ANALYSIS OF POTENTIAL CARCINOGENIC CONTAMINANTS IN GRAIN AND VEGETABLES WITH FOCUS ON HEAVY METALS

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1 ANALYSIS OF POTENTIAL CARCINOGENIC CONTAMINANTS IN GRAIN AND VEGETABLES WITH FOCUS ON HEAVY METALS N. Belc, V. Ionescu, M. Ionescu, E. Iorga and G. Mustatea National R&D Institute for Food Bioresources, 6 Dinu Vintila Street, , Bucharest, Romania

2 INTRODUCTION Heavy metals are significant environmental pollutants. The main cause of emissions of heavy metals into the environment is represented by anthropogenic activities. Heavy metals such as arsenic, cadmium, chromium and lead are harmful to humans and they are known to be potential carcinogens. However, the mechanism underlying heavy metal - induced cancer remains unclear. The main objective of this study was to measure the levels of trace elements (lead, cadmium, chromium and arsenic) knowing to have carcinogenic potential in 10 plant species (vegetables and grains), and to evaluate them according to current guidelines in European Union and worldwide.

3 INTRODUCTION As & As compounds Cd & Cd compounds Cr(VI) compounds Pb compounds (inorganic) IARC GHS NTP ACGIH EU Group 1 Cat. 1A Known A1 Cat. 1 Group 2A Group 2B Group 3 Group 4 Cat. 1B Reasonably suspected A2 Cat. 2 Cat. 2 A3 Cat. 3 IARC International Agency for Research on Cancer (part of WHO); GHS Globally Harmonized System; NTP U.S. National Toxicology Program; ACGIH American Conference of Governmental Industrial Hygienists; EU European Union A4 A5 Group 1 The agent is definitely carcinogenic to humans. Group 2A The agent is probably carcinogenic to humans.

SAMPLES The sampling was done in the most unfavorable

heavy traffic and, in some cases, close to the garbagge

1 8 14 samples 3, 5 2 samples 6 2 samples 5, 6 2

4 SAMPLES The sampling was done in the most unfavorable areas, sorrounding Bucharest - very crowded, with very heavy traffic and, in some cases, close to the garbagge collection areas samples 3, 5 2 samples 6 2 samples 5, 6 2 samples 1, 6, 8 4 samples 4, 5, 6 6 samples 3, 5, 7 19 samples 4 1 sample 4, 6, 7 3 samples 6 1 sample

SAMPLE ANALYSIS 54 samples from 8 areas were analyzed (Ilfov

samples 6 salad samples 3 radishes samples 2 cabbage samples 2 green

were collected in 2 different periods (May and July 2014) from the

5 SAMPLE ANALYSIS 54 samples from 8 areas were analyzed (Ilfov District): 20 cereal samples (14 wheat, 4 barley; 2 oat) 19 tomatoes samples 6 salad samples 3 radishes samples 2 cabbage samples 2 green onion samples 1 spinach sample 1 green garlic sample Some samples were collected in 2 different periods (May and July 2014) from the same places, using GPS coordinates. Samples were dried before being analyzed.

MATERIALS AND METHODS Samples were weighted into acid-washed PTFE tubes, combined with 3 ml of ultrapure nitric acid (Merck) and 2 ml of hydrogen peroxide solution 30%

After cooling, each sample was transferred in clean volumetric flasks and the volumes were increased to 50 ml with ultra pure water (Milli-Q).

6 MATERIALS AND METHODS Samples were weighted into acid-washed PTFE tubes, combined with 3 ml of ultrapure nitric acid (Merck) and 2 ml of hydrogen peroxide solution 30% (Merck). Heating in a microwave digestion system (MWS-2 Berghof). Microwave digestion steps: 145 ⁰C for 10 minutes, 160 ⁰C for 10 minutes and 190 ⁰C for 20 minutes. After cooling, each sample was transferred in clean volumetric flasks and the volumes were increased to 50 ml with ultra pure water (Milli-Q). All samples were analyzed for Pb, Cd, Cr and As content using a graphitefurnace atomic absorption spectrometer (AAnalyst 600, Perkin Elmer). Microwave oven (MWS-2 Berghof) GF-AAS (AAnalyst 600, Perkin Elmer)

