玉米中赭曲霉毒素A辐照降解技术研究
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摘要
目前,农产品中真菌毒素的污染严重危害人和动物的健康,受到广泛的关注。食品辐照技术是20世纪发展起来的一种新兴技术,经过长时间的研究与实践应用,食品辐照技术已被证明是一种高效、无污染的保藏手段。由于食品辐照技术在处理过程中不需要加入任何化学药品,不会产生二次污染,另外辐照一般在常温常压条件下进行,工艺简单,效率较高。食品辐照技术被认为是解决农产品真菌毒素污染、保障食用安全的重要的手段。
本论文的研究目的是探讨OTA的辐照降解效果,确定辐解产物的结构以及对产物进行毒性评价,从而证实辐照技术能够有效地控制农产品中的OTA,为辐照技术应用于其他真菌毒素降解提供一定的参考价值。本文选择较有代表性的真菌毒素-OTA为研究对象,选取水溶液为模型,以玉米为基质,研究了水溶液中OTA的辐照降解效果、辐解产物的结构以及辐解产物的毒性记录、玉米中OTA的辐照降解效果、经过辐照后玉米营养成分的变化以及辐解产物是否有毒等关键问题。具体研究结论如下:
(1)探讨了γ射线辐照对水溶液中OTA的降解效果。用OTA标准溶液确定高效液相色谱检测技术的线性关系和检出限,采用外标法定量,研究了辐照对水溶液中OTA的降解效果。结果表明,水溶液中OTA的降解率随着辐照剂量的增加而增加,但降解率与辐照剂量之间不呈现显著的正比关系,降解率随辐照剂量的变化可以由多项式方程来拟合:y=ax~2+bx+c;在4 kGy的辐照剂量下,OTA的降解率可达90%,此后降解率的增长随着辐照剂量的增加而趋于缓慢;且水溶液中OTA初始浓度越低,相同辐照剂量下降解率越高;有机溶剂浓度不同,OTA的降解率也不同,有机溶剂的浓度越高,其OTA的降解率越低。
(2)解析出水溶液中OTA的5种主要辐解产物并对产物的毒性进行评价。以LC/TOFMS作为分析OTA辐解产物的方法解析出水溶液中OTA的5种主要辐解产物结构,其中一种辐解产物也是OTA的水解产物,3种产物是OTA类似物,剩下的一种产物没有报道。通过检索相关毒性数据库评价产物的毒性,结果发现,5种产物中,只有一种产物有毒性记录,且毒性远远低于OTA的毒性,其他4种产物均没有毒性记录。可以推测,在痕量水平下,产物的毒性可以忽略。
(3)研究了γ射线辐照对玉米中OTA的降解效果。以污染OTA严重的玉米为研究对象,其中,玉米中OTA的含量为71μg/kg,采用γ射线辐照,以高效液相色谱法定量检测,研究了不同辐照剂量下玉米中OTA的降解率以及经过辐照后玉米营养成分的变化。结果表明,随着辐照剂量的增加,玉米中OTA的降解率逐渐增加,在10kGy时,玉米中OTA的降解率可达到50%。经过高、中、低剂量辐照后的玉米的营养成分,水分、粗蛋白、粗脂肪和淀粉的含量与辐照前没有发生显著的变化,表明辐照处理能够降解玉米中OTA,且不会降低玉米的营养成分。
(4)由于玉米成分的复杂性,考虑到经过辐照后无法解析玉米中OTA辐解产物的结构,故而采用毒理学实验的方法,确定OTA以及辐解产物的毒性。结果显示:OTA对小鼠生长状况有一定的影响,例如小鼠的体重增加量和食物利用率减少,而OTA降解产物对上述指标基本无影响;OTA还会造成小鼠血液中细胞数量和一些生化指标的显著变化,破坏了血小板和淋巴细胞,造成血液中AST、GLU、BUN显著升高,LDH显著降低;OTA对小鼠的肝脏,肾脏,肺脏有一定的损害,但不会损伤脾脏和小肠,而未发现OTA降解产物对小鼠脏器的损伤。实验结果表明,OTA对小鼠有一定的毒害作用,而OTA降解产物不会对小鼠造成损伤,说明辐照能够有效地降解OTA,且降解后的产物没发现有毒害作用。
目前,国外对于农产品中OTA的辐照降解研究尚处于初级阶段,且只注重OTA的降解率,而对降解产物的结构及毒性均没有开展相关的研究;国内在此领域的研究更是一片空白。本研究不仅研究了辐照处理OTA的降解效果和规律,还详尽地对主要辐解产物进行结构解析,同时对辐解产物的毒性进行了评价,这为今后研究其他真菌毒素物理消解方法奠定了理论基础。
The mycotoxin in agricultural products has concerned us more and more as its great harm for the health of humans and animals. The technology of food irradiation, a boom technology in 20th century, is proved to be a measure of preservation with high efficiency and no pollution after long-term researches and applications. Food irradiation could be an important technology to deal with mycotoxin pollution in agriculture products in order to ensure safety, there is no add-in of chemicals in the process of food irradiation, so this technology could not cause pollution repeated, what’s more, the irradiation is carried out under normal pressure and temperature which can bring out high efficiency.
