Transfer of Cesium and Potassium from Grapes to Wine

Transcription

1 Transfer of Cesium and Potassium from Grapes to Wine Nami Goto-Yamamoto, 1 * Kazuya Koyama, 1 Kaori Tsukamoto, 1 Hiroshi Kamigakiuchi, 1 Masanori Sumihiro, 1 Masaki Okuda, 1 Tomokazu Hashiguchi, 1 Katsumi Matsumaru, 1 Haruhito Sekizawa, 2 and Hitoshi Shimoi 1 Abstract: The food-processing transfer parameters of radioactive and stable cesium and radioactive potassium were determined from grapes to wine. The concentration of cesium in the pomace was higher than that in juice, as was the case of potassium. During white and rosé wine fermentation, cesium concentration did not change significantly and potassium concentration decreased. These results suggest that the absorbance of cesium by yeast is much lower than that of potassium in the winemaking environment. The food-processing retention factors (Fr, content in wine/ content in grape) of radiocesium and stable cesium for red wine were generally higher than those for white wine, reflecting the yields of wine and the extraction of cesium during maceration. Key words: radiocesium, radionuclide, cesium, food-processing transfer parameters The accident at the Fukushima Daiichi Nuclear Power Plant following the Great East Japan earthquake and tsunami in March 2011 led to a large release of radionuclides, of which contamination to food has been of great concern. Cesium (Cs) and iodine (I) were the major radionuclides released through the accident, and radioactive Cs is a long-lasting problem in regard to food contamination because of its relatively long half-life ( 134 Cs, 2 years; 137 Cs, 30.2 years). After the accident, much research was carried out to clarify the transfer of radioactive Cs from environment to raw foodstuffs (Nakanishi et al. 2013, Nemoto and Abe 2013) and its transfer from raw foodstuffs to processed foods and beverages (Tagami et al. 2012, Tagami and Uchida 2013, Okuda et al. 2013, Hachinohe et al. 2013). For grapes, it was reported after the 1986 nuclear power plant accident in Chernobyl, Ukraine, that grapevines absorb radiocesium through roots and leaves (Zehnder et al. 1995, Carini and Lombi 1997). In addition, after the accident in Fukushima, it was suggested that radiocesium passed through the bark of grapevines and peach trees (Takada 2013), as the accident occurred before the budburst of deciduous fruit trees. The inspection of radionuclides in raw foodstuffs by Fukushima prefectural institutions showed that the radiocesium concentration of 53% of grapes harvested in 2011 in Fukushima Prefecture was lower than the detection limit of the standard determination condition (~10 Bq/kg) and that the 1 National Research Institute of Brewing, Kagamiyama, Higashi-Hiroshima, Hiroshima, Japan; and 2 Fukushima Agricultural Technology Centre, 116 Shimonakamichi, Takakura, Hiwada, Koriyama, Fukushima, Japan. *Corresponding author (gotoh_n@nrib.go.jp) Manuscript submitted June 2013, revised Nov 2013, accepted Nov 2013 Copyright 2014 by the American Society for Enology and Viticulture. All rights reserved. doi: /ajev maximum concentrations of 134 Cs and 137 Cs were 57 and 64 Bq/kg, respectively (summarized in Nihei 2013). The provisional legal limit in raw foodstuffs until April 2012 in Japan was 500 Bq/kg, and the legal limit for general foodstuffs from April 2012 is 100 Bq/kg. To discuss the transfer of radionuclides from raw foodstuffs to processed foods, the International Atomic Energy Agency (IAEA 2010) recommended the following food-processing transfer parameters: food-processing retention