7 RESULTS AND DISCUSSION Parameters of calibration curves Element Domain Correlation coefficient Slope Detection limit Pb 0 80 μg/l μg/l Cd 0 5 μg/l μg/l Cr 0 40 μg/l μg/l As 0 10 μg/l μg/l Limits for heavy metals content in EU, China and Australia Product Pb (mg/kg) Cd (mg/kg) Cr (mg/kg) As (mg/kg) EU Cod GB Aus EU Cod GB Aus EU Cod GB Aus EU Cod GB Aus Wheat Oat Barley Radish Onion Garlic Tomato Cabbage Salad Spinach EU Regulation (EC) No. 1881/2006; Cod - Codex Alimentarius Commission - Codex STAN 193: ; GB - GB 2762 Standard China; Aus - Food Standards Australia and New Zealand - Standard 1.4.1

Heavy metals content of grain samples Sample no. Sample Sampling area Collecting period Humidity (%) Pb Heavy metals content (mg/kg wet weight) EU EU Cd limit limit Cr As < 0.0002 0.23 0.

03 5 Wheat (4) 2 05.2014 82.2 < 0.002 < 0.0002 0.20 0.02 6 Wheat (4) 2 07.2014 46.4 < 0.002 < 0.0002 0.2 < 0.0013 < 0.001 7 Wheat (10) 5 05.2014 82.0 < 0.002 < 0.0002 0.01 0.04 8 Wheat (15) 4 05.

2014 76.1 0.68 < 0.0002 0.12 0.02 13 Barley (25) 6 07.2014 10.8 < 0.002 < 0.0002 0.01 0.01 14 Wheat (26) 6 05.2014 78.8 < 0.002 < 0.0002 0.18 0.01 0.2 15 Wheat (26) 6 07.2014 50.2 < 0.002 < 0.0002 < 0.

8 Heavy metals content of grain samples Sample no. Sample Sampling area Collecting period Humidity (%) Pb Heavy metals content (mg/kg wet weight) EU EU Cd limit limit Cr As < Wheat (1) < Wheat (1) < < < Barley (2) < < Wheat (3) < < Wheat (4) < < Wheat (4) < < < < Wheat (10) < < Wheat (15) < < < Wheat (16) < < < Oat (24) < Oat (24) < < < Barley (25) < Barley (25) < < Wheat (26) < < Wheat (26) < < < < Barley (27) < < Wheat (28) < < Wheat (29) < < Wheat (29) < < < Wheat (32) < <

9 Lead content of all cereal samples is bellow 0.2 mg/kg limit stated by Regulation (EC) No. 1881/2006. Cadmium content of all cereal samples is bellow 0.2 mg/kg limit (for wheat) and 0.1 mg/kg limit (for barley and oat) stated by Regulation (EC) No. 1881/2006. For Chromium and Arsenic there is no imposed limit in EU. The level of these two metals in all tested cereal samples is bellow 1.5 mg/kg (for Chromium) and 0.2 mg/kg (for Arsenic), limits stated by GB 2762 Standard (China). According with the heavy metal levels in analyzed cereals the descendent order was: Wheat: Cr t > As > Pb > Cd Oat: Cr t > As > Pb > Cd Barley: Pb > Cr t > As > Cd

10 Heavy metals content of tomatoes samples Sample no. Sample Sampling area Collecting period Humidity (%) Pb Heavy metals content (mg/kg wet weight) EU EU Cd limit limit Cr As Tomato (5) - leaves Tomato (5) unriped fruits < < Tomato (5) riped fruits < < < Tomato (5) peel < < < < Tomato (5) - juice < < < Tomato (6) unriped fruits < < Tomato (6) riped fruits < < < Tomato (7) leaves < < Tomato (7) riped fruits < Tomato (8) - leaves < < Tomato (8) unriped fruits < < Tomato (8) riped fruits < < < Tomato (8) peel < < < < Tomato (8) - juice < < < Tomato (9) leaves < < Tomato (9) unriped fruits < < Tomato (17) leaves & < < < fruits 18 Tomato (30) leaves < < Tomato (30) unripe fruits < < <

11 Lead and Cadmium content of all tomatoe samples is bellow 0.1 mg/kg and 0.05 mg/kg, respectively, limits stated by Regulation (EC) No. 1881/2006. For Chromium and Arsenic there is no imposed limit in EU. The level of these two metals in all tested cereal samples is bellow 1.0 mg/kg (for Chromium) and 0.5 mg/kg (for Arsenic), limits stated by GB 2762 Standard (China). Heavy metals content in leaves is higher than in fruit tomatoes. The descendent order of heavy metals level in analyzed tomatoes was: Leaves: Pb > Cr t > As > Cd Unriped tomatoes: Cr t > As > Pb > Cd Riped tomatoes: As > Cr t > Pb > Cd

Heavy metals content of vegetables samples Sample no.

