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Food Additives & Contaminants: Part B: Surveillance Publication details, including instructions for authors and subscription information: http://www.tandfonline.com/loi/tfab20
Survey of nitrite content in foods from north-east China a
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Y. Yuan , T. Zhang , H. Zhuang , K. Wang , Y. Zheng , H. Zhang , B. Zhou & J. Liu
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Department of Food Quality and Safety , College of Quartermaster Technology, Jilin University , Changchun 130062, China Published online: 18 Mar 2010.
To cite this article: Y. Yuan , T. Zhang , H. Zhuang , K. Wang , Y. Zheng , H. Zhang , B. Zhou & J. Liu (2010) Survey of nitrite content in foods from north-east China, Food Additives & Contaminants: Part B: Surveillance, 3:1, 39-44, DOI: 10.1080/19440040903514515 To link to this article: http://dx.doi.org/10.1080/19440040903514515
PLEASE SCROLL DOWN FOR ARTICLE Taylor & Francis makes every effort to ensure the accuracy of all the information (the “Content”) contained in the publications on our platform. However, Taylor & Francis, our agents, and our licensors make no representations or warranties whatsoever as to the accuracy, completeness, or suitability for any purpose of the Content. Any opinions and views expressed in this publication are the opinions and views of the authors, and are not the views of or endorsed by Taylor & Francis. The accuracy of the Content should not be relied upon and should be independently verified with primary sources of information. Taylor and Francis shall not be liable for any losses, actions, claims, proceedings, demands, costs, expenses, damages, and other liabilities whatsoever or howsoever caused arising directly or indirectly in connection with, in relation to or arising out of the use of the Content. This article may be used for research, teaching, and private study purposes. Any substantial or systematic reproduction, redistribution, reselling, loan, sub-licensing, systematic supply, or distribution in any form to anyone is expressly forbidden. Terms & Conditions of access and use can be found at http:// www.tandfonline.com/page/terms-and-conditions
Food Additives and Contaminants: Part B Vol. 3, No. 1, March 2010, 39–44
View Dataset Survey of nitrite content in foods from north-east China Y. Yuan, T. Zhang, H. Zhuang, K. Wang, Y. Zheng, H. Zhang, B. Zhou and J. Liu* Department of Food Quality and Safety, College of Quartermaster Technology, Jilin University, Changchun 130062, China
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(Received 1 September 2009; final version received 27 November 2009) This study reports a survey of nitrite in a variety of foods consumed in north-east China and estimates the intake of nitrite for the north-east Chinese consumer. A total of 642 food categories including rice and rice products, flour and flour products, soybean and products, vegetables, fruit, preserved vegetables, cured meat products, dairy products, fish products, salt, and soy sauce were analysed for their content of nitrite. Nitrite content was quite different both between different food categories and within the same food category, ranging from not determined (n.d.) to 19.7 mg kg1. A great variation in the content of nitrite was found for all the food products. The average content of nitrite was highest in cured meat products (14.3 mg kg1). Next to that, the nitrite content was high in the order of preserved vegetables (4.1 mg kg1), soybean products (3.5 mg kg1), and dairy products (1.9 mg kg1). The lowest average values of nitrite were detected in soy sauce, rice and rice products, salt and fish products, the contents being 0.1, 0.3, 0.3, and 0.6 mg kg1. Calculations on the basis of these results and including dietary surveys show that the average intake of nitrite in north-east China from food was 0.03 mg kg1 body weight for an average Chinese person weighing 60 kg, and the data are lower than the established acceptable daily intake (ADI) for nitrite. Cured meat products are normally the major contributor to average nitrite intake of the north-east Chinese population. The second contributor is vegetables. Keywords: risk assessment; nitrite
Introduction Nitrite and nitrate occur naturally in food as a consequence of the nitrogen cycle whereby nitrogen is fixed by bacteria. Additionally, nitrate and nitrite are often used as food additives in the form of sodium or potassium salt in modern food processing, especially in meat products, where they fulfil the function of preservation, antimicrobial agent and colour fixative (Reinik et al. 2005). Nitrate and nitrite contamination of vegetables is produced mainly by chemical fertilizers, especially nitrogenous fertilizers (Zhou et al. 2000). The nitrate ion has a low level of acute toxicity, but if transformed into nitrite it may constitute a health problem (Reinik et al. 2005). Research has shown that 5–8% of the nitrate from the diet may be reduced to nitrite by the microflora in the oral cavity (Gangolli et al. 1994; World Health Organization (WHO) 1996; Mensinga et al. 2003). Nitrite is known to be a precursor of toxic and carcinogenic N-nitrosamines (Bassir and Maduagwu 1978) and high dietary intakes of nitrate and nitrite will induce gastric cancer based on epidemiology and clinical studies (Bartsch et al. 1990; Joossens et al. 1996; Sen and Baddoo 1997). Only a few studies have been conducted to estimated the consumption of food additives including *Corresponding author. Email: [email protected] ISSN 1939–3210 print/ISSN 1939–3229 online ß 2010 Taylor & Francis DOI: 10.1080/19440040903514515 http://www.informaworld.com
nitrate and nitrite in China, especially in the north-east (Zhou et al. 2000; Zhong et al. 2002), and have rather focused on the nitrate and nitrite content of vegetables. This research has paid little attention to the contents and effect of cured meat products and other typical Chinese foods on the Chinese population. Thus, it is necessary to learn about the content of nitrite in Chinese foods. This paper presents the results of the content of nitrite in different food categories from north-east China, including rice and rice products, flour and flour products, soybean and products, vegetables, fruit, preserved vegetables, cured meat products, dairy products, fish products, salt, and soy sauce. The daily intake of nitrite from food (exogenous intakes) from north-east Chinese markets was then estimated according to the data obtained here and combined with the average daily consumption of foods. The data could help customers avoid the adverse effects of nitrite consumption.