factor, Fr = total activity in wine (Bq)/total activity in grapes (Bq); processing factor, Pf = concentration in wine (Bq/kg)/ concentration in grapes (Bq/kg); and processing efficiency, Pe = weight of wine/weight of grapes. Thus, Fr = Pf Pe. For wines, there are reviews of the transfer of radionuclides from grapes to wine after the accident in Chernobyl (Green 2001, IAEA 2010) and it was shown that the Fr of 134 Cs and 137 Cs in wine was 0.33 to However, the original reports were proceedings of meetings and are difficult to access, and we cannot know the change in the radionuclide concentrations during winemaking. Thus, we carried out small-scale vinification tests using grapes containing a small amount of radioactive Cs to show the change in its concentration during winemaking. In addition, the change in the concentration of radioactive potassium ( 40 K), which is a congener of Cs, was determined. It was reported that the Fr of 137 Cs and that of 40 K of vegetables through washing, boiling, and pickling were highly correlated (R = 0.966) (Tagami and Uchida 2013). A similar result was reported for rice polishing and washing (Okuda et al. 2013). We also examined the change in the concentration of stable cesium, 133 Cs, during winemaking. In general, isotopes behave similarly in terms of their physical properties and chemical reactivity (IAEA 2010). The behavior of stable 133 Cs is likely a useful indicator for predicting that of radioactive Cs in winemaking. Small-scale winemaking trials using seven red and white winegrape cultivars were conducted to investigate the transfer of 133 Cs into wine. 143

2 144 Goto-Yamamoto et al. Materials and Methods Grapes. Fukushima Prefecture is a prosperous fruitgrowing region, although it is not a major wine region in Japan. Thus, two black skin table-grape cultivars, Vitis labruscana Campbell Early and Takao, were used in this study. Takao is a seedless variety bred from a seedling of Kyoho, a cross between V. labruscana cv. Ishihara-Wase, which is a tetraploid bud mutant of Campbell Early, and V. vinifera cv. Centennial. These grapes were harvested in September 2012 in Fukushima Prefecture and trace concentrations of radiocesium were detected in the inspection by the prefecture. Grapes of V. vinifera cvs. Chardonnay, Sauvignon blanc, Cabernet Sauvignon, Merlot, and Koshu and Vitis labruscana cvs. Muscat Bailey A and Black Queen were harvested at full ripeness in 2012 in an experimental vineyard in Higashi-Hiroshima, Japan. The berries were destemmed from the clusters taken from different vines with three biological replicates. Approximately 30 berries from each of the three replicates were manually peeled with a scalpel to dissect skins and seeds. The whole berries, skins, and seeds were weighed and frozen under liquid nitrogen for the storage at -80 C. Winemaking. Blush-type rosé wine and red wine were made in a small scale using Campbell Early and Takao. For rosé winemaking, ~3.4 kg of destemmed grape berries were crushed and pressed by hand, and 150 mg/kg potassium metabisulfite was added while crushing. Sugar concentration was adjusted to 22% (w/v) with sucrose, and 200 mg/l of wine yeast X5 (Laffort, Bordeaux, France) was added following the instructions of the manufacturer. The must in a glass bottle was fermented in a room kept at 15 C, and 125 mg/l Fermaid K (Lallemand, Montréal, QC, Canada) was supplied on the second day of fermentation. wine must was made from ~3.0 kg of grape berries in a similar way and fermented by wine yeast RX60 (Laffort) at 25 C. wine must was pressed