The effect ofγ-irradiation on degradation of Ochratoxin A was researched on the basis of products structure and their toxicity in order to provide reference for application of irradiation in degradation of mycotoxin, which could control OTA pollution in agricultural products. OTA was taken as an example of mycotoxin, then the effects ofγ-irradiation on degradation of Ochratoxin A in aqueous solution, products structure and toxicity in aqueous solution, the effects ofγ-irradiation on degradation of Ochratoxin A in maize, products structure and toxicity in maize, the change of maize quality and the toxicity of products were evaluated. The main research results are followed:
(1) OTA in aqueous solution was irradiated byγ-rays to evaluate the irradiation degradation effects. The standard aqueous solution of OTA was applied to determine the linear relationship and detection limit of HPLC, with the external standard method for quantitative analysis, which was used to investigate the effects of absorbed doses on the degradation of OTA in aqueous solution. The results showed that the degradation rate of OTA in aqueous solution increases with the absorbed dose in certain range with no proportional. The degradation rate changed with irradiation doses by polynomial equation to fitting: y=ax~2+bx+c. The degradation rate reached the maximum, 90%, at 4 kGy, and then it tended to be stable. What’s more, the degradation rate was higher with lower concentration of OTA in aqueous solution. As a result, 60Coγ-rays could have good effect on the degradation of OTA in aqueous solution with lower absorbed dose.
Five main radiolysis products of OTA degradation were analyzed and their toxicity was also evaluated. LC/TOF MS (Liquid -Chromatography-Quadrupole-Time-of-Flight Mass Spectrometry) was applied for the analysis of five radiolysis products. The results showed that one product was OTA hydrolysate, three products were OTA analogs, and the product left hadn’t been reported. The search results in related toxicity databases indicated that there was only one product with toxicity much lower than OTA and other four products no toxicity. It was speculated that the toxicity of radiolysis products of OTA degradation could be ignored.
To evaluate the effects ofγ-irradiation on degradation of OTA in maize, maize samples contaminated with OTA was irradiated byγray firstly, while content of OTA in maize was 71μg/kg, then the quantity of OTA in samples was assayed with HPLC which was compared at different doses, and the nutritional components were also detected. The result showed that he content of OTA in maize decreased significantly, and the rate of degradation of OTA could reach 50% at the dose of 10kGy. The nutritional components were not destroyed byγ-irradiation.γ-irradiation could degrade OTA in maize with fine rate of degradation and not decrease the quality of maize.
Considered that the products structure of OTA irradiated was difficult to analyze for complicated components in maize, toxicology experiments were arranged to determine the toxicity of OTA and its products after irradiation. Results: OTA could affect growing status of mice such as the augment of body weight and the rate of food utilization, compared with that the radiolysis products of OTA degradation brought out no effects. OTA could destroy blood platelet and lymphocyte of mice, increase AST, GLU, BUN in mice serum and decrease LDH. Some viscera of mice could be shattered by OTA such as liver, kidney and lung, except spleen. But the radiolysis products of OTA degradation have no effects on all the viscera. It was indicated that OTA has toxicity on mice and the radiolysis products of OTA degradation has no such toxicity. Therefore, OTA could be irradiated with high efficiency and no toxicity.
