Health Risk Assessment of Heavy Metals Through the Consumption of Vegetables in the National Capital Region, India

This study aimed to evaluate the heavy metal contamination in the vegetables growing in the national region and to assess the health risk in human resulting from consumption of these vegetables. A total of 99 vegetable samples were collected from the selected study area during the winter season and were analyzed for heavy metals contamination by atomic absorption spectrophotometer. The relative abundance of heavy metal in the vegetable samples was in the following order: Fe>Al>Mn>Zn>Cu>Cr>Ni>Pb>Cd>Co. The mean concentration of Al, Cd, Co, Cr, Cu, Fe, Mn, Ni, Pb, and Zn, in the vegetables was 158.01 mg kg -1 dw -1 , 0.23 mg kg -1 dw -1 , 0.04 mg kg -1 dw -1 , 3.70 mg kg -1 dw -1 7.82 mg kg -1 dw -1 , 297.87 mg kg -1 dw -1 , 39.81 mg kg -1 dw -1 , 1.78 mg kg -1 dw -1 , 0.52 mg kg -1 dw -1 , and 32.21 mg kg -1 dw -1 for respectively. The statistical analysis supported the formation of two primary clusters Al-Fe-Pb and Cu-Zn, indicating their common source of origin. Most of the vegetable samples exceeded the permissible limit of heavy metals prescribed by the Food and agricultural organization/ World health organization (FAO/WHO) standards. The total target hazard quotient was greater than 1 for all types of vegetables, indicating appreciable health risk due to the consumption of these vegetables in the study area. a health risk due to the high contents of Al, Cr, Fe, and Pb. The target hazard quotient for aluminium, copper, iron, manganese, and cadmium was greater than 1 for more than 50% of the collected vegetable sample. The total target hazard (TTHQ) value was greater than one in all the vegetables suggesting potential non-carcinogenic risk due to the presence of multi-metal in vegetable samples. TTHQ value for adults and children was 23.45 and 26.95 for the mustard which is 23 and 27 times higher than the threshold value. Thus, we can say that the consumption of mustard and other leafy vegetables are not safe in the study areas.

1 gram dried sample of vegetable was digested by adding 15 ml of a tri-acid mixture (HNO3, H2SO4, and HClO4 in 5:1:1 ratio) at 80 o C until a transparent solution was obtained. After cooling, the digested sample was finally maintained to 50 ml with double distilled water and was filtered using Whatman No. 42 filter paper (Sharma et al. 2008). Concentrations of Al, Cd, Cu, Co, Cr, Fe, Mn, Ni, Pb, and Zn in the filtrate of digested vegetable samples were estimated by using an atomic absorption spectrophotometer (AAS) (Model Lab India 8000A). The instrument was calibrated with a blank and reference standard of heavy metals solution of different concentrations ranging between 0.02-10 ppm at standard conditions (r 2 =0.999). Blank and standard were run after every 10 samples on the instrument. All the chemicals and standards used were of Merck Company.

Data analysis
The Statistical Program for Social Sciences (SPSS version 22.0) was used to find out the Pearson's correlation coefficient (r), principal component analysis (PCA), and hierarchical cluster analysis (CA) to understand the correlation between heavy metals.

Health risk assessment
Target hazard quotient (THQ) Health risk can be estimated by calculating the target hazard quotient (THQ) caused by heavy metals exposure by ingestion. The equation used for estimating the THQ for non-carcinogenic risk are given below (Proshad et al. 2019). (1) The total target hazard quotient for all studied metals in a vegetable can be calculated as: (2) Where -Exposure frequency (365 days/year for Non-carcinogenic risk and 350 days/year for carcinogenic risk), -Exposure duration (70 Years), -Ingestion rate of soil (345 g /person /day for adult and 232 g /person /day for children), -Concentration of the contaminant in vegetable (mg/kg.dw), -Oral reference dose of heavy metals (mg/kg/day), -Average body weight (55.9 kg for adult and 32.7 kg for children), -Averaging time (365 days/year ×number of exposure years (70 years)), Oral reference dose values (mg kg -1 day -1 ) for Al, Cd, Co, Cr, Cu, Fe, Mn, Ni, Pb and Zn were 1, 0.001, 0.043, 1.5, 0.040, 0.7, 0.014,0.02, 0.0035, and 0.300, respectively (IRIS, 1995;US-EPA, 2006, 2008Verma et al. 2015;Antoine et al. 2017).

Metal pollution index (MPI)
The Metal pollution index was used to assess the total metal content for a vegetable by calculating the geometric mean concentrations of all the studied metals in the vegetable as per equation (4) (Usero et al. 1997). (4) Where, : Concentration of n th metal in the sample

Result and discussion:
The relative abundance of heavy metal in the vegetable samples was in the following order: Fe > Al > Mn > Zn > Cu > Cr > Ni > Pb > Cd > Co. Metal concentrations were compared to the permissible limit of food and agricultural organization (FAO)/ World health organization (WHO) standards, as well as, the concentrations reported in earlier studies (Table 1). Mean concentration of metals in different types of vegetables collected from NCR, India is shown in fig. 2 (a-e).

