本文關(guān)鍵詞： 黑水虻 抗菌肽 誘導(dǎo) 抑菌活性 生物學(xué)活性　出處：《遵義醫(yī)學(xué)院》2012年碩士論文　論文類型：學(xué)位論文

【摘要】：目的：采用不同誘導(dǎo)方法對(duì)黑水虻(Hermetia illucens L.)幼蟲抗菌肽進(jìn)行誘導(dǎo),并對(duì)所提取粗提物進(jìn)行活性研究,探討黑水虻幼蟲抗菌肽粗提物的抑菌活性及生物學(xué)活性。期待通過(guò)本研究豐富昆蟲抗菌肽的種類,并為黑水虻抗菌肽的研發(fā)及利用提供有效的實(shí)驗(yàn)方法。 方法：本研究利用大腸桿菌菌液針刺浸泡誘導(dǎo)法誘導(dǎo)黑水虻幼蟲,通過(guò)研磨提取法提取幼蟲抗菌肽粗提物確定最佳誘導(dǎo)齡期；采用大腸桿菌菌液、金黃色葡萄球菌菌液注射誘導(dǎo)法,大腸桿菌、金黃色葡萄球菌菌液針刺浸泡誘導(dǎo)法,不同功率、不同時(shí)間超聲波誘導(dǎo)法對(duì)黑水虻5齡幼蟲進(jìn)行抗菌肽誘導(dǎo)。利用研磨提取法提取抗菌肽粗提物,并對(duì)粗提物分別進(jìn)行抑菌活性研究。選用抑菌效果最佳的抗菌肽粗提物進(jìn)行生物學(xué)活性鑒定及利用反相高效液相色譜(RP—HPLC)對(duì)粗提物進(jìn)行初步檢測(cè)。 結(jié)果：研究表明,黑水虻幼蟲產(chǎn)生抗菌肽的最佳誘導(dǎo)齡期為5齡；不同誘導(dǎo)方法產(chǎn)生抗菌肽對(duì)大腸桿菌抑菌活性不同,大腸桿菌菌液針刺浸泡60s,誘導(dǎo)后飼養(yǎng)24h提取的抗菌肽抑菌活性最好；其次為超聲波100W誘導(dǎo)20min產(chǎn)生抗菌肽抑菌活性；其余誘導(dǎo)方法產(chǎn)生抗菌肽也有一定的抑菌效果但與未誘導(dǎo)組相比差異不顯著。生物活性研究表明,黑水虻抗菌肽經(jīng)25℃、37℃、50℃處理10min后仍具有較為理想的抑菌效果；反復(fù)凍融次數(shù)少于4次時(shí)對(duì)抗菌肽抑菌活性無(wú)顯著影響,凍融6次以上抑菌活性減小較為明顯；pH值在5.0-9.0范圍仍可保持抑菌活性,pH5或pH9的環(huán)境中無(wú)明顯抑菌活性。利用反相高效液相色譜(RP—HPLC)得到抗菌肽獨(dú)立峰。 結(jié)論：1.誘導(dǎo)黑水虻幼蟲產(chǎn)生抗菌肽的最佳齡期為5齡。2.不同誘導(dǎo)方法產(chǎn)生抗菌肽對(duì)大腸桿菌抑菌活性不同。大腸桿菌菌液針刺浸泡60s,誘導(dǎo)后飼養(yǎng)24h提取的抗菌肽抑菌活性最好；其次抑菌活性較高的為100W超聲波誘導(dǎo)20min產(chǎn)生抗菌肽。3.黑水虻抗菌肽粗提物在25℃、37℃、50℃時(shí)均有較好的活性,低溫反復(fù)凍融次數(shù)少于4次時(shí)可保持較好抑菌活性；pH值的適宜范圍為5.0-9.0。4.利用反相高效液相色譜(RP—HPLC)得到抗菌肽粗提物獨(dú)立峰。
[Abstract]:Objective: to induce the antimicrobial peptides of Hermetia illucens L. by different methods, and to study the activity of the extracted crude extracts. To study the antimicrobial activity and biological activity of the crude extract of antimicrobial peptides from the larvae of Tabanus nigra. It is expected that this study will enrich the species of insect antimicrobial peptides and provide an effective experimental method for the development and utilization of antimicrobial peptides. Methods: in this study, the best inducing age was determined by the extraction of antimicrobial peptides from the larvae by the method of lapping extraction, the best inducing age was determined by the method of needle immersion induction of Escherichia coli bacteria, and the optimum induction age was determined by the method of grinding extraction. Staphylococcus aureus liquid injection induction method, Escherichia coli, Staphylococcus aureus liquid acupuncture immersion induction method, different power, The antimicrobial peptides of the 5th instar larvae of the black water fly were induced by ultrasonic induction at different times, and the crude antimicrobial peptides were extracted by the grinding extraction method. The antimicrobial activity of crude extract was studied respectively. The crude antimicrobial peptide extract with the best antibacterial effect was selected for biological activity identification and RP-HPLC was preliminarily detected by RP-HPLC. Results: the results showed that the best inducing age of antimicrobial peptide production was 5 years old, and the antimicrobial activity of different induction methods was different to Escherichia coli. The bacteriostatic activity of antibacterial peptides extracted from Escherichia coli was the best after incubation for 60 s, and the second was the antibacterial activity of antibacterial peptides induced by ultrasonic wave 100W for 20 minutes. The antimicrobial peptides produced by the other methods also had a certain bacteriostatic effect, but there was no significant difference compared with the uninduced group. The bioactivity study showed that the antimicrobial peptides of the black water gadfly still had ideal bacteriostatic effect after being treated at 25 鈩，

本文編號(hào)：1494491