by hand after fermentation. After the addition of 100 mg/l potassium metabisulfite, red and rosé wines were racked and centrifuged at 5000 g for 15 min. Must settling before rosé wine fermentation and fining and filtration of wine were omitted. To examine the effect of yeast strains on the radiocesium concentrations of finished wine, the two yeast strains were used for rosé wine of Takao with two replications. The other winemaking experiments were triplicated using the same grape. To examine the transfer of stable cesium ( 133 Cs) from grapes to wine, red and white wines were made in a small scale using winegrapes with three replicates. For red winemaking, ~0.9 kg lots of destemmed grape berries of Cabernet Sauvignon, Merlot, Muscat Bailey A, and Black Queen were crushed. Sugar concentration was adjusted to 23% (w/v) with sucrose. Six hours after the musts were sulfited to 50 mg/l, wine yeast RX60 was inoculated. The must in a glass bottle was fermented in a room kept at 25 C for 8 days. wine was pressed by hand after fermentation. For white winemaking, the destemmed grape berries of Chardonnay, Sauvignon blanc, and Koshu were crushed and pressed by hand. The juices were settled and centrifuged at 4500 g for 15 min. Then wine yeast X16 (Laffort) was inoculated, and fermentation was conducted in a room kept at 15 C. To ensure that fermentation had completed, the glucose and fructose concentrations in the musts were checked with a d-glucose/d-fructose kit (Roche, Basel, Switzerland) and 150 mg/l potassium metabisulfite was then added. Both red and white wines were racked and centrifuged at 5000 g for 10 min. Radioactivity analysis. The radioactivities (Bq/kg) of 134 Cs, 137 Cs, and 40 K were determined using a Ge gamma-ray spectrometer (Seiko EG&G Ortec, Chiba, Japan) with a Marinelli beaker (~2 kg sample) for 3,000 sec of counting or with a U-8 container (~100 g sample) for 25,000 sec. A U-8 container was used for pomace, since its volume was small. Following the definition of IAEA (2010), food-processing transfer parameters of radionuclides were obtained (see the introduction). Analysis of stable cesium. The concentration of 133 Cs in the samples was determined using inductively coupled plasma mass spectrometry (ICP-MS) (Agilent 7700x, Tokyo, Japan) after wet-ashing using a microwave digestion system (MLS-1200MEGA, Milestone, Bergamo, Italy). Frozen samples of whole berries, skins, and seeds were ground to a fine powder using a mortar and pestle in liquid nitrogen. For microwave digestion, ~0.3 g of the ground powder or 2 ml of wine sample was accurately weighed into a tetrafluoromethaxil digestion vessel. An acid mixture (5 ml 61% HNO 3 and 1 ml 30% H 2 O 2 ) was then added. Programmed microwave irradiation was applied in sequential steps at the wattage (W) and times indicated: step 1, 250 W/3 min; step 2, 0 W/2 min; step 3, 250 W/5 min; step 4, 500 W/5 min; step 5, 0 W/2 min; step 6, 500 W/5 min. After the first digestion, the vessels were cooled to room temperature and another acid mixture (2 ml 61% HNO 3 and 1 ml 30% H 2 O 2 ) was added. The microwave program described above was then repeated. After completion of the second digestion, samples were transferred to calibrated flasks and diluted to 50 ml with ultrapure water. Samples were then filtered through 0.45 µm cellulose acetate syringetip filters (DISMIC-25CS, Advantec, Tokyo, Japan) before analysis by ICP-MS. 133 Cs losses as a result of cellulose acetate filtration were negligible ( ± 3.35 ng/l without filtration and ± 3.95 ng/l with filtration, n = 5). The