Materials and methods Chemicals All solvents used in the experiments were analytical grade. The distilled water was free of ions.
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Sampling All samples selected in this survey were purchased at local supermarkets and stores in Changchun city in 2009. A total of 642 samples covered eleven categories of popular foods in north-east China, including rice and rice products, flour and flour products, soybean and products, vegetables, fruit, preserved vegetables, cured meat products, dairy products, fish products, salt, and soy sauce. The samples were stored in a freezer at 18 C until analysis.
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Sample preparation and nitrite analysis Nitrite contents were determined according to the Chinese National Standards Analytical Method (GB/T 5009.33-2008) with slight modification. Before analysis, non-edible parts of the sample were removed, and the whole sample was chopped and frozen. Just before analysis, the samples were homogenized in a blender. Food samples (2.0–10.0 g) were blended with 80 ml of distilled water in a 100 ml volumetric flask. Samples were treated ultrasonically for 30 min, and heated in a water bath at 75 C for up to 5 min, and then made up with distilled water to 100 ml and shaken. The mixtures were centrifuged at 10,000 rpm for 10 min and the supernatant removed for ultra-filtration. The filtrate was used for further nitrite analysis. Nitrite in the extract was reacted with 0.5% sulphanilamide at a concentration of 4 g l1 for 3 min and 0.25% N-(1-naphthyl)ethylene-diamine with a concentration of 2 g l1 was added to the extract and left to stand for 15 min. The absorbance of the violet azo compound was then measured at 540 nm. Chemical analysis for each sample of food was repeated three times. The method was shown to provide an accurate result by analysis of the standard solution of nitrite (ID: GSBZ50006-88) with a concentration of 0.050 mg l1 from the Chinese CRM/RM Information Center (Beijing, China). The result for the nitrite concentration was measured at 0.050, 0.050, 0.051, 0.050, 0.051 and 0.050 mg l1 for six analyses.
Statistical analysis Statistical analysis was performed with a Student’s t-test with SPSS 15.0 software. Analysis of variance (ANOVA) was tested on a significance level of p ¼ 0.05.
Results and discussion Analytical quality assurance The performance of the method was established in terms of its limit of quantification (LOQ), linearity, repeatability or run-to-run precision. Nitrite
(1.0–20.0 mg l1) was spiked in different blank matrices including water, spinach, dried tofu and ham, and recovery values were 89–102%. The LOQ in water and spinach matrix were 0.009 and 0.02 mg kg1, respectively. The good linearity of the method was achieved in a range from zero to 5.00 mg kg1 (r40.99). Precision was evaluated in terms of intra-day repeatability and inter-day reproducibility as RSD%. Both spinach and ham were tested. They had nitrite contents of 2.0 and 13.5 mg kg1, respectively. RSD ranged from 1.6% to 3.5% for the intra-day precision tests (n ¼ 5) and from 2.4% to 6.3% for the inter-day precision tests (n ¼ 15). A standard curve was made before and after analysis. To ensure the stability of the method, one in every ten samples analysed was a standard and this had to be within the 95% confidence interval of the standard curve.
Nitrite content in foods The study was focused on Chinese foods available in Chinese markets. Altogether, 642 food samples were divided into eleven kinds of food categories. These samples were included a wide range of food categories in the north-east China market and a lot of them were traditional Chinese foods. For food category and the numbers of samples within different ranges of nitrite content, see Table 1. Nitrite content was quite different both between different food categories and within the same food category, ranging from not determined (n.d.) to 19.7 mg kg1. The average content of nitrite was highest in cured meat products. Next to that, the nitrite content was high in the sequential order: preserved vegetables, soybean products, and dairy products. The average concentrations of nitrite were 14.3, 4.1, 3.5, and 1.9 mg kg1, respectively. The lowest average values of nitrite were detected in soy sauce, rice and rice products, salt and fish products, the contents being 0.1, 0.3, 0.3, and 0.6 mg kg1. There was a great variation in nitrite contents between the brands in each food groups, as easily seen from the high standard deviations shown in Table 1. Since nitrite can be formed via the reduction of nitrate if the foods are stored incorrectly (Zhong et al. 2002), the high variability of the data is mainly a result of variable food brands, the use of nitrate-based fertilizers, variable sampling places in this survey, and storing places. Ham and sausage were analysed for their nitrite content, and the results are shown in Table 1. The average contents of nitrite in ham and sausage were 16.1 and 12.5 mg kg1, respectively. Nitrite is used as an antimicrobial agent to provide protection against toxic microorganisms and as colouring or flavouring agents to produce the characteristic pink colour, texture and flavour in meat products (Reinik et al. 2005). Since the toxic nature of nitrite and the
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Table 1. Numbers of samples within different ranges of nitrite content (mg kg1).