Aluminium
A significant amount of aluminium was detected in all the vegetable samples with minimum and maximum value observed in pumpkin (13.85 mg/kg) and mustard (333.88 mg/kg). We could not found permissible limit for aluminium exposure through vegetable consumption in literature. Average concentration of aluminum in the studied vegetable samples was in the decreasing order of mustard

Chromium
All the studied leafy vegetables exceeded the permissible limit of chromium (2.3 mg/kg) set by FAO/WHO, except carrot leaves, chenopodium, and onion leaves. Among root vegetables, radish and turnip exceeded the permissible limits of chromium while in fruity vegetable only brinjal exceeded the permissible limit. In potato also the chromium concentration was higher than the permissible limit of WHO (Khan et al. 2015b). The value of chromium was the minimum in onion bulbs (1.37 mg/kg) and the highest was found in mustard (6.38 mg/kg). The mean concentration of Cr in the vegetables was in decreasing order of mustard > radish >

Copper
Copper concentration was found to be lowest in cabbage (3.02 mg/kg) and the highest in pumpkin (20.4 mg/kg

Manganese
Manganese concentration was found to be highest in spinach (86.99 mg/kg) and the lowest concentration in potato (8.7 mg/kg

Nickel
Nickel acts as a micronutrient; it catalyzes nitrogen metabolism and is a constituent of the urease enzyme molecule (Nagajyoti et al. 2010). The mean concentration of Nickel exceeded the permissible limit (1.5 mg/kg) in all the collected leafy vegetables except cabbage, carrot leaves, and onion leaves (Nisa et al. 2020). The mean concentrations were above the permissible limit in pumpkin, round gourd, potato, and onion bulb. The lowest mean concentration was found in beetroot (0.8 mg/kg) and the highest was in chenopodium (6.13 mg/kg). Concentration of Ni in the vegetables was in decreasing order of chenopodium > pumpkin > mustard > onion bulb > round guard > spinach > potato > fenugreek > beet leaves > coriander leaves > brinjal > carrot leaves > carrot > cauliflower > radish > cabbage > onion leaves > turnip > beetroot. The average concentration of Ni reported in mustard in the present study is two times higher than the concentration reported in wastewater irrigated mustard from Varanasi, India (

Lead
Very high concentrations of lead were found in mustard, fenugreek, coriander leaves, and spinach, whereas in cauliflower, radish, brinjal, potato, and onion bulb it also exceeded the permissible limit of FAO/WHO standards. Permissible limit of Pb for leafy vegetables is 0.3 mg/kg and 0.1 mg.kg for tuberous and bulb vegetables ( FAO/WHO, 2017). The minimum average concentration of lead was found in onion bulbs (0.07 mg/kg) while the maximum was in mustard (2.4 mg/kg). Pb concentration in the studied vegetable samples was followed the order as chenopodium > pumpkin > mustard > onion bulb > round guard > spinach > potato > fenugreek > beet leaves >coriander leaves > brinjal > carrot leaves > carrot > cauliflower > radish > cabbage > onion leaves > turnip > beetroot. Pb concentration in spinach, potatoes, cabbage, and cauliflower in present study was higher than the concentration reported by (Bo et al. 2009). Prenatal exposure to lead causes abnormality in offspring, reduces the Ca 2+ ion absorption in the body and affects the soft tissues like the brain, heart, kidney, and central nervous system; and is also known as mutagen, teratogen, and carcinogens (Bhowmik

Zinc
Mean concentration of zinc was found minimum in onion leaves (15.55 mg/kg) and the maximum was in Pumpkin (70.35 mg/kg).
Pumpkin exceeded the permissible limits of Zn (60 mg kg -1 ) prescribed by FAO/WHO standards (Sharma et al. 2008). Zn concentration in the studied vegetable samples followed the order as pumpkin > round guard > chenopodium > coriander leaves > mustard > spinach > fenugreek > beet leaves > cauliflower > beetroot > brinjal > cabbage > turnip > onion bulb > radish > carrot leaves > carrot > potato > onion leaves. The average concentration of Zn reported in radish and mustard was 2 times higher than the concentration reported in Varanasi, India (Verma et al. 2015), and in potato it was 4 times higher than the concentration reported in India and China (Bo et al. 2009;Verma et al. 2015). Zn concentration in our study is higher than the concentration reported in cabbage be seen that most of the leafy vegetables are grouped together like mustard, fenugreek, coriander leaves, carrot leaves, and fruit vegetables that are pumpkin and round guard were placed in the same group ( fig. 3b).

