accepted : Arsenic accumulation in pumpkin through contaminated groundwater and varietal evaluation thereof in Gangetic alluvium of West Bengal RAJIB KUNDU, SUKANTA PAL 1 AND APARAJITA MAJUMDER 1 Members of the Research Forum Associate Author : 1 ICAR Niche Area of Excellence, Arsenic Research Laboratory, Directorate of Research, Bidhan Chandra Krishi Viswavidyalaya, Kalyani, NADIA (W.B.) INDIA Author for correspondence : RAJIB KANDU ICAR Niche Area of Excellence, Arsenic Research Laboratory, Directorate of Research, Bidhan Chandra Krishi Viswavidyalaya, Kalyani, NADIA (W.B.) INDIA Abstract : Arsenic is the most hazardous toxic metalloid available in the natural ecological unit and widely distributed in various parts of the world. Near about 50 million people are suffering from its toxicity in Gangetic alluvium of India particularly in West Bengal. The emerging areas of arsenic problem in agricultural system through use of contaminated irrigation water and entry of toxin in vegetables have been largely ignored. Arsenic uptake by plants and its translocation to the edible parts were observed to vary with crops even across the cultivars. With this background, the experiment was laid out in RBD with five selected pumpkin cultivars each replicated four times to study the arsenic accumulation and varietal tolerance of pumpkin and arsenic intake and dietary risk assessment through fruits at different sites of framers fields in Nadia district of West Bengal during summer of and The arsenic accumulation was estimated by using atomic absorption spectrophotometer (AAS) coupled with FIAS The locally grown cultivar was observed to maximum arsenic accumulation with lower yield across the experimental sites, whereas, higher yielding capacity with least arsenic loading in fruit was recorded in cultivars Kali kumro and Ambili. Arsenic accumulation in different parts of pumpkin remained in an order of root > stem > leaf > fruit across the cultivars. Key words : Arsenic, Uptake, Risk assessment, Pumpkin, Vegetable, Cultivar How to cite this article : Kundu, Rajib, Pal, Sukanta and Majumder, Aparajita (2012). Arsenic accumulation in pumpkin through contaminated groundwater and varietal evaluation thereof in Gangetic alluvium of West Bengal, Asian J. Hort., 7(1) : Pumpkin (Cucurbita moscbata Duch. ex Poir.) is one of the most popular summer vegetable, grown all over India, on a commercial scale for its immature and mature fruits. They are used as fresh vegetable, processed food and stock feed, sometimes young tender tops of shoots and leaves are also cooked as vegetable. Because of its high carotene content and good keeping quality, it is considered as a vegetable of immense value (Thamburaj and Singh, 2005). Under intensive cropping, pumpkin is successfully grown as a remunerative vegetable in Gangetic alluvium of West Bengal due to its low-cost of production and long keeping quality, but over the past decade, arsenic contamination in ground water has been reported from these areas (Mitra et al., 2002; Pandey et al., 2002). The word arsenic itself now-a-days sounds as a serious threat and cure to the human race because of its capability of causing terrible health hazards to human being (Srivastava et al., 2001; Rahman, 2002). The World Health Organization (WHO) ranked this calamity as the largest poisoning of a population in history (Smith et al., 2000). Out of 20 countries in different parts of the world where groundwater arsenic contaminations and human suffering have been reported so far, the magnitude is considered to be the maximum in Bangladesh, followed by West Bengal, India (Sanyal, 2005). It causes serious problem in Gangetic alluvium of India and Bangladesh through contamination of groundwater and drinking water (WHO, 2001). The emerging areas of arsenic hazards in agricultural systems through use of contaminated irrigation water and entry of toxin in crops has been largely avoided. Increased arsenic levels in ground water-irrigated soil in West Bengal were well documented by Sanyal and Nasar (2002) and uptake of arsenic by crop plants grown in soils contaminated with high concentration of arsenic HIND AGRICULTURAL RESEARCH AND TRAINING INSTITUTE