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Rice and rice products Flour and flour products Soybean and products Vegetable Fruits Preserved vegetables Cured meat products Dairy products Fish products Salt Soy sauce Total
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0.01–0.5
0.5–1.0
1.0–5.0
5.0–10.0
10–20
420
Number
0 0 0 0 1 0 0 0 0 0 0 1
11 12 0 170 20 1 0 9 3 6 6 238
0 0 0 105 3 3 0 0 1 0 0 112
1 6 24 79 24 61 3 15 2 0 0 215
0 0 3 3 0 23 11 0 0 0 0 40
0 0 0 0 0 2 31 0 0 0 0 33
0 0 0 0 0 0 3 0 0 0 0 3
12 18 27 357 48 90 48 24 6 6 6 642
formation of nitrosamines by nitrite in cured meat products, many governments have established maximum levels for nitrite in many kinds of food. The European Parliament and its Council Directive have established maximum levels for nitrate and nitrite of 150 and 300 mg kg1, respectively. Residual amounts should not exceed 50 mg kg1 nitrite in non-heattreated meat products and 100 mg kg1 nitrite in all other meat products, except Wiltshire bacon and some other similar products which are limited to 175 mg kg1 nitrite (Honikel 2008). In China, the tolerance limit for nitrite is 20 mg kg1 for ham and sausage (National Standards 2005). In the present survey, there were three samples exceeding the Chinese tolerance limit. Since the difference in using nitrite and nitrate and using methods for those two kinds of additives, a little difference could found between the present study and that of Sen and Baddoo (1997) in sausage and ham in Canada. The average content of nitrite in their studies was 26 mg kg1 in sausage and 24 mg kg1 in ham, respectively. However, in the present survey the maximum contents of residual nitrite was 19.7 mg kg1 in ham and 15.9 mg kg1 in sausage, respectively. Since some manufacturers add less nitrite but more nitrate as a nitrite reserve (Hsu et al. 2009), this might also explain the differences in the findings by different researchers. Preserved vegetables are traditional Chinese food and many north-east Chinese people like to eat preserved vegetables in their daily lives, especially in winter. In the present survey, nitrite was found in preserved vegetables (4.1 mg kg1 on average) such as pickled vegetables, pickled Chinese cabbage and Chinese pickled vegetables. The average contents of nitrite were 2.6, 2.7 and 4.3 mg kg1, respectively. In China, the tolerance limit for nitrite is 20 mg kg1 in preserved vegetables (National Standards 2005). In the present survey, none of the samples contained over 20 mg kg1. But studies showed that nitrates in soil are absorbed into certain vegetables, for instance radish
and cabbage, which have a higher content of nitrate. The pickling process makes the conversion from nitrates to nitrites possible, resulting from the reduction by Escherichia, Proteus, Salmonella and so forth. Nitrite content increases as temperature increases in the pickling process. Also, salt concentration in the substrate has a significant influence. Nitrites reach a maximum at about 37 C when 5% salt is available. The nitrite content varies, increasing during the second to the fourth days, reaching a maximum during the seventh to eighth days, while dropping markedly 20 days later. While the decomposition by lactic acid bacteria to the nitrite in preserved vegetables was divided into enzymolysis and acid splitting, in the early period of fermentation, when the fermentation soup’s pH was greater than 4.5, enzymolysis was the main controlling factor, but when the pH was less than 4.0 because the amount of acid was increasing, acid splitting became the controlling factor. The nitritedecomposing ability of lactobacillus was stronger than lactococcus because of its acid-producing ability (Hashimo 2001; Ji 2007). As vegetables are consumed in large amounts in north-east China, the number of samples included in the study was the largest. A total of 357 vegetables were analysed for nitrite content. The average content of nitrite in vegetables was 0.9 mg kg1, and cherry tomato had the highest nitrite content (2.7 mg kg1) compared with other vegetables, and the maximum content was reached at 4.1 mg kg1. Spinach also had a high content of nitrite (2.1 mg kg1 on average) in the selected vegetables, and the maximum content was reached at 4.8 mg kg1. The finding correlated well with the study of Chung et al. (2003), who also found a high nitrite content in spinach (1.0 mg kg1 on average) in Korea. The difference between north-east China and Korea may be because of the varieties and use of nitrate-based fertilizers (Hsu et al. 2009). A high standard deviation could also be found in nitrite content from different vegetable varieties because
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Y. Yuan et al.
cultivar and harvest date can affect the nitrite levels of selected vegetables (Amr and Hadidi 2001). This might explain the high variability between findings presented in the present study also contributing to a high standard deviation as observed in Table 1. Garlic contained the lowest amount of nitrite in this study (0.1 mg kg1 on average), which was lower that that in the study of Chung et al. (2003) which demonstrated a nitrite content in garlic of 0.2 mg kg1 on average. The difference may be due to cultivation conditions. A total of 48 fruits were also analysed for nitrite content, including banana, pear, apple, and watermelon. The average content of nitrite was 1.3 mg kg1; apple had the highest nitrite content in the selected fruits (2.0 mg kg1 on average). Thus, it can be concluded that apple contributed to the highest dietary nitrite intake from fruits. Next were watermelon and banana. The average concentrations of nitrite were 2.0 and 1.7 mg kg1, respectively. Dogan et al. (2008) also found that nitrite could exist in apple and pear products. Okafor and Ogbonna (2003) found some nitrite content in fruit juices. Hord et al. (2009) reported that about 0.09 mg kg1 of nitrite could be detected in banana. The present study has found for the first time nitrite content in watermelon. The results might be the accumulation of nitrate in the soil or nitrate-rich soil, which would transfer the nitrite during growth of the watermelon (Onyesom and Okoh 2006). For statistical evaluation of the data about the content of nitrite obtained in different food categories of north-east Chinese food, a box-and-whisker plot (generated by OriginPro 7.5 software) was used. The plot is a powerful statistical tool that displays the median, range, minimum, maximum, mean, 25th percentile and 75th percentile. It is necessary since the analytical results are not normally distributed, especially in this kind of survey. Nitrite contents in different food categories are varied, and the median and inter-quartile range are two appropriate statistics to describe the central tendency and the spread around the median, respectively. Figure 1 shows that the nitrite content in different food categories ranges quite markedly. Nitrite levels in most food categories were less than 5 mg kg1, except for cured meat products from the median of the data. And from the median, one can also see that most of the data lie toward the lower rather than the higher end of the range. This box plot offered a great deal more insight than a lone average, and much more than an average complemented by the low and high data.