Foreign researches on OTA irradiation are just primary currently, which focus only on the rate of OTA irradiation, and there are no related studies on the products structure of OTA irradiated and their toxicity all over the world. The effects and disciplinarian ofγ-irradiation on degradation of Ochratoxin A were explored in our research with detail, as well as the analysis of these products and toxicology experiments, which can provide academic foundation for irradiation of other mycotoxin.
本论文的研究目的是探讨OTA的辐照降解效果,确定辐解产物的结构以及对产物进行毒性评价,从而证实辐照技术能够有效地控制农产品中的OTA,为辐照技术应用于其他真菌毒素降解提供一定的参考价值。本文选择较有代表性的真菌毒素-OTA为研究对象,选取水溶液为模型,以玉米为基质,研究了水溶液中OTA的辐照降解效果、辐解产物的结构以及辐解产物的毒性记录、玉米中OTA的辐照降解效果、经过辐照后玉米营养成分的变化以及辐解产物是否有毒等关键问题。具体研究结论如下:
(1)探讨了γ射线辐照对水溶液中OTA的降解效果。用OTA标准溶液确定高效液相色谱检测技术的线性关系和检出限,采用外标法定量,研究了辐照对水溶液中OTA的降解效果。结果表明,水溶液中OTA的降解率随着辐照剂量的增加而增加,但降解率与辐照剂量之间不呈现显著的正比关系,降解率随辐照剂量的变化可以由多项式方程来拟合:y=ax~2+bx+c;在4 kGy的辐照剂量下,OTA的降解率可达90%,此后降解率的增长随着辐照剂量的增加而趋于缓慢;且水溶液中OTA初始浓度越低,相同辐照剂量下降解率越高;有机溶剂浓度不同,OTA的降解率也不同,有机溶剂的浓度越高,其OTA的降解率越低。
(2)解析出水溶液中OTA的5种主要辐解产物并对产物的毒性进行评价。以LC/TOFMS作为分析OTA辐解产物的方法解析出水溶液中OTA的5种主要辐解产物结构,其中一种辐解产物也是OTA的水解产物,3种产物是OTA类似物,剩下的一种产物没有报道。通过检索相关毒性数据库评价产物的毒性,结果发现,5种产物中,只有一种产物有毒性记录,且毒性远远低于OTA的毒性,其他4种产物均没有毒性记录。可以推测,在痕量水平下,产物的毒性可以忽略。
(3)研究了γ射线辐照对玉米中OTA的降解效果。以污染OTA严重的玉米为研究对象,其中,玉米中OTA的含量为71μg/kg,采用γ射线辐照,以高效液相色谱法定量检测,研究了不同辐照剂量下玉米中OTA的降解率以及经过辐照后玉米营养成分的变化。结果表明,随着辐照剂量的增加,玉米中OTA的降解率逐渐增加,在10kGy时,玉米中OTA的降解率可达到50%。经过高、中、低剂量辐照后的玉米的营养成分,水分、粗蛋白、粗脂肪和淀粉的含量与辐照前没有发生显著的变化,表明辐照处理能够降解玉米中OTA,且不会降低玉米的营养成分。
(4)由于玉米成分的复杂性,考虑到经过辐照后无法解析玉米中OTA辐解产物的结构,故而采用毒理学实验的方法,确定OTA以及辐解产物的毒性。结果显示:OTA对小鼠生长状况有一定的影响,例如小鼠的体重增加量和食物利用率减少,而OTA降解产物对上述指标基本无影响;OTA还会造成小鼠血液中细胞数量和一些生化指标的显著变化,破坏了血小板和淋巴细胞,造成血液中AST、GLU、BUN显著升高,LDH显著降低;OTA对小鼠的肝脏,肾脏,肺脏有一定的损害,但不会损伤脾脏和小肠,而未发现OTA降解产物对小鼠脏器的损伤。实验结果表明,OTA对小鼠有一定的毒害作用,而OTA降解产物不会对小鼠造成损伤,说明辐照能够有效地降解OTA,且降解后的产物没发现有毒害作用。
目前,国外对于农产品中OTA的辐照降解研究尚处于初级阶段,且只注重OTA的降解率,而对降解产物的结构及毒性均没有开展相关的研究;国内在此领域的研究更是一片空白。本研究不仅研究了辐照处理OTA的降解效果和规律,还详尽地对主要辐解产物进行结构解析,同时对辐解产物的毒性进行了评价,这为今后研究其他真菌毒素物理消解方法奠定了理论基础。
The mycotoxin in agricultural products has concerned us more and more as its great harm for the health of humans and animals. The technology of food irradiation, a boom technology in 20th century, is proved to be a measure of preservation with high efficiency and no pollution after long-term researches and applications. Food irradiation could be an important technology to deal with mycotoxin pollution in agriculture products in order to ensure safety, there is no add-in of chemicals in the process of food irradiation, so this technology could not cause pollution repeated, what’s more, the irradiation is carried out under normal pressure and temperature which can bring out high efficiency.