analytical conditions for ICP-MS were as follows: RF power, 1.55 kw; RF matching, 1.80 V; sampling position, 8.0 mm; carrier gas flow, 0.85 L/min; dilution gas flow, 0.40 L/min; extract 1, 0 V; extract 2, -160 V; non-gas mode. The integration time was 0.99 sec, and the number of replications was five. 115 Indium was used as an internal standard to compensate for changes in the analytical signals during the operation of ICP-MS. The Fr and Pf of 133 Cs were calculated using the total amount and the concentration of 133 Cs, respectively, instead of total radioactivity and concentration of radionuclides. Results and Discussion Small-scale vinification. The analytical data of wine made from Campbell Early and Takao were determined and were within those of standard wines (Table 1). Although both cultivars are table grapes, Campbell Early is often used for winemaking in Japan. Two yeast strains were used for rosé winemaking from Takao, but no difference was observed in

3 Radiocesium Transfer from Grapes to Wine 145 the residual radioactive Cs concentration or other analytical data (data not shown). Thus, the average of four replications is shown in all the tables in this study. Small-scale vinification trials using seven wine cultivars were carried, and the alcohol concentrations of the finished wine were from 11.9 to 12.8%. Change in radioactive Cs and K during winemaking. The grapes of Campbell Early and Takao used in this study contained low concentrations of 134 Cs and 137 Cs: Campbell Early contained 2.5 ± 0.3 and 3.7 ± 0.4 Bq/kg, respectively, and Takao contained 3.1 ± 0.1 and 5.3 ± 0.5 Bq/kg, respectively. Thus, they are expressed in total radiosecium concentration. Since these grapes were harvested in 2012, one year after the accident, the radioactive Cs was likely transferred through the roots from the soil and/or through the surface of vines contaminated with radiocesium after the accident. Also, the grapes contained 40 K, which is a natural isotope of stable potassium, 39 K. For rosé winemaking, the concentrations of radiocesium in the pomace were 1.55 to 2.05 times higher than the pressed juice (Table 2). Thus, the concentration of radioactive Cs was decreased by pressing. On the other hand, the concentration of radioactive Cs did not change significantly during fermentation. For red winemaking, the concentrations of radioactive Cs in the wine were almost the same as those of crushed grapes. Mass balance of radioactive Cs (Table 3) showed that a higher ratio of radioactive Cs was transferred into red wine than into rosé wine. Similarly to Cs, the concentration of 40 K was reduced by juice pressing, which means both radioactive Cs and K distributed in a higher concentration in the pomace than in the juice. However, the concentration of 40 K decreased to 49.4 to 56.5% of that of the juice during the fermentation and racking of rosé wine. Also, during red winemaking, the concentration Table 1 Analytical data of wine. Takao rosé wine was made from one lot of juice with four replications. The other vinification was carried out in triplicate from crushing and pressing. Values are means ± standard deviations. Wine/cultivar Alcohol (% v/v) Extract (% w/v) ph Total acid a (g/100 ml) Campbell Early 12.8 ± ± ± ± Takao 13.0 ± ± ± ± Campbell Early 11.2 ± ± ± ± Takao 12.4 ± ± ± ± a Expressed as tartaric acid. Table 2 Changes in the concentration of radioactive cesium and potassium and in the sample weight during winemaking. wine of Takao was made from one lot of juice with four replications and its sample weight was expressed for one fermentation bottle. Radioactivities of pomace of rosé wine of Campbell Early and red wine of Takao were determined after combining and mixing all the pomace (n = 1 for Campbell Early and n = 2 for Takao). The other vinification and determination were carried out in triplicate. 