Exposure estimation and risk assessment of nitrite in Chinese foods Nitrite can react with nitrosatable compounds, such as amides and amines, to form N-nitroso compounds.
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Nitrite content (mg kg–1)
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0 A
B
C
D
E F G H Food categories
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J
K
Figure 1. Nitrite contents in north-east Chinese foods. A, Rice and rice products; B, flour and flour products; C, soybean and products; D, vegetables; E, fruits; F, preserved vegetables; G, poultry; H, dairy products; I, fish and products; J, salt; and K, soy sauce. The centre horizontal line of the box is the median of the data; the top and bottom of the box are the 25th and 75th percentiles (quartiles); and the ends of the whiskers are the 10th and 90th percentiles. Any points outside are considered outliers, which are labelled as ‘X’. The small square of each box means the average of the data.
Some N-nitroso compounds are potent carcinogens in animal species, and therefore can be carcinogenic to man (Onyesom and Okoh 2006). Once the nitrite contents in food were established, one can estimate the intake of these compounds based on national dietary surveys. The average content of nitrite in foods was used in combination with the average daily consumption of foods. The average consumption of different kind foods in China was completed with The Nutrition and Health Status of the Chinese People (Ministry of Health, Ministry of Science and Technology 2002). In the present survey the average dietary exposure to nitrite in the north-east Chinese population was estimated to be 1.826 mg day1 and was 0.030 mg kg1 body weight for an average Chinese person weighing 60 kg. The maximum intake of nitrite was 0.088 mg kg1 body weight. Simultaneously, the contributions of different food categories to the total nitrite intake were ranked. Cured meat products constituted the major contribution at 1.138 mg day1 (0.019 mg kg1 body weight, about 62.3% of total nitrite) to the average nitrite intake of the north-east Chinese because of their higher nitrite content. Vegetables contributed the second highest intake of nitrite at 0.248 mg day1 (0.004 mg kg1 body weight, about 13.6% of total nitrite). The other food items, such as rice and products, flour and flour products, and fruits contributed a little to the nitrite intake.
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Food Additives and Contaminants: Part B Gangolli et al. (1994) estimated that the mean daily intakes of nitrite were 1.5 mg kg1 both in the United States and the United Kingdom. In 1994, Van Vliet et al. (1997) estimated the mean intake of nitrite in the Dutch population to be 0.1 mg day1 per person. While in the study of Cornee´ et al. (1992), the mean intake of nitrite in the French was estimated to be 1.88 mg day1 per adult, with 43% of the intake from vegetables, 28% from cured meat, and 16% from cereals. In the present study, cured meat products constituted the major contribution (about 62.3% of total nitrite intake). While according to Japanese studies (Murata and Ishinage 2001; Murata et al. 2002), meat products provided 98% of nitrite intake. The difference may be from the different dietary customs of Western and Eastern customs. The acceptable daily intake (ADI) for nitrite suggested by the FAO/WHO is up to 0.13 mg kg1 day1. It was proposed that this value be changed to 0.07 mg NO2 kg1 day1 based on a more recent study. The Joint FAO/WHO Expert Committee on Food Additives (JECFA) and the Scientific Committee on Food (SCF) have proposed an ADI for NO2 of zero to 0.06 mg kg1 body weight day1. Although the average and maximal nitrite intakes are lower than the ADI suggested by the World Health Organization (WHO), the maximal nitrite intake is a little higher than the ADI suggested by the WHO and JECFA. For those who had consumed too much cured meat products and vegetables, a high nitrite intake must be taken into account.
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Gangolli SD, Van den Brandt PA, Feron VJ, Janzowski C, Koeman JH, Speijers GJA. 1994. Assessment nitrate, nitrite and N-nitroso compounds. Eur J Pharmacol Environ Toxicol Pharmacol. Section. 292:1–28. Hashimo T. 2001. The cause of the abnormal accumulation of nitrite in pickles of Chinese cabbage (Brassica pekinesis Rupr.). J Jpn Soc Food Sci Tech. 48:409–415. Honikel KO. 2008. The use and control of nitrate and nitrite for the processing of meat products. Meat Sci. 78:68–76. Hord NG, Tang Y, Bryan N. 2009. Food sources of nitrates and nitrites: the physiologic context for potential health benefits. Am J Clin Nutr. 90:1–10. Hsu J, Arcot J, Alice Lee N. 2009. Nitrate and nitrite quantification from cured meat and vegetables and their estimated dietary intake in Australians. Food Chem. 115:334–339. Ji FD. 2007. Study on changes of microorganism and the law of formation and reduction of nitrite during vegetable fermentation [Master’s degree in Engineering]. [Beijing (China)]: China Agricultural University. Joossens JV, Hill MJ, Elliott P, Stamler R, Stamler J, Lesaffre E, Dyer A, Nochols R, Kesteloot H. 1996. Dietary salt, nitrate and stomach cancer mortality in 24 countries. Int J Epidemiol. 25:494–504. Mensinga TT, Speijers GJA, Meulenbelt J. 2003. Health implication of exposure to environmental nitrogenous compounds. Toxicol Rev. 22:41–51. Ministry of Health, Ministry of Science and Technology. 2002. The nutrition and health status of the Chinese people. Beijing (China): Ministry of Health, Ministry of Science and Technology. Murata M, Ishinage M. 2001. Daily intakes of nitrate and nitrite in middle-aged men by the duplicate portion method. Shokuhin Eiseigaku Zasshi. 