3 0.2 1 Salad (11) 05.2014 5 93.6 0.02 2 Cabbage (12) 05.2014 5 91.0 < 0.002 < 0.0002 0.01 0.04 3 Salad (13) 05.2014 5 90.2 < 0.002 0.02 < 0.0013 0.

03 6 Cabbage (19) 05.2014 3 87.7 < 0.002 0.3 < 0.0002 0.2 < 0.0013 0.01 7 Spinach (20) 05.2014 6 91.3 < 0.002 0.01 0.02 0.01 8 Radish (21) 05.

2014 6 92.3 0.2 0.01 0.09 0.04 0.3 0.2 11 Salad (23) 05.2014 6 93.4 0.04 < 0.0002 0.37 0.03 12 Radish (31) 05.2014 7 95.3 < 0.002 < 0.

12 Heavy metals content of vegetables samples Sample no. Sample Sampling period Sampling area Humidity (%) Pb Heavy metals content (mg/kg wet weight) EU EU Cd limit limit Cr As Salad (11) Cabbage (12) < < Salad (13) < < Green onion (14) < < < Salad (18) < < Cabbage (19) < < < Spinach (20) < Radish (21) < < Green onion (22) < < Salad (23) Salad (23) < Radish (31) < < < Radish (33) < < < Salad (34) < Green garlic (35) < <

13 Lead content of radish, onion and garlic samples is bellow 0.1 mg/kg limit and the lead content of cabbage, salad and spinach samples is bellow 0.3 mg/kg limit, stated by Regulation (EC) No. 1881/2006. Cadmium content for all tested vegetables samples is bellow limits stated by Regulation (EC) No. 1881/2006: 0.1 mg/kg for radish, 0.05 mg/kg for onion and garlic and 0.2 mg/kg for cabbage, salad and spinach. For Chromium and Arsenic there is no imposed limit in EU. The level of these two metals in all tested vegetables samples is bellow 1.0 mg/kg (for Chromium in radish, onion and garlic) and 0.5 mg/kg (for Arsenic and Chromium in cabbage, salad and spinach), limits stated by GB 2762 Standard (China). The descendent order of heavy metals level in the analyzed vegetables was: Salad: Cr t > Pb > As > Cd Green onion: As > Cr t > Pb > Cd Radish: Cr t > As > Pb > Cd Cabbage: As > Cr t > Pb > Cd Spinach: Cr t > As = Cd > Pb Green garlic: As > Cd > Pb > Cr t

14 CONCLUSSIONS (I) Heavy metals content (lead, cadmium, total chromium and arsenic) was determined from 10 plant species (cereals: wheat, oat and barley and vegetables: tomatoes, radish, cabbage, green onion, spinach, salad and green garlic) collected from 8 areas located sorrounding Bucharest. Samples were collected from the most unfavorable area considering a high potential of chemical contamination. The sampling was done in two different period of the year (May and July) from the same place, using GPS coordinates. Samples were analyzed after drying and microwave digestion using atomic absorption spectrometry technique (GF-AAS).

15 CONCLUSSIONS (II) Heavy metals contamination of cereals and vegetables cannot be underestimated as these foodstuffs are very important components of human diet. Vegetables take up metals by absorbing them from contaminated soils, as well as from deposits on different parts of the vegetables exposed to the air from polluted environment, so regular monitoring of these metals in vegetables is essential. Although the content of lead and cadmium in cereals was very low (taking as a sample entire plant), it is also necessary to be analysed only the grain because they are used in feed and food industry. Leafy vegetables (especially salad) had proved to had the highest content of heavy metals (even if it was bellow the limits). No contamination was found in all tested samples.

16 PERSPECTIVES Considering chemical contamination of food as a hidden contamination that it can t be seen or tasted, the perspectives in the future are related to discovering/innovation of the new tools in order to control/monitor the level of the most toxicogenic chemical contaminants and assure food safety. So, the perspectives can be: Analysis of heavy metals contamination on the entire food chain: soil, water and plant is needed. Extending the analyses to other potential carcinogenic contaminants along the food chain will be envisaged. Understanding the way of accumulation in order to find prevention measures that can be taken into account. Developing of new and more efficient safety management tools.

17 Gabriel Mustatea, PhD Nastasia Belc, PhD

Lead and Cadmium Levels in Some Commercially Available Local and Imported Peach Juice Samples in Alkoms City

Human Journals Research Article September 2018 Vol.:13, Issue:2 All rights are reserved by Zaid M. Najah et al. Lead and Cadmium Levels in Some Commercially Available Local and Imported Peach Juice Samples