134 Cs Cs (Bq/kg) 40 K (Bq/kg) Sample wt (kg) Campbell Early Takao Campbell Early Takao Campbell Early Takao Crushed grapes 6.42 ± ± ± Pomace ± Juice 5.99 ± ± ± After racking 5.30 ± ± ± ± ± ± 0.01 Crushed grapes 6.00 ± ± ± ± ± ± 0.02 Pomace 9.42 ± ± ± ± ± ± 0.01 After racking 6.11 ± ± ± ± ± ± 0.00 Table 3 Mass balance (%) of radioactive cesium and potassium during winemaking. 134 Cs Cs (%) 40 K (%) Campbell Early Takao Campbell Early Takao Crushed grapes Pomace Juice 64.6 ± ± After racking 50.9 ± ± ± ± 4.4 Crushed grapes Pomace 25.7 ± ± After racking 79.3 ± ± ± ± 14.5

4 146 Goto-Yamamoto et al. of 40 K decreased to 46.7 to 55.1% of that of the crushed berries. These results indicate that Cs was not absorbed by the yeast significantly, while K was absorbed to a large extent. The yeast Saccharomyces cerevisiae BY4741 was reported to absorb Cs in YPD medium (1% yeast extract, 2% peptone, and 2% glucose) with agitation through both the same mechanism as K + uptake and the different mechanism (Heuck et al. 2010). However, the uptake of Cs was not active in the winemaking environment of this study. A similar result to this study of wine was observed during the production of sake (Okuda et al. 2012, 2013). Food-processing transfer parameters. The Fr of wine obtained in this study was to (Table 4). The Pf was ~1.0 in red wine and ~0.8 to 0.9 in rosé wine, indicating that maceration during red winemaking increased Cs concentration due to extraction from the skin and seeds. The higher Fr values of red wines resulted from their higher Pf and Pe values than those of rosé wine. Distribution of stable cesium in berry tissues and transfer into wine. To examine the 133 Cs distribution in the berries, the berries were dissected into skins and seeds and 133 Cs content in the tissues were measured. 133 Cs concentrations in the berry skin were 1.6 to 3.4 times higher than those of the whole berry; those in the seeds were 1.6 to 2.4 times higher (Table 5). Because of the small percentage of these tissues within the whole berries, 133 Cs contents in these tissues per Wine/cultivar Table 4 Food-processing transfer parameters of rosé and red wines. Fr Pf 134 Cs Cs 134 Cs Cs 40 K Campbell Early ± ± ± ± Takao ± ± ± ± Campbell Early ± ± ± ± Takao ± ± ± ± Pe Table Cs concentrations and the sample weights of the raw materials and wine of seven winegrape cultivars. Values are means ± standard deviations (n = 3). Chardonnay Sauvignon blanc Koshu Cabernet Sauvignon Merlot Black Queen Muscat Bailey A Cs concn (µg/kg) Whole berry 8.18 ± ± ± ± ± ± ± 1.26 Skin ± ± ± ± ± ± ± 9.67 Seed ± ± ± ± ± ± ± 4.68 Wine 6.28 ± ± ± ± ± ± ± 2.42 Sample wt (kg) Whole berry ± ± ± ± ± ± ± Skin ± ± ± ± ± ± ± Seed ± ± ± ± ± ± ± Wine ± ± ± ± ± ± ± Mass balance (%) Whole berry Skin 13.7 ± ± ± ± ± ± ± 2.5 Seed 5.9 ± ± ± ± ± ± ± 0.1 Wine 45.8 ± ± ± ± ± ± ± 4.6 Table 6 Food-processing transfer parameters of 133 Cs during winemaking. Values are means ± standard deviations (n = 3). Wine/cultivar Fr Pf Pe White Chardonnay ± αβ a ± αβ ± α Sauvignon blanc ± α ± α ± α Koshu ± βγ ± βγ ± α Cabernet Sauvignon ± γδ ± αβγ ± β Merlot ± ε ± γ ± β Black Queen ± δε ± γ ± β Muscat Bailey A ± δε ± γ ± β a Different letters (α, β, γ, δ, ε) indicate that values are significantly different at p < 0.05 by Tukey s HSD.