Metal Pollution Index (MPI)
A metal pollution index is an effective tool for assessing metal pollution in vegetables. Among all collected different vegetable samples, the minimum value of MPI was found in brinjal while maximum in carrot leaves ( fig. 5). MPI in the different collected vegetables from NCR, India followed the decreasing order as carrot leaves, mustard, spinach, chenopodium, beet leaves, turnip, potato, round guard, coriander leaves, carrot, beetroot, fenugreek, cauliflower, onion leaves, pumpkin, onion bulb, cabbage, radish and brinjal with MPI values as 11. 32, 10.32, 9.12, 8.55, 8.07, 7.8, 7.68, 7.04, 6.41,6.06, 6.05, 5.9, 5.29, 4.77, 4.55, 3.6, 3.59, 3.43 and 3.30, respectively. Higher values of MPI for mustard, spinach, and chenopodium pointed towards high potential health risks due to the consumption of these leafy vegetables. MPI among vegetables followed the decreasing order as Fe> Al> Zn> Mn> Cu> Cr> Ni> Pb> Co> Cd. Thus, the highest metal pollution in the vegetables is due to iron and the lowest is due to cadmium.

Health risk assessment
The target hazard quotient (THQ) for non-carcinogenic risk of the studied metal from consuming vegetables for adults and children are shown in the fig. 6 (a-e). The estimation of THQ provides an indication of the potential risk probability in a population exposed to the toxicant. Among the studied metals THQ value of Al exceeded the threshold value of one for adults and children in mustard, fenugreek, coriander leaves, spinach, carrot leaves, radish, carrot, and turnip while THQ was greater than 1 for beetroot in children only. THQ was greater than 1 for Cd in adults and children both by the use of mustard, beet leaves, spinach, and carrot leaves, while it was greater than 1 for children only due to the consumption of coriander leaves and chenopodium. THQ was less than 1 for Cobalt in all the vegetable samples. THQ for Cr was greater than 1 for beetroot and carrot in adults and children. THQ value for Ni and Pb was >1 in Chenopodium and mustard for adults and children. THQ due to Zn was greater than 1 in radish, carrot, turnip, pumpkin, and round guard for adults and children while it was greater than 1 in children in case of coriander leaves, chenopodium, and beetroot.
THQ was >1 for Cu, Fe, and Mn in Mustard, fenugreek, beet leaves, coriander leaves, spinach, carrot leaves, and chenopodium for adults and children. THQ was >1 for Cu in radish and onion bulb and for Fe in onion leaves, brinjal, and potato for children. Cu showed higher THQ values for round guard and potato, and Mn has THQ> 1 for brinjal and pumpkin. The total target hazard quotient expresses the non-carcinogen effect of multi-metals in a vegetable sample. The highest TTHQ value was observed for mustard in adults (23.45) and children (26.96). The high value indicating the potential non-carcinogen may occur due to the consumption of mustard in the population. TTHQ value due to consumption of vegetables in the study area was in the following decreasing order mustard> spinach> coriander> carrot leaves> chenopodium> fenugreek>beet leaves> pumpkin> turnip> brinjal> round guard> carrot> beetroot> cauliflower> radish> onion bulb> cabbage> potato> onion leaves. The hazard index (HI) is the sum of individual THQ for vegetables. In the present study, HI was 140.85 (>1) for adults and 170.11 (>1) for children. Thus potential health risks from exposure to the vegetable are of great concern in the study area. The result of present study is consistent with the result of Hussain and Qureshi (2019).

Conclusion
The present study revealed that Fe, Co, Pb, Ni, Cd, and Zn have crossed the permissible limit set by FAO/WHO in most of the vegetable samples, indicating potential health hazards to humans due to consumption of these vegetables. Fe was the most prominent metal found in the vegetables followed by Al, Mn, and Zn while Co was the least reported metal in the present study. Multivariate analysis showed that Pb, Cr, Zn, Cu, Co, Cd, and Ni in the vegetable samples were mainly contributed by industrial activities. High metal pollution indices values (MPI) for leafy vegetables are pointing towards their high tendency of metal accumulation. Mustard consumption may pose a health risk due to the high contents of Al, Cr, Fe, and Pb. The target hazard quotient for aluminium, copper, iron, manganese, and cadmium was greater than 1 for more than 50% of the collected vegetable sample. The total target hazard (TTHQ) value was greater than one in all the vegetables suggesting potential non-carcinogenic risk due to the presence of multi-metal in vegetable samples. TTHQ value for adults and children was 23.45 and 26.95 for the mustard which is 23 and 27 times higher than the threshold value. Thus, we can say that the consumption of mustard and other leafy vegetables are not safe in the study areas.

Ethical approval and consent to participate
Not Applicable

Consent for Publication
Not Applicable

Availability of data and materials
All data generated or analyzed during this study are available from first author or corresponding author on request.

Competing interests
The authors declare they have no competing interests.

Funding
This work was not supported by any funding however facilities provided in lab of CEEES, DCR university of Science and Technology, Murthal are highly appreciated.

Authors' contributions
JR provided and analysed data for the present study, SC helped in interpretation and writing of manuscript and TA provided consultation advice for project. All authors read and approved the final manuscript.