【摘要】：霉菌毒素是霉菌在基質(zhì)(飼料、飼料原料等)上生長(zhǎng)的過(guò)程中所產(chǎn)生的有毒的次級(jí)代謝產(chǎn)物,包括霉菌使基質(zhì)(飼料)的成分轉(zhuǎn)變而形成的有毒物質(zhì)。霉菌毒素能夠干擾許多靶器官和系統(tǒng),尤其是肝、腎、神經(jīng)系統(tǒng)、內(nèi)分泌系統(tǒng)和免疫系統(tǒng),引發(fā)人類和動(dòng)物疾病。研究表明當(dāng)植物受到霉菌毒素的侵害時(shí),會(huì)進(jìn)行自我保護(hù),將非極性的毒素與糖、氨基酸或硫酸鹽等結(jié)合,轉(zhuǎn)化為極性更強(qiáng)的代謝產(chǎn)物,儲(chǔ)存在液泡中或結(jié)合在器官、組織、細(xì)胞器等的生物大分子上,從而產(chǎn)生霉菌毒素的結(jié)合態(tài)。常規(guī)的霉菌毒素檢測(cè)方法只能檢測(cè)到游離態(tài)的霉菌毒素,而檢測(cè)不到結(jié)合態(tài)的霉菌毒素,因而這部分結(jié)合態(tài)的毒素也被稱為隱蔽性毒素。然而研究表明結(jié)合態(tài)的霉菌毒素經(jīng)動(dòng)物消化后會(huì)被部分或全部釋放出來(lái),危害畜禽的健康。因此,建立一種快速真實(shí)地評(píng)定飼料及飼料原料中霉菌毒素(游離態(tài)和結(jié)合態(tài))的含量顯得尤為重要。本研究通過(guò)3個(gè)試驗(yàn)分別研究了霉菌毒素的體內(nèi)外釋放測(cè)定,在此基礎(chǔ)上分析了現(xiàn)有的霉菌毒素吸附劑對(duì)黃曲霉毒素B1、玉米赤霉烯酮和嘔吐毒素的吸附效率,并開展了新型吸附劑的開發(fā)研究。為了能夠真實(shí)地評(píng)定結(jié)合態(tài)和游離態(tài)的霉菌毒素,本試驗(yàn)建立了體外消化評(píng)定法。通過(guò)體外消化法模擬動(dòng)物體內(nèi)的消化過(guò)程對(duì)發(fā)霉玉米進(jìn)行消化。玉米樣品在60目的篩下物樣品,先后用胃蛋白酶和胰蛋白酶進(jìn)行體外消化,然后再用ELISA的方法對(duì)消化前后的霉菌毒素進(jìn)行檢測(cè)。試驗(yàn)結(jié)果表明玉米中的霉菌毒素AFB1 (P0.001)、 DON (P0.01)、ZEN (P0.05)經(jīng)過(guò)體外消化后毒素含量會(huì)明顯升高。采用了高效液相色譜法對(duì)ELISA檢測(cè)的結(jié)果進(jìn)行了驗(yàn)證,試驗(yàn)結(jié)果表明表明玉米中的霉菌毒素AFB1 (P0.05)、DON (P0.001)、ZEN (P0.001)經(jīng)過(guò)體外消化后毒素含量會(huì)明顯升高,高效液相色譜法進(jìn)一步驗(yàn)證了玉米中的霉菌毒素AFB1、DON、ZEN經(jīng)過(guò)體外消化后毒素含量會(huì)明顯升高。為了驗(yàn)證體外消化試驗(yàn)結(jié)果的真實(shí)性,還進(jìn)行了肉雞體內(nèi)消化試驗(yàn)來(lái)驗(yàn)證體外消化試驗(yàn)的結(jié)果。試驗(yàn)挑選10只體重基本一致的健康A(chǔ)A肉仔雞,單籠飼養(yǎng),禁食12h,自由飲水,自由采食含1%三氧化二鉻的發(fā)霉玉米4h,采集十二指腸、空腸、回腸內(nèi)容物。分別檢測(cè)飼料、十二指腸內(nèi)容物、空腸內(nèi)容物、回腸內(nèi)容物中的三氧化二鉻(濕法消化法)及霉菌毒素AFB1、DON、ZEN含量(ELISA)。試驗(yàn)結(jié)果表明發(fā)霉玉米經(jīng)肉雞體內(nèi)消化后AFB1 (P0.05)、DON (P0.05)、ZEN (P0.01)的含量顯著升高。上述結(jié)果表明：霉菌毒素在玉米體內(nèi)存在結(jié)合態(tài)和游離態(tài)兩種形式,結(jié)合態(tài)霉菌毒素經(jīng)消化后會(huì)被充分的釋放出來(lái),轉(zhuǎn)變?yōu)橛坞x態(tài)的霉菌毒素。為了真實(shí)地評(píng)價(jià)霉菌毒素吸附劑的吸附效果,本試驗(yàn)還通過(guò)體外消化評(píng)定法對(duì)市場(chǎng)上收集的10種霉菌毒素吸附劑的吸附效果進(jìn)行了評(píng)定,試驗(yàn)結(jié)果表明3#、9#吸附劑對(duì)三種毒素均有較好的吸附效果,其它吸附劑對(duì)各個(gè)毒素的吸附效果各有側(cè)重,3#、9#是酵母細(xì)胞產(chǎn)品,這說(shuō)明酵母細(xì)胞產(chǎn)品對(duì)霉菌毒素的吸附效果較好。為了有效地去除飼料中的霉菌毒素,本課題研發(fā)了一種新型經(jīng)濟(jì)有效的霉菌毒素吸附劑,并用體外消化評(píng)定法對(duì)該吸附劑的吸附效果進(jìn)行評(píng)價(jià)。試驗(yàn)結(jié)果表明：該吸附劑對(duì)黃曲霉毒素B1(AFB1)的吸附率達(dá)49.17%,對(duì)嘔吐毒素(DON)的吸附率達(dá)67.27%。
[Abstract]:The mycotoxin is a toxic secondary metabolite produced during the growth of a mold on a substrate (feed, feed, etc.), including a toxic substance formed by the transformation of the components of the substrate (feed) by mold. The mycotoxin can interfere with many of the target organs and systems, in particular the liver, the kidney, the nervous system, the endocrine system and the immune system, triggering human and animal diseases. The research shows that when the plant is attacked by the mycotoxin, self-protection is carried out, the non-polar toxin is combined with sugar, amino acid or sulfate and the like, converted into a more polar metabolite, stored in the vacuole or combined with the biological macromolecules such as organs, tissues, organelles, and the like, so as to produce the bound state of the mycotoxin. The conventional mycotoxin detection method can only detect the mycotoxin of the free state, but the mycotoxin of the bound state can not be detected, and thus the toxin of the bound state is also referred to as the hidden toxin. However, the study shows that the bound mycotoxin can be partially or completely released after the animal is digested, which is harmful to the