The effect ofγ-irradiation on degradation of Ochratoxin A was researched on the basis of products structure and their toxicity in order to provide reference for application of irradiation in degradation of mycotoxin, which could control OTA pollution in agricultural products. OTA was taken as an example of mycotoxin, then the effects ofγ-irradiation on degradation of Ochratoxin A in aqueous solution, products structure and toxicity in aqueous solution, the effects ofγ-irradiation on degradation of Ochratoxin A in maize, products structure and toxicity in maize, the change of maize quality and the toxicity of products were evaluated. The main research results are followed:
(1) OTA in aqueous solution was irradiated byγ-rays to evaluate the irradiation degradation effects. The standard aqueous solution of OTA was applied to determine the linear relationship and detection limit of HPLC, with the external standard method for quantitative analysis, which was used to investigate the effects of absorbed doses on the degradation of OTA in aqueous solution. The results showed that the degradation rate of OTA in aqueous solution increases with the absorbed dose in certain range with no proportional. The degradation rate changed with irradiation doses by polynomial equation to fitting: y=ax~2+bx+c. The degradation rate reached the maximum, 90%, at 4 kGy, and then it tended to be stable. What’s more, the degradation rate was higher with lower concentration of OTA in aqueous solution. As a result, 60Coγ-rays could have good effect on the degradation of OTA in aqueous solution with lower absorbed dose.
Five main radiolysis products of OTA degradation were analyzed and their toxicity was also evaluated. LC/TOF MS (Liquid -Chromatography-Quadrupole-Time-of-Flight Mass Spectrometry) was applied for the analysis of five radiolysis products. The results showed that one product was OTA hydrolysate, three products were OTA analogs, and the product left hadn’t been reported. The search results in related toxicity databases indicated that there was only one product with toxicity much lower than OTA and other four products no toxicity. It was speculated that the toxicity of radiolysis products of OTA degradation could be ignored.
To evaluate the effects ofγ-irradiation on degradation of OTA in maize, maize samples contaminated with OTA was irradiated byγray firstly, while content of OTA in maize was 71μg/kg, then the quantity of OTA in samples was assayed with HPLC which was compared at different doses, and the nutritional components were also detected. The result showed that he content of OTA in maize decreased significantly, and the rate of degradation of OTA could reach 50% at the dose of 10kGy. The nutritional components were not destroyed byγ-irradiation.γ-irradiation could degrade OTA in maize with fine rate of degradation and not decrease the quality of maize.
Considered that the products structure of OTA irradiated was difficult to analyze for complicated components in maize, toxicology experiments were arranged to determine the toxicity of OTA and its products after irradiation. Results: OTA could affect growing status of mice such as the augment of body weight and the rate of food utilization, compared with that the radiolysis products of OTA degradation brought out no effects. OTA could destroy blood platelet and lymphocyte of mice, increase AST, GLU, BUN in mice serum and decrease LDH. Some viscera of mice could be shattered by OTA such as liver, kidney and lung, except spleen. But the radiolysis products of OTA degradation have no effects on all the viscera. It was indicated that OTA has toxicity on mice and the radiolysis products of OTA degradation has no such toxicity. Therefore, OTA could be irradiated with high efficiency and no toxicity.
Foreign researches on OTA irradiation are just primary currently, which focus only on the rate of OTA irradiation, and there are no related studies on the products structure of OTA irradiated and their toxicity all over the world. The effects and disciplinarian ofγ-irradiation on degradation of Ochratoxin A were explored in our research with detail, as well as the analysis of these products and toxicology experiments, which can provide academic foundation for irradiation of other mycotoxin.
引文
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