42:215–219. Murata M, Kishida N, Ishinage M. 2002. Survey of the daily intake of nitrate and nitrite in school children by the duplicate portion method. J Food Hyg Soc Jpn. 43:57–61. National Standards. 2005. Maximum levels of contaminants in foods. GB/T 2762–2005. Beijing (China): National Standards. National Standards Analytical Method. 2008. Determination of nitrite and nitrate in foods. GB/T 5009.33-2008. Beijing (China): National Standards. Okafor PN, Ogbonna UI. 2003. Nitrate and nitrite contamination of water sources and fruit juices marketed in South-Eastern Nigeria. J Food Composit Anal. 16:213–218. Onyesom I, Okoh PN. 2006. Quantitative analysis of nitrate and nitrite contents in vegetables commonly consumed in Delta State, Nigeria. Br J Nutr. 96:902–905. Reinik M, Tamme T, Roasto M, Juhkam K, Jurtsˇ enko S, Tenn´o T, Kiis A. 2005. Nitrites, nitrite and N-nitrosoamines in Estonian cured meat products: intake by Estonian children and adolescents. Food Addit Contamin. 22:1098–1105. Sen NP, Baddoo PA. 1977. Trends in the levels of residual nitrite in Canadian cured meat products over the past 25 years. J Agricult Food Chem. 45:4714–4718. Van Vliet JJ, Vaessen HA, Van de Burg G, Schothorst RC. 1997. Twenty-four-hour duplicate diet study 1994; nitrate
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and nitrite: method development and intake per person per day. Cancer Lett. 114:305–307. World Health Organization (WHO). 1996. Toxicological evaluation of certain food additives and contaminants. Food Additives Series No. 35. Geneva (Switzerland): WHO. p. 269–360.
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Food Additives & Contaminants: Part B: Surveillance Publication details, including instructions for authors and subscription information: http://www.tandfonline.com/loi/tfab20
Survey of nitrite content in foods from north-east China a
a
a
a
a
a
a
Y. Yuan , T. Zhang , H. Zhuang , K. Wang , Y. Zheng , H. Zhang , B. Zhou & J. Liu
a
a
Department of Food Quality and Safety , College of Quartermaster Technology, Jilin University , Changchun 130062, China Published online: 18 Mar 2010.
To cite this article: Y. Yuan , T. Zhang , H. Zhuang , K. Wang , Y. Zheng , H. Zhang , B. Zhou & J. Liu (2010) Survey of nitrite content in foods from north-east China, Food Additives & Contaminants: Part B: Surveillance, 3:1, 39-44, DOI: 10.1080/19440040903514515 To link to this article: http://dx.doi.org/10.1080/19440040903514515
PLEASE SCROLL DOWN FOR ARTICLE Taylor & Francis makes every effort to ensure the accuracy of all the information (the “Content”) contained in the publications on our platform. However, Taylor & Francis, our agents, and our licensors make no representations or warranties whatsoever as to the accuracy, completeness, or suitability for any purpose of the Content. Any opinions and views expressed in this publication are the opinions and views of the authors, and are not the views of or endorsed by Taylor & Francis. The accuracy of the Content should not be relied upon and should be independently verified with primary sources of information. Taylor and Francis shall not be liable for any losses, actions, claims, proceedings, demands, costs, expenses, damages, and other liabilities whatsoever or howsoever caused arising directly or indirectly in connection with, in relation to or arising out of the use of the Content. This article may be used for research, teaching, and private study purposes. Any substantial or systematic reproduction, redistribution, reselling, loan, sub-licensing, systematic supply, or distribution in any form to anyone is expressly forbidden. Terms & Conditions of access and use can be found at http:// www.tandfonline.com/page/terms-and-conditions
Food Additives and Contaminants: Part B Vol. 3, No. 1, March 2010, 39–44
View Dataset Survey of nitrite content in foods from north-east China Y. Yuan, T. Zhang, H. Zhuang, K. Wang, Y. Zheng, H. Zhang, B. Zhou and J. Liu* Department of Food Quality and Safety, College of Quartermaster Technology, Jilin University, Changchun 130062, China
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(Received 1 September 2009; final version received 27 November 2009) This study reports a survey of nitrite in a variety of foods consumed in north-east China and estimates the intake of nitrite for the north-east Chinese consumer. A total of 642 food categories including rice and rice products, flour and flour products, soybean and products, vegetables, fruit, preserved vegetables, cured meat products, dairy products, fish products, salt, and soy sauce were analysed for their content of nitrite. Nitrite content was quite different both between different food categories and within the same food category, ranging from not determined (n.d.) to 19.7 mg kg1. A great variation in the content of nitrite was found for all the food products. The average content of nitrite was highest in cured meat products (14.3 mg kg1). Next to that, the nitrite content was high in the order of preserved vegetables (4.1 mg kg1), soybean products (3.5 mg kg1), and dairy products (1.9 mg kg1). The lowest average values of nitrite were detected in soy sauce, rice and rice products, salt and fish products, the contents being 0.1, 0.3, 0.3, and 0.6 mg kg1. Calculations on the basis of these results and including dietary surveys show that the average intake of nitrite in north-east China from food was 0.03 mg kg1 body weight for an average Chinese person weighing 60 kg, and the data are lower than the established acceptable daily intake (ADI) for nitrite. Cured meat products are normally the major contributor to average nitrite intake of the north-east Chinese population. The second contributor is vegetables. Keywords: risk assessment; nitrite