5 Radiocesium Transfer from Grapes to Wine 147 berry were relatively low, and ~80% of 133 Cs was distributed in the pulp in all the cultivars examined in this study. The distributions of 133 Cs in the skins were lower than those of radioactive Cs in the pomace, probably due to the different methods of preparation. The Fr of 133 Cs during winemaking of these grape cultivars was to (Table 6). These values were close to those of 134 Cs and 137 Cs in wine reported by IAEA (2010), as well as those of radiocesium in this study (Table 4), validating the use of stable 133 Cs to predict the behavior of radioactive Cs during winemaking. In addition, the Fr of all red grape cultivars other than Cabernet Sauvignon was significantly higher than that of all white cultivars (Table 6). Similar to the results of rosé wine (Table 4), the Pe of white cultivars was significantly lower than that of red cultivars. The Pf of red cultivars except Cabernet Sauvignon was ~1.0 and significantly higher than that of Chardonnay and Sauvignon blanc. This result indicated that Cs was extracted from the skin and seeds and possibly from the solid parts of pulp during maceration. Similar to the results of radioactive Cs during red and rosé winemaking, the higher Fr values of red wines resulted from their higher Pf and Pe values than those of white wines. Conclusion Radioactive Cs was contained both in the juice and pomace of grape berries, and its concentration in the pomace was ~1.5 to 2.0 times higher than that in the juice. Decrease of radioactive Cs during fermentation was much less than that of K and negligible. Thus, Fr of radioactive Cs from grapes to wine depends on the yield of juice or wine as well as on the extraction from the pomace during the winemaking process. Consistent with the result of the radioactive Cs, the Fr and Pf of stable Cs in red wine tended to be higher than in white wine, validating the use of stable Cs to predict the behavior of radioactive Cs during winemaking. Literature Cited Carini, F., and E. Lombi Foliar and soil uptake of 134 Cs and 85 Sr by grape vines. Sci. Total Environ. 207: Green, N The effect of storage and processing on radionuclide content of fruit. J. Environ. Radioact. 52: Hachinohe, M., et al Distribution of radioactive cesium ( 134 Cs plus 137 Cs) in a contaminated Japanese soybean cultivar during the preparation of tofu, natto, and nimame (boiled soybean). J. Food Prot. 76: Heuck, S., U.C. Gerstmann, B. Michalke, and U. Kanter Genome-wide analysis of caesium and strontium accumulation in Saccharomyces cerevisiae. Yeast 27: IAEA Handbook of Parameter Values for the Prediction of Radionuclide Transfer in Terrestrial and Freshwater Environments. Technical reports series 472. International Atomic Energy Agency, Vienna, Austria. Nakanishi, T.M., N.I. Kobayashi, and K. Tanoi Radioactive cesium deposition on rice, wheat, peach and soil after nuclear accident in Fukushima. J. Radioanal. Nucl. Chem. 296: Nemoto, K., and J. Abe Radiocesium absorption by rice in paddy field ecosystems. In Agricultural Implications of the Fukushima Nuclear Accident. T.M. Nakanishi and K. Tanoi (eds.), pp Springer Japan, Tokyo. Nihei, N Radioactivity in agricultural products in Fukushima. In Agricultural Implications of the Fukushima Nuclear Accident. T.M. Nakanishi and K. Tanoi (eds.), pp Springer Japan, Tokyo. Okuda, M., M. Joyo, M. Tokuoka, T. Hashiguchi, N. Goto-Yamamoto, H. Yamaoka, and H. Shimoi The transfer of stable 133 Cs from rice to Japanese sake. J. Biosci. Bioeng. 114: Okuda, M., T. Hashiguchi, M. Joyo, K. Tsukamoto, M. Endo, K. Matsumaru, N. Goto-Yamamoto, H. Yamaoka, K. Suzuki, and H. Shimoi The transfer of radioactive cesium and potassium from rice to sake. J. Biosci. Bioeng. 116: Tagami, K., S. Uchida, and N. Ishii Extractability of radiocesium from processed green tea leaves with hot water: The first emergent tea leaves harvested after the TEPCO s Fukushima Daiichi Nuclear Power accident. J. Radioanal. Nucl. Chem. 292: Tagami, K., and S. Uchida Comparison of food processing retention factors of 137 Cs and 40 K in vegetables. J. Radioanal. Nucl. Chem. 295: Takada, D Distribution of radiocesium from the radioactive fallout in fruit trees. In Agricultural Implications of the Fukushima Nuclear Accident. T.M. Nakanishi and K. Tanoi (eds.), pp Springer Japan, Tokyo. Zehnder, H.J., P. Kopp, J. Eikenberg, U. Feller, and J.J. Oertli Uptake and transport of radioactive cesium and strontium into grapevines after leaf contamination. Radiat. Phys. Chem. 46:61-69.