2 RAJIB KUNDU, SUKANTA PAL AND APARAJITA MAJUMDER and irrigated with arsenic contaminated groundwater (Dahal et al., 2008; Bhattacharya et al., 2009). This requires an immediate attention as arsenic finds its way into the food web, other than drinking water, accompanied with possible biomagnifications of the food chain (Roychowdhury et al., 2002). Recent studies have shown that the contribution of food-chain towards arsenic pollution in human is many folds greater than that of the drinking water (Roychowdhury et al., 2003; Díaz et al., 2004). Presence of arsenic in vegetable crops and its translocation to the edible parts were observed to vary with crops (Alam et al., 2003; Das et al., 2004) and even among the cultivars of the same crop (Kundu et al., 2010; Rahman et al., 2007). In this context, a thorough understanding of arsenic-plant-soil interaction is necessary in order to examine arsenic uptake by plants grown in soils contaminated with high concentration of arsenic, and irrigated with arsenic contaminated groundwater as well as entry of arsenic in food chain. Rice-based cropping system is the most predominant in this state; however, rainy season rice is not a major problem due to its rainfed cultivation (Duxbury et al., 2003), whereas winter and summer crops, mainly cultivated through irrigated ground water through shallow tube-well, creates a great problem in point of arsenic uptake (Ghosh et al., 2004; Huang et al., 2006). Keeping this view, the present investigation has been undertaken to i) study the arsenic accumulation in pumpkin, known to have a high water requirement and possibilities to accumulate more toxin from the system ii) the varietal tolerance and relative pattern of arsenic uptake by selected cultivars and iii) arsenic intake and dietary risk assessment through pumpkin fruits. RESEARCH METHODS Study area: The experiment was conducted at two different sites of framers fields in village Nonaghata (latitude N, longitude E and altitude 7.8m above sea level), Haringhata Block during the summer season of and under alluvial soil of Nadia district, West Bengal. Many wells contaminated with arsenic have been used as irrigation source in this zone. The experimental site was subtropical humid climate with an average rainfall ranging between 1200 mm to 2500 mm and mean minimum and maximum temperature were 12 0 C and 40 0 C, respectively. Experimental design: The experiment was laid out in Randomized Block Design (RBD) replicated four times having five selected pumpkin cultivars viz., Pusa vikas, Kali kumro, Arka chandan, Ambili and locally grown cultivar. The experiment was conducted for two consecutive years in the same piece of land without disturbing the experimental layout. The experimental soil was silty clay loam in texture having ph 7.21, EC 0.27 dsm -1, organic carbon 0.53%, available nitrogen 167 kg ha -1, available phosphorus 34 kg ha -1 and available potassium 139 kg ha -1 by chemical analysis (Jackson, 1967). Arsenic content in the irrigation water and soil of the experimental remained to the tune of mg l -l to mg l -l and 9.31 mg kg -1 to 9.43 mg kg -1 in site I and site II, respectively. Crop management: The pumpkin, one of the common and remunerative cucurbitaceous vegetable of West Bengal, both fruit and leaves are eaten as fried and cooked, was planted in the end of April with a spacing 3m x 2m and recommended dose of fertilizers (75: 80: 80 kg NPK ha -1, well decomposed farmyard manure was applied as basal along with ½ N and full dose of P and K at the time of final land preparation. Remaining N was to be applied in two equal split doses at the time of vining and at the time of full blooming). The sources of NPK were urea, SSP and MOP. The irrigation was given immediately after germination of seeds, subsequent irrigations were applied once at 5-7 days intervals and arsenic contaminated shallow tubewell (STW) water used as the source of irrigation. Sampling: Irrigation water samples have been collected from the shallow tube well pumps which are used for irrigation in the study area. Prior to sample collection, the pumps were kept running for about minutes in order to get an uniform rate of discharging water. Then the water samples were collected in plastic container and preserved with concentrated HNO 3. The plant samples were taken from the experimental farmer s field at different stages, vegetative and mature stage. First, the plant samples were washed with pure water, to remove the soil particles attached to the plant body, and then with ultrapure de-ionized water. The root, stem, leaves and fruits were collected separately and cut into small pieces. Then the samples were air dried and grinded and kept in the sample container. Arsenic concentration determination: The plant samples were weighted to 1g and digested with tri-acid mixture (HNO 3 : H 2 SO 4 : HClO 4 : 10:1:4, v/v) until a clear solution was obtained. These digested were adequate filtered by using Whatman No. 42 filter paper. 10 ml of the filtrate was taken and in 50 ml volumetric flask, 5 ml of concentrated HCl and 1 ml of mixed reagent [5% KI (w/v) + 5% Ascorbic acid (w/v)] were added to it, kept for 45 minutes to ensure complete reaction and the volume was made up to 50 ml. The total arsenic content in the solution was determined by using atomic absorption spectrophotometer (AAS), Perkin Elmer Analyst 200 coupled with flow injection analysis system (FIAS 400) where the carrier solution was 10% v/v HCl, following Olsen method as described by McLaren et al. (1998). A set of standard solutions of 2.5, 5, 10, and 20 mg l -1 As were used for calibration. Asian J. Hort., 7(1) June, 2012 :