Introduction Nitrite and nitrate occur naturally in food as a consequence of the nitrogen cycle whereby nitrogen is fixed by bacteria. Additionally, nitrate and nitrite are often used as food additives in the form of sodium or potassium salt in modern food processing, especially in meat products, where they fulfil the function of preservation, antimicrobial agent and colour fixative (Reinik et al. 2005). Nitrate and nitrite contamination of vegetables is produced mainly by chemical fertilizers, especially nitrogenous fertilizers (Zhou et al. 2000). The nitrate ion has a low level of acute toxicity, but if transformed into nitrite it may constitute a health problem (Reinik et al. 2005). Research has shown that 5–8% of the nitrate from the diet may be reduced to nitrite by the microflora in the oral cavity (Gangolli et al. 1994; World Health Organization (WHO) 1996; Mensinga et al. 2003). Nitrite is known to be a precursor of toxic and carcinogenic N-nitrosamines (Bassir and Maduagwu 1978) and high dietary intakes of nitrate and nitrite will induce gastric cancer based on epidemiology and clinical studies (Bartsch et al. 1990; Joossens et al. 1996; Sen and Baddoo 1997). Only a few studies have been conducted to estimated the consumption of food additives including *Corresponding author. Email: [email protected] ISSN 1939–3210 print/ISSN 1939–3229 online ß 2010 Taylor & Francis DOI: 10.1080/19440040903514515 http://www.informaworld.com
nitrate and nitrite in China, especially in the north-east (Zhou et al. 2000; Zhong et al. 2002), and have rather focused on the nitrate and nitrite content of vegetables. This research has paid little attention to the contents and effect of cured meat products and other typical Chinese foods on the Chinese population. Thus, it is necessary to learn about the content of nitrite in Chinese foods. This paper presents the results of the content of nitrite in different food categories from north-east China, including rice and rice products, flour and flour products, soybean and products, vegetables, fruit, preserved vegetables, cured meat products, dairy products, fish products, salt, and soy sauce. The daily intake of nitrite from food (exogenous intakes) from north-east Chinese markets was then estimated according to the data obtained here and combined with the average daily consumption of foods. The data could help customers avoid the adverse effects of nitrite consumption.
Materials and methods Chemicals All solvents used in the experiments were analytical grade. The distilled water was free of ions.
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Sampling All samples selected in this survey were purchased at local supermarkets and stores in Changchun city in 2009. A total of 642 samples covered eleven categories of popular foods in north-east China, including rice and rice products, flour and flour products, soybean and products, vegetables, fruit, preserved vegetables, cured meat products, dairy products, fish products, salt, and soy sauce. The samples were stored in a freezer at 18 C until analysis.
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Sample preparation and nitrite analysis Nitrite contents were determined according to the Chinese National Standards Analytical Method (GB/T 5009.33-2008) with slight modification. Before analysis, non-edible parts of the sample were removed, and the whole sample was chopped and frozen. Just before analysis, the samples were homogenized in a blender. Food samples (2.0–10.0 g) were blended with 80 ml of distilled water in a 100 ml volumetric flask. Samples were treated ultrasonically for 30 min, and heated in a water bath at 75 C for up to 5 min, and then made up with distilled water to 100 ml and shaken. The mixtures were centrifuged at 10,000 rpm for 10 min and the supernatant removed for ultra-filtration. The filtrate was used for further nitrite analysis. Nitrite in the extract was reacted with 0.5% sulphanilamide at a concentration of 4 g l1 for 3 min and 0.25% N-(1-naphthyl)ethylene-diamine with a concentration of 2 g l1 was added to the extract and left to stand for 15 min. The absorbance of the violet azo compound was then measured at 540 nm. Chemical analysis for each sample of food was repeated three times. The method was shown to provide an accurate result by analysis of the standard solution of nitrite (ID: GSBZ50006-88) with a concentration of 0.050 mg l1 from the Chinese CRM/RM Information Center (Beijing, China). The result for the nitrite concentration was measured at 0.050, 0.050, 0.051, 0.050, 0.051 and 0.050 mg l1 for six analyses.
Statistical analysis Statistical analysis was performed with a Student’s t-test with SPSS 15.0 software. Analysis of variance (ANOVA) was tested on a significance level of p ¼ 0.05.
Results and discussion Analytical quality assurance The performance of the method was established in terms of its limit of quantification (LOQ), linearity, repeatability or run-to-run precision. Nitrite
(1.0–20.0 mg l1) was spiked in different blank matrices including water, spinach, dried tofu and ham, and recovery values were 89–102%. The LOQ in water and spinach matrix were 0.009 and 0.02 mg kg1, respectively. The good linearity of the method was achieved in a range from zero to 5.00 mg kg1 (r40.99). Precision was evaluated in terms of intra-day repeatability and inter-day reproducibility as RSD%. Both spinach and ham were tested. They had nitrite contents of 2.0 and 13.5 mg kg1, respectively. RSD ranged from 1.6% to 3.5% for the intra-day precision tests (n ¼ 5) and from 2.4% to 6.3% for the inter-day precision tests (n ¼ 15). A standard curve was made before and after analysis. To ensure the stability of the method, one in every ten samples analysed was a standard and this had to be within the 95% confidence interval of the standard curve.
