ARTICLE pubs.acs.org/est
Effects of Arsenic Compound Amendment on Arsenic Speciation in Rice Grain Tomohito Arao,*,† Akira Kawasaki,† Koji Baba,† and Shingo Matsumoto‡ † ‡
National Institute for Agro-Environmental Sciences, 3-1-3 Kannondai, Tsukuba, Ibaraki 305-8604, Japan Shimane University, 2059 Kamihonjomachi, Matsue, Shimane 690-1102, Japan
bS Supporting Information ABSTRACT: Rice consumption is a major source of arsenic for Asian populations. Arsenic is present in rice grain both as inorganic arsenic and as dimethylarsinic acid (DMA). It is unclear whether DMA in rice is taken up from the soil or synthesized in planta. We investigated the effect of DMA, methylarsonic acid (MMA) and arsenite amendment on arsenic speciation in rice grain grown in soil and in solution culture. We also investigated the methylation of arsenic in solution culture under suppression of bacterial activity. When rice was grown under flooded conditions after the heading stage, DMA amendment to the soil resulted in higher DMA concentration in brown rice and rice straw. In the solution culture, not only DMA amendment but also MMA or arsenite amendment increased the DMA concentration in brown rice and rice straw. DMA was detected in the solution amended by MMA or arsenite with young rice plants. When the solution included the antibacterial agent chloramphenicol, DMA concentration in the solution decreased dramatically. When only the soil was incubated with MMA or arsenite, only a slight amount of DMA was detected in the soil. These results suggest that rice rhizosphere associated bacteria would be involved in the formation of DMA in brown rice.
’ INTRODUCTION Arsenic (As) is a carcinogen and the intake of inorganic As in rice is a significant risk factor for cancer in populations for which rice is a staple foodstuff.1 In some cases, human As intake from the consumption of rice exceeds that from drinking water.2 For inorganic As, a provisional tolerable weekly intake (PTWI) of 15 μg kg-1 body weight was established by the Joint FAO/WHO Expert Committee on Food Additives (JECFA).3 The Committee withdrew this PTWI at its 72nd meeting.4 Inorganic As causes cancer of the lung, urinary tract, and skin, and a range of adverse effects have been reported at exposures lower than those reviewed by the JECFA. Therefore, the European Food Safety Authority’s Panel on Contaminants in the Food Chain concluded that the JECFA PTWI of 15 μg kg-1 body weight is no longer appropriate.5 Contamination by As occurs to a greater extent in paddy rice than in other upland crops because anaerobic conditions in paddy soil lead to arsenite mobilization and thus enhanced bioavailability to rice.6 The Ministry of Agriculture, Forestry, and Fisheries of Japan analyzed the As content of staple crops in Japan7 and found that As concentration in brown rice ranged from 0.04 to 0.33 mg kg-1, with an average value of 0.16 mg kg-1 (n = 199). As concentration in most other agricultural products was below the detection limit. Rice is therefore a major source of dietary intake of inorganic As in the Japanese population. Rice is the staple food for about half the world’s population. There is an urgent need to understand how rice absorbs and metabolizes As in order to develop mitigation strategies against widespread contamination in the food chain.8 Arsenic is present in rice grain both as inorganic As (mainly arsenite) and as dimethylarsinic acid (DMA). Inorganic As is generally considered to be more toxic than methylated As compounds.9 The grain of aerobically grown rice contains As almost r 2011 American Chemical Society
exclusively in the form of inorganic As, whereas in anaerobically grown rice, DMA accounts for the majority of the total As.10,11 It has not yet been resolved whether DMA in rice is taken up from the soil or synthesized from inorganic As in planta. One possible explanation for the greater percentage of DMA in anaerobically grown rice than in aerobically grown rice is that DMA is synthesized in the soil or rhizoplane and translocates to the grain. The main objective of the present study was to investigate the effect of DMA, methylarsonic acid (MMA) and arsenite amendment on arsenic speciation in rice grain grown in soil and in solution culture. We also investigated the methylation of As in solution culture under suppression of bacterial activity.