3 ARSENIC ACCUMULATION IN PUMPKIN THROUGH CONTAMINATED GROUNDWATER & VARIETAL EVALUATION Arsenic intake through food and risk assessment: A dietary exposure assessment is the process of estimating how much of a food chemical a population, or population sub group, consumes. Dietary exposure to (or intake of) food chemicals is estimated by combining food consumption data with food chemical concentration data. The process of doing this is called dietary modeling. Dietary exposure = food chemical concentration x food consumption The dietary intake and the risks associated with the arsenic contamination of food materials were assessed in per cent of provisional tolerable weekly intake (PTWI) of the contaminant. Provisional tolerable weekly intake (PTWI) of Arsenic 15 µg/ body weight (900 µg for an adult of 60 kg body weight) as described by FAO and WHO (2000). Statistical analysis: Analysis of variance method (Gomez and Gomez, 1976) was used for statistical analysis. The significance of different sources of variation was tested by error mean square with the help of Fisher s F test at probability level of For comparison of F value and computation of critical different (CD) at 5% level of significance, Fisher and Yates, table were consulted. RESEARCH FINDINGS AND DISCUSSION The results obtained from the present investigation as well as relevant discussion have been summarised under following sub heads: Arsenic accumulation pattern: The arsenic concentration in the different parts of the pumpkin plants (Fig.1) reflects that the maximum accumulation was observed in the roots as compared to any other plant parts. Arsenic accumulation in different parts of pumpkin remained in an order of root > stem > leaf > fruit across the tested cultivars, across the experimental sites, irrespective of the time of sampling (vegetative stage or maturity). Comparison of arsenic accumulation of different plant parts Fig.1 : Arsenic accumulation pattern of pumpkin (average) at maturity of pumpkin clearly showed that translocation of arsenic in edible part is relatively lower than the any other plant parts. In a similar study, significantly lower amount of uptake and translocation of this toxic metalloid in the rice grain compared with the uptake in the rice straw and root parts have been reported by Liu et al. (2007). Such findings are quite consistent with general observations like in general, the distribution of arsenic in plant parts is found to be in the order: below ground parts > areal parts (Sanyal, 2005). The arsenic concentration was found to vary between different parts of the plants. Higher amount of arsenic was reported to accumulate in the root of the plant as compared with other parts (Norra et al., 2005; Rahman et al., 2007; Bhattacharya et al., 2009). Again, arsenic concentration of different plant parts of pumpkin varies over the experimental sites; whereas, site-ii recorded the maximum values across the plant parts and cultivars. It is possibly due to the reason that the different source of irrigation water and cultivated soil. Similar findings also reported by Bhattacharya et al. (2010), the arsenic-contaminated irrigation water and soil considerably influenced in the accumulation of arsenic in different crops and vegetables. Arsenic accumulation at vegetative stage: At the vegetative stage of pumpkin (60 days after sowing), arsenic accumulation by different plant parts varied significantly across the tested cultivars (Table 1). The maximum arsenic accumulation in roots was observed in the cultivar Arka chandan (2.14 and 2.45 mg kg -1 in site-i and site-ii, respectively) and it was statistically at par with the cultivar Pusa vikas and Locally grown cultivar, whereas, least arsenic loading by roots recorded under the cultivar Kali kumro (1.37 and 1.61 mg kg -1 in site-i and site-ii, respectively) and it had no significant difference with the cultivar Ambili among the all tested cultivars. In case of pumpkin stem, locally grown cultivar (1.69 and 1.91 mg kg -1 in site-i and site-ii, respectively) accumulated maximum amount of this toxic metalloid. Across tested cultivars, Pusa vikas (1.30 mg kg -1 ) and Arka chandan (1.55 mg kg -1 ) loaded least amount of arsenic in its stem in site-i and site-ii, respectively. Finally in leaves of the locally grown cultivar (1.21 and 