Nitrite content in foods The study was focused on Chinese foods available in Chinese markets. Altogether, 642 food samples were divided into eleven kinds of food categories. These samples were included a wide range of food categories in the north-east China market and a lot of them were traditional Chinese foods. For food category and the numbers of samples within different ranges of nitrite content, see Table 1. Nitrite content was quite different both between different food categories and within the same food category, ranging from not determined (n.d.) to 19.7 mg kg1. The average content of nitrite was highest in cured meat products. Next to that, the nitrite content was high in the sequential order: preserved vegetables, soybean products, and dairy products. The average concentrations of nitrite were 14.3, 4.1, 3.5, and 1.9 mg kg1, respectively. The lowest average values of nitrite were detected in soy sauce, rice and rice products, salt and fish products, the contents being 0.1, 0.3, 0.3, and 0.6 mg kg1. There was a great variation in nitrite contents between the brands in each food groups, as easily seen from the high standard deviations shown in Table 1. Since nitrite can be formed via the reduction of nitrate if the foods are stored incorrectly (Zhong et al. 2002), the high variability of the data is mainly a result of variable food brands, the use of nitrate-based fertilizers, variable sampling places in this survey, and storing places. Ham and sausage were analysed for their nitrite content, and the results are shown in Table 1. The average contents of nitrite in ham and sausage were 16.1 and 12.5 mg kg1, respectively. Nitrite is used as an antimicrobial agent to provide protection against toxic microorganisms and as colouring or flavouring agents to produce the characteristic pink colour, texture and flavour in meat products (Reinik et al. 2005). Since the toxic nature of nitrite and the
Food Additives and Contaminants: Part B
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Table 1. Numbers of samples within different ranges of nitrite content (mg kg1).
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Rice and rice products Flour and flour products Soybean and products Vegetable Fruits Preserved vegetables Cured meat products Dairy products Fish products Salt Soy sauce Total
50.01
0.01–0.5
0.5–1.0
1.0–5.0
5.0–10.0
10–20
420
Number
0 0 0 0 1 0 0 0 0 0 0 1
11 12 0 170 20 1 0 9 3 6 6 238
0 0 0 105 3 3 0 0 1 0 0 112
1 6 24 79 24 61 3 15 2 0 0 215
0 0 3 3 0 23 11 0 0 0 0 40
0 0 0 0 0 2 31 0 0 0 0 33
0 0 0 0 0 0 3 0 0 0 0 3
12 18 27 357 48 90 48 24 6 6 6 642
formation of nitrosamines by nitrite in cured meat products, many governments have established maximum levels for nitrite in many kinds of food. The European Parliament and its Council Directive have established maximum levels for nitrate and nitrite of 150 and 300 mg kg1, respectively. Residual amounts should not exceed 50 mg kg1 nitrite in non-heattreated meat products and 100 mg kg1 nitrite in all other meat products, except Wiltshire bacon and some other similar products which are limited to 175 mg kg1 nitrite (Honikel 2008). In China, the tolerance limit for nitrite is 20 mg kg1 for ham and sausage (National Standards 2005). In the present survey, there were three samples exceeding the Chinese tolerance limit. Since the difference in using nitrite and nitrate and using methods for those two kinds of additives, a little difference could found between the present study and that of Sen and Baddoo (1997) in sausage and ham in Canada. The average content of nitrite in their studies was 26 mg kg1 in sausage and 24 mg kg1 in ham, respectively. However, in the present survey the maximum contents of residual nitrite was 19.7 mg kg1 in ham and 15.9 mg kg1 in sausage, respectively. Since some manufacturers add less nitrite but more nitrate as a nitrite reserve (Hsu et al. 2009), this might also explain the differences in the findings by different researchers. Preserved vegetables are traditional Chinese food and many north-east Chinese people like to eat preserved vegetables in their daily lives, especially in winter. In the present survey, nitrite was found in preserved vegetables (4.1 mg kg1 on average) such as pickled vegetables, pickled Chinese cabbage and Chinese pickled vegetables. The average contents of nitrite were 2.6, 2.7 and 4.3 mg kg1, respectively. In China, the tolerance limit for nitrite is 20 mg kg1 in preserved vegetables (National Standards 2005). In the present survey, none of the samples contained over 20 mg kg1. But studies showed that nitrates in soil are absorbed into certain vegetables, for instance radish
and cabbage, which have a higher content of nitrate. The pickling process makes the conversion from nitrates to nitrites possible, resulting from the reduction by Escherichia, Proteus, Salmonella and so forth. Nitrite content increases as temperature increases in the pickling process. Also, salt concentration in the substrate has a significant influence. Nitrites reach a maximum at about 37 C when 5% salt is available. The nitrite content varies, increasing during the second to the fourth days, reaching a maximum during the seventh to eighth days, while dropping markedly 20 days later. While the decomposition by lactic acid bacteria to the nitrite in preserved vegetables was divided into enzymolysis and acid splitting, in the early period of fermentation, when the fermentation soup’s pH was greater than 4.5, enzymolysis was the main controlling factor, but when the pH was less than 4.0 because the amount of acid was increasing, acid splitting became the controlling factor. The nitritedecomposing ability of lactobacillus was stronger than lactococcus because of its acid-producing ability (Hashimo 2001; Ji 2007). As vegetables are consumed in large amounts in north-east China, the number of samples included in the study was the largest. A total of 357 vegetables were analysed for nitrite content. The average content of nitrite in vegetables was 0.9 mg kg1, and cherry tomato had the highest nitrite content (2.7 mg kg1) compared with other vegetables, and the maximum content was reached at 4.1 mg kg1. Spinach also had a high content of nitrite (2.1 mg kg1 on average) in the selected vegetables, and the maximum content was reached at 4.8 mg kg1. The finding correlated well with the study of Chung et al. (2003), who also found a high nitrite content in spinach (1.0 mg kg1 on average) in Korea. The difference between north-east China and Korea may be because of the varieties and use of nitrate-based fertilizers (Hsu et al. 2009). A high standard deviation could also be found in nitrite content from different vegetable varieties because