health of the livestock and poultry. Therefore, it is very important to establish a fast and realistic evaluation of the content of mycotoxins (free and bound) in feed and feed materials. In this study, the in vitro release of mycotoxin was studied by three experiments, and the adsorption efficiency of the existing mycotoxin adsorbent to the aflatoxin B1, the zearalenone and the Vomiting toxin was analyzed, and the development and research of the new type of adsorbent were also carried out. In order to be able to truly assess the bound and free mycotoxins, an in vitro digestion and evaluation method was established. the musty corn was digested by an in vitro digestion process to simulate the digestion process in the animal. The corn sample was digested with pepsin and trypsin in vitro at 60 mesh, and then the mycotoxin before and after digestion was detected by ELISA. The results showed that the content of mycotoxin AFB1 (P0.01), DON (P0.01) and ZEN (P0.05) in maize increased significantly after in vitro digestion. The results of ELISA were validated by high performance liquid chromatography. The results showed that the content of mycotoxin AFB1 (P0.05), DON (P0.01) and ZEN (P0.001) in maize increased significantly after in vitro digestion. The content of mycotoxin AFB1, DON and ZEN in maize was increased significantly after in vitro digestion with high performance liquid chromatography. In order to verify the authenticity of the in vitro digestion test results, the in vivo digestion test was conducted to verify the results of the in vitro digestion test. 10 healthy AA broilers with almost the same body weight were selected for feeding, fasting for 12h, free drinking water, and free feed of musty corn with 1% chromium oxide for 4h, and the contents of the duodenum, the jejunum and the ileum were collected. The contents of the feed, the contents of the duodenum, the contents of the jejunum, the chromium trioxide in the contents of the ileum (wet digestion method) and the mycotoxin AFB1, DON and ZEN (ELISA) were respectively detected. The results showed that the content of AFB1 (P0.05), DON (P0.05) and ZEN (P0.01) increased significantly after the body was digested with musty corn. The above results show that the mycotoxin is in the form of binding and free state in the corn, and the bound mycotoxin can be fully released after being digested and transformed into a free-form mycotoxin. In order to evaluate the adsorption effect of the mycotoxin adsorbent, the adsorption effect of 10 kinds of mycotoxin adsorbent collected on the market was evaluated by in vitro digestion and evaluation. The results showed that 3 # and 9 # adsorbent had good adsorption effect on the three kinds of toxins. The effect of other adsorbents on the adsorption of each toxin is different, and the 3 # and 9 # are the product of the yeast cell, which indicates that the yeast cell product has good adsorption effect on the mycotoxin. In order to effectively remove the mycotoxin from the feed, a new type of economical and effective mycotoxin adsorbent was developed and the adsorption effect of the adsorbent was evaluated by in vitro digestion and evaluation. The results showed that the adsorption rate of Aflatoxin B1 (AFB1) was 49. 17%, and the adsorption rate of VON was 67. 27%.
【學(xué)位授予單位】：山東農(nóng)業(yè)大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2015
【分類號(hào)】：S816;S859.8

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