1.35 mg kg -1 in site-i and site-ii, respectively) accumulated the significantly highest amount of arsenic, whereas, the least arsenic loading was showed under the cultivar Kali kumro (0.73 and 0.97 mg kg -1 in site-i and site-ii, respectively) followed by the cultivar Ambili. Arsenic accumulation at maturity: At the maturity of pumpkin ( days after sowing, depending upon cultivars), the degree of arsenic loading also differed significantly across the cultivars (Table 2). At this stage, arsenic accumulation in below ground parts of pumpkin that means roots recorded the more or less similar trends as followed by its vegetative stage. The cultivar Pusa vikas, showed the significantly highest amount of arsenic in its stem Asian J. Hort., 7(1) June, 2012 :

4 RAJIB KUNDU, SUKANTA PAL AND APARAJITA MAJUMDER Table 1 : Arsenic accumulation (mg kg -1 ) by different plant parts of pumpkin cultivars at vegetative stages (pooled data) Root Stem Leaf Cultivar Site I Site II Site I Site II Site I Site II Pusa vikas Kali kumro Arka chandan Ambili Local cultivar S E. (±) C.D. (P=0.05) Arsenic content in the irrigation water and soil of the experimental remained to the tune of mg l -l to mg l -l and 9.31 mg kg -1 to 9.43 mg kg -1 in site I and site II, respectively. Table 2 : Arsenic accumulation (mg kg -1 ) by different plant parts and yield (kg ha -1 ) of pumpkin cultivars at maturity (pooled data) Arsenic accumulation (mg kg -1 ) Yield Cultivar Root Stem Leaf Fruit (kg ha -1 ) Site I Site II Site I Site II Site I Site II Site I Site II Site I Site II Pusa vikas Kali kumro Arka chandan Ambili Local cultivar S Em (±) CD (P=0.05) Arsenic content in the irrigation water and soil of the experimental remained to the tune of mg l -l to mg l -l and 9.31 mg kg -1 to 9.43 mg kg -1 in site I and site II, respectively and leaves (2.11 and 2.56 mg kg -1 by stem and 1.72 and 2.03 mg kg -1 by leaves in site-i and site-ii, respectively), whereas, among the all cultivars, least arsenic loading by stem and leaves were observed under the cultivar Kali kumro (1.77 and 1.90 mg kg -1 by stem and 1.07 and 1.41 mg kg -1 by leaves in site-i and site-ii, respectively) closely followed by the cultivar Ambili. Finally in the fruits of the cultivar Arka chandan (1.39 and 1.47 mg kg -1 in site-i and site-ii, respectively) loaded significantly highest concentration of arsenic, whereas, the significantly least accumulation of this toxic metalloid was found under the cultivar Kali kumro (0.42 and 0.55 mg kg -1 in site-i and site-ii, respectively) followed by the cultivar Ambili across the tested cultivars. The arsenic accumulation levels was observed to increase with the advancement of growth, when such increase in arsenic load was maximally manifested in pumpkin root. obtained in locally grown cultivar (2451 and 2513 kg ha -1 in site-i and site-ii, respectively), followed by the cultivar Arka chandan (2879 and 2785 kg ha -1 in site-i and site-ii, respectively). The arsenic uptake (g ha -1 ) by fruit of the selected cultivars in both the experimental sites was presented in Fig. 2. The arsenic uptake by fruit was observed to increase in the site-ii, across the tested cultivars, when the maximum uptake recorded by the cultivars Arka chandan across the experimental sites (40.02 and g ha - 1in site-i and site-ii, Yield and arsenic uptake: Fruit yield (kg ha -1 ) of pumpkin was recorded at the time of harvest, and differed significantly with the different cultivars (Table 2) and did not reasonable vary over experimental sites. Across the tested cultivars, Ambili produced the maximum fruit yield (3517 and 3633 kg ha -1 in site-i and site-ii, respectively) which had no significant difference with the cultivar Kali kumro (3353 and 3471 kg ha -1 in site-i and site-ii, respectively). On the other hand, significantly least fruit yield Fig. 2 : Arsenic uptake (g ha -1 ) by different cultivars of pumpkin at maturity Asian J. Hort., 7(1) June, 2012 :

5 ARSENIC ACCUMULATION IN PUMPKIN THROUGH CONTAMINATED GROUNDWATER & VARIETAL EVALUATION THEREOF IN GANGETIC ALLUVIUM OF WEST BENGAL Table 3 : Arsenic intake and dietary risk assessment through pumpkin Food item Weekly consumption (g) Max. total as conc. (µg.kg -1 ) Max. weekly total As intake (µg) Max. weekly inorganic as intake (µg) Per cent PTWI Pumpkin (Site-I) Pumpkin (Site-II) Rice** 10965** 357** 3911** 1956** 217** **Final Report of Total diet study in West Bengal (2010) respectively), followed by Pusa vikas and locally grown cultivar. The cultivar Kali kumro showed the least arsenic uptake across the experimental sites (40.02 and g ha -1 in site-i and site-ii, respectively), followed by the cultivar Ambili. The higher uptake of arsenic in fruits of Arka chandan