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cultivar and harvest date can affect the nitrite levels of selected vegetables (Amr and Hadidi 2001). This might explain the high variability between findings presented in the present study also contributing to a high standard deviation as observed in Table 1. Garlic contained the lowest amount of nitrite in this study (0.1 mg kg1 on average), which was lower that that in the study of Chung et al. (2003) which demonstrated a nitrite content in garlic of 0.2 mg kg1 on average. The difference may be due to cultivation conditions. A total of 48 fruits were also analysed for nitrite content, including banana, pear, apple, and watermelon. The average content of nitrite was 1.3 mg kg1; apple had the highest nitrite content in the selected fruits (2.0 mg kg1 on average). Thus, it can be concluded that apple contributed to the highest dietary nitrite intake from fruits. Next were watermelon and banana. The average concentrations of nitrite were 2.0 and 1.7 mg kg1, respectively. Dogan et al. (2008) also found that nitrite could exist in apple and pear products. Okafor and Ogbonna (2003) found some nitrite content in fruit juices. Hord et al. (2009) reported that about 0.09 mg kg1 of nitrite could be detected in banana. The present study has found for the first time nitrite content in watermelon. The results might be the accumulation of nitrate in the soil or nitrate-rich soil, which would transfer the nitrite during growth of the watermelon (Onyesom and Okoh 2006). For statistical evaluation of the data about the content of nitrite obtained in different food categories of north-east Chinese food, a box-and-whisker plot (generated by OriginPro 7.5 software) was used. The plot is a powerful statistical tool that displays the median, range, minimum, maximum, mean, 25th percentile and 75th percentile. It is necessary since the analytical results are not normally distributed, especially in this kind of survey. Nitrite contents in different food categories are varied, and the median and inter-quartile range are two appropriate statistics to describe the central tendency and the spread around the median, respectively. Figure 1 shows that the nitrite content in different food categories ranges quite markedly. Nitrite levels in most food categories were less than 5 mg kg1, except for cured meat products from the median of the data. And from the median, one can also see that most of the data lie toward the lower rather than the higher end of the range. This box plot offered a great deal more insight than a lone average, and much more than an average complemented by the low and high data.
Exposure estimation and risk assessment of nitrite in Chinese foods Nitrite can react with nitrosatable compounds, such as amides and amines, to form N-nitroso compounds.
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Nitrite content (mg kg–1)
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20
10
0 A
B
C
D
E F G H Food categories
I
J
K
Figure 1. Nitrite contents in north-east Chinese foods. A, Rice and rice products; B, flour and flour products; C, soybean and products; D, vegetables; E, fruits; F, preserved vegetables; G, poultry; H, dairy products; I, fish and products; J, salt; and K, soy sauce. The centre horizontal line of the box is the median of the data; the top and bottom of the box are the 25th and 75th percentiles (quartiles); and the ends of the whiskers are the 10th and 90th percentiles. Any points outside are considered outliers, which are labelled as ‘X’. The small square of each box means the average of the data.
Some N-nitroso compounds are potent carcinogens in animal species, and therefore can be carcinogenic to man (Onyesom and Okoh 2006). Once the nitrite contents in food were established, one can estimate the intake of these compounds based on national dietary surveys. The average content of nitrite in foods was used in combination with the average daily consumption of foods. The average consumption of different kind foods in China was completed with The Nutrition and Health Status of the Chinese People (Ministry of Health, Ministry of Science and Technology 2002). In the present survey the average dietary exposure to nitrite in the north-east Chinese population was estimated to be 1.826 mg day1 and was 0.030 mg kg1 body weight for an average Chinese person weighing 60 kg. The maximum intake of nitrite was 0.088 mg kg1 body weight. Simultaneously, the contributions of different food categories to the total nitrite intake were ranked. Cured meat products constituted the major contribution at 1.138 mg day1 (0.019 mg kg1 body weight, about 62.3% of total nitrite) to the average nitrite intake of the north-east Chinese because of their higher nitrite content. Vegetables contributed the second highest intake of nitrite at 0.248 mg day1 (0.004 mg kg1 body weight, about 13.6% of total nitrite). The other food items, such as rice and products, flour and flour products, and fruits contributed a little to the nitrite intake.
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Food Additives and Contaminants: Part B Gangolli et al. (1994) estimated that the mean daily intakes of nitrite were 1.5 mg kg1 both in the United States and the United Kingdom. In 1994, Van Vliet et al. (1997) estimated the mean intake of nitrite in the Dutch population to be 0.1 mg day1 per person. While in the study of Cornee´ et al. (1992), the mean intake of nitrite in the French was estimated to be 1.88 mg day1 per adult, with 43% of the intake from vegetables, 28% from cured meat, and 16% from cereals. In the present study, cured meat products constituted the major contribution (about 62.3% of total nitrite intake). While according to Japanese studies (Murata and Ishinage 2001; Murata et al. 2002), meat products provided 98% of nitrite intake. The difference may be from the different dietary customs of Western and Eastern customs. The acceptable daily intake (ADI) for nitrite suggested by the FAO/WHO is up to 0.13 mg kg1 day1. It was proposed that this value be changed to 0.07 mg NO2 kg1 day1 based on a more recent study. The Joint FAO/WHO Expert Committee on Food Additives (JECFA) and the Scientific Committee on Food (SCF) have proposed an ADI for NO2 of zero to 0.06 mg kg1 body weight day1. Although the average and maximal nitrite intakes are lower than the ADI suggested by the World Health Organization (WHO), the maximal nitrite intake is a little higher than the ADI suggested by the WHO and JECFA. For those who had consumed too much cured meat products and vegetables, a high nitrite intake must be taken into account.
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