’ MATERIALS AND METHODS Pot Experiments. Pot experiments, with three replications each, were performed in 2009 in a greenhouse at ambient temperatures under sunlight. Wagner pots (1/5000 a, Fujiwara Scientific Co., Tokyo, Japan) were filled with 2.5 kg of soil B 11 collected from the plowed layer of a paddy field. Soil B contained 3.4% total C, 0.32% total N, 0.66 mg kg-1 total Cd, and 48 mg kg-1 total As, and it had a pH of 5.5. Rice seedlings (Oryza sativa L. cv. Koshihikari) were germinated on perlite and transplanted to the soil samples on 11 May 2009. A compound fertilizer containing 0.2 g of N, 0.04 g of P, and 0.08 g of K was supplied to each pot by basal application. Ammonium sulfate containing 0.2 g of N was also supplied to each pot by top dressing 60 days after transplantation of the rice seedlings. The rice was grown under flooded Received: October 7, 2010 Accepted: December 22, 2010 Revised: December 8, 2010 Published: January 19, 2011 1291
dx.doi.org/10.1021/es1033316 | Environ. Sci. Technol. 2011, 45, 1291–1297
Environmental Science & Technology conditions for five weeks from transplanting, followed by intermediate irrigation to 13 July 2009. Three water-management treatments were examined in the experiment: Treatment A involved flooding from 13 July to 28 August; treatment B, under intermediate irrigation from 13 July to the time of heading (emergence of rice ear) and flooding from heading to 28 August; treatment C, under intermediate irrigation from 13 July to 28 August. From 28 August, the rice was grown under drainage conditions before harvest. Redox potential (Eh) was measured at a depth of 10 cm. For intermediate irrigation planted pots were irrigated daily. Irrigation volume was decided according to the daily weather such that the Eh value would not be below zero. The heading days occurred between August 2nd and August 6th. Under treatment B and C, 1.8 mg of DMA per kilogram of soil was added by aqueous solution of DMA to the top layer of soil on 13 July. A soil-water sampler (DIK8393, Daiki Rika Kogyo Co., Saitama, Japan) was buried in the middle of the soil in each pot for collecting soil solution. The soil solution was sampled and diluted with 10% HNO3 at a ratio of 9:1 immediately after collection and filtering through a 0.45 μm filter. Solution Culture and Soil Incubation. Solution culture experiments with three replications were performed in a greenhouse at 22-28 °C under sunlight. Rice (O. sativa L. cv. Koshihikari) was cultivated in Soil A11 under field conditions in Tsukuba, Japan. Rice seedlings were transplanted on 11 May 2009 and were grown under flooded conditions for five weeks from transplanting, followed by intermediate irrigation to 8 July. The rice plants were transferred from the plots to Wagner pots (1/5000 a) on 1 July 2009 and grown in nutrient solution of 0.091 mM KNO3, 0.183 mM Ca(NO3)2, 0.274 mM MgSO4, 0.1 mM KH2PO4, 0.183 mM (NH4)2SO4, and 20 μM Fe(III)-EDTA. At the time of heading, the nutrient solution was renewed and amended with arsenic species. DMA-d6 and MMA-d3 (Supporting Information (SI) Figure S2) were purchased from Hayashi Pure Chemical Industries (Osaka, Japan). NaAsO2 was purchased from Aldrich (St. Louis, MO). Concentrations of As chemicals were determined by ICP-MS (ELAN DRC-e, PerkinElmer SCIEX, DE, USA). Nutrient solution a2 or a3 contained 1.3 μM DMA-d6 or 1.3 μM MMA-d3, respectively. Nutrient solution a4 contained 1.3 μM DMA-d6, 1.3 μM MMA-d3 and 1.3 μM NaAsO2. Nutrient solution a5 contained 13 μM NaAsO2. The solution remaining five days after the addition of the arsenicals was collected and filtered through a 0.45 μm filter and the nutrient solution was renewed. Five days later, the nutrient solution was renewed again. Another five days later, the seedlings were transferred to nutrient solution without the arsenic and were grown for six weeks. The nutrient solution was renewed once every seven days. The rice grain and straw were harvested at the time of maturity. Solution culture experiments and soil incubation experiments with three replications were performed in a greenhouse at 2228 °C under sunlight. Soil A was collected from the plowed layer of the plots for the paddy field and sieved to