may be responsible for the lower observed fruit yields of the cultivar, although there may be several other factors controlling yields. Arsenic intake and dietary risk assessment: In risk assessment, exposure estimates are compared with reference health standards, where available, to assess the potential risk to health associated with changes to the food supply. The dietary intake and the risks associated with the arsenic contamination of food materials were assessed in per cent of provisional tolerable weekly intake (PTWI) of the contaminant. Provisional tolerable weekly intake (PTWI) of inorganic arsenic 15 µg/ body weight (900 µg for an adult of 60 kg body weight) as described by FAO and WHO (2000). It was also reported that in fruits and vegetables, inorganic arsenic accounted for approximately one-half of the total arsenic (Schoof et al., 1999). Ingestion of significant proportion of inorganic arsenic intake (1596 µg/week/adult and 1688 µg/ week/adult in site-i and site-ii, respectively) through consumption of pumpkin was observed through the present investigation which poses a risk of 177 per cent and 188 per cent (in site-i and site-ii, respectively) of PTWI through dietary intake of inorganic arsenic which is much beyond the WHO permissible limit but still pretty low as compared to such risk of exposures to contaminated rice (Table 3). This field based experiment clearly revealed that the continuous application of arsenic contaminated irrigation water increased with the age of crops, which may be the possible reason for increasing level of arsenic loading according to the age of the pumpkin plants. Again, Arsenic intake and dietary risk assessment through pumpkin varied over the source of irrigation water and cultivated soil. Similar findings was also reported by Bhattacharya et al. (2010), the arseniccontaminated irrigation water and soil considerably influenced in the accumulation of arsenic in different crops and vegetables. The cultivars Kali kumro and Ambili accumulated least amount of arsenic in their fruits with maximum yielding capacity across the cultivars, whereas, the locally grown cultivar loaded moderate amount of arsenic in its fruit and produced least fruit yield. On the other hand, the cultivar Arka chandan showed highest recovery of this toxic metalloid with lower yielding capacity. So, yield of pumpkin may not affected by the level of arsenic concentration in its environment but the quality of fruits and aerial parts (use of leafy vegetable) is highly affected. Preference should be given for those cultivars having higher production ability with least accumulation of arsenic in its edible parts. Chaturbedi (2006), Rahman et al. (2007) and Kundu et al. (2010) reported that arsenic concentration appearing toxicity was widely varied with plant genotypes probably due to varietal differences in arsenic translocation and phyto-extraction or phytomorphological potential of the varieties. Present observations also support the fact when cultivar Kali kumro was found to accumulate least arsenic and cultivar Arka chandan the highest under same growing condition. Conclusion: In Eastern India, particularly West Bengal pumpkin is a commonly consumed cucurbitaceous vegetable. Though arsenic accumulation in pumpkin fruits relatively was lower than the other plant parts, but the leaves and top of the shoots are also used as cooked vegetable, so, there is need for appropriate agricultural management practices towards minimize the arsenic loading from pumpkin. The accumulation of toxic heavy metals like arsenic remained also significantly higher in pumpkin and other vegetables than cereals and other crops and also varies across the cultivars. Selection of the appropriate cultivars of pumpkin which loaded lesser amount of arsenic without hampering its production rate in arsenic affected belt have an immense ecological importance, as it might have a beneficial role towards mankind. Pumpkin accumulates much higher arsenic in edible portion than rice, still the risk associated with dietary exposure to arsenic contaminated pumpkin is less than rice which is due to far less weekly consumption of the former though the level of arsenic ingestion through pumpkin far exceeded the WHO permissible limit for clinically safe food item. Such food items are largely ignored in toxicological investigation and demand due attention of the researchers. Acknowledgement: We are thankful for financial assistance by ICAR Niche Area of Excellence Arsenic Management Options including Organic Agricultural Systems in West Bengal, Directorate of Research, BCKV, Kalyani, Nadia, West Bengal, India. Asian J. Hort., 7(1) June, 2012 :

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