本文關(guān)鍵詞：基于表面增強(qiáng)拉曼散射光譜的氨基酸和多肽檢測(cè)方法研究　出處：《吉林大學(xué)》2017年博士論文　論文類型：學(xué)位論文

  更多相關(guān)文章： SERS 偶氮偶聯(lián) 茚三酮 磷酸化 氨基酸 多肽

【摘要】：本論文針對(duì)目前缺乏快速靈敏的氨基酸和多肽檢測(cè)手段等問題,建立了幾種基于表面增強(qiáng)拉曼散射(SERS)技術(shù)的分析方法。將SERS技術(shù)與化學(xué)衍生法結(jié)合,利用衍生反應(yīng)物或生成物的SERS信號(hào)間接分析氨基酸和多肽;將SERS基底改進(jìn)成銀@二氧化鈦(Ag@Ti O2)納米材料,通過磷酸基團(tuán)對(duì)Ti O2的親和作用以及銀的拉曼增強(qiáng)效應(yīng)不僅對(duì)磷酸化氨基酸進(jìn)行了富集而且實(shí)現(xiàn)了SERS檢測(cè)。本論文的創(chuàng)新點(diǎn)主要包括:(1)將偶氮偶聯(lián)反應(yīng)和SERS技術(shù)結(jié)合,以對(duì)巰基苯胺(PATP)分子作為重氮組分,組氨酸(His)和酪氨酸(Tyr)作為偶聯(lián)組分進(jìn)行偶氮偶聯(lián)反應(yīng),生成最大吸收在500 nm左右的偶氮生色分子。以表面增強(qiáng)共振拉曼散射(SERRS)檢測(cè)代替吸光度檢測(cè),根據(jù)其SERRS信號(hào)進(jìn)行His和Tyr的定量分析、混合物鑒別和實(shí)際樣品分析。優(yōu)點(diǎn)是無需復(fù)雜的樣品前處理、實(shí)驗(yàn)過程快速簡便、選擇性好和靈敏度高。(2)利用上述基于SERRS技術(shù)和偶氮偶聯(lián)反應(yīng)的方法,以磺胺酸分子作為重氮組分,分別與His、Tyr和促甲狀腺激素釋放激素(TRH)偶聯(lián)生成具有SERRS活性的偶氮分子,進(jìn)行定量分析。運(yùn)用該方法實(shí)現(xiàn)了正常人和甲亢患者血清中TRH的檢測(cè),該方法具有簡單快速、超靈敏性、選擇性以及實(shí)用性和可靠性的優(yōu)勢(shì)。(3)基于經(jīng)典的氨基酸總量檢測(cè)法-茚三酮比色法,將過量茚三酮試劑與混合氨基酸樣品作用后的反應(yīng)液負(fù)載在銀納米粒子(Ag NPs)自組裝膜上進(jìn)行SERS測(cè)試,過量茚三酮的SERS強(qiáng)度變化反映氨基酸總量變化。該化學(xué)衍生方法成熟可靠,操作簡便,SERS技術(shù)的應(yīng)用大大提高了其檢測(cè)靈敏度。(4)基于半導(dǎo)體納米材料Ti O2對(duì)磷酸基團(tuán)的特異捕獲能力,制備了新型雙功能Ag@Ti O2核殼SERS活性基底,該納米材料既能富集磷酸化氨基酸或多肽又能夠增強(qiáng)待測(cè)分子的拉曼信號(hào)。該方法為實(shí)際樣品中磷酸化多肽的檢測(cè)及探究蛋白質(zhì)磷酸化作用提供了新的思路和手段。
[Abstract]:This paper aims at the lack of rapid and sensitive detection methods for amino acids and peptides. Several analytical methods based on surface-enhanced Raman scattering (SERS) technique were established. The SERS technique was combined with chemical derivation method. Amino acids and peptides were analyzed indirectly by SERS signal of derivative reactants or products. The SERS substrate was modified to form silver @ titanium dioxide (AgTiO _ 2) nanomaterials. The affinity of phosphate group to TIO 2 and Raman enhancement of silver not only enriched phosphorylated amino acids but also achieved SERS detection. The azo coupling reaction was combined with SERS technique. The azo coupling reaction was carried out with p-mercaptoaniline PATP as diazo component, histidine Hisand tyrosine tyrosine tyrosine Tyr as coupling component. Azo chromogenic molecules with maximum absorption of about 500 nm were formed. Surface enhanced Resonance Raman scattering (SERRS) detection was used instead of absorbance detection. According to its SERRS signal, the quantitative analysis of His and Tyr, the identification of mixture and the analysis of actual samples are carried out. The advantages are that there is no need for complex sample pretreatment, and the experimental process is fast and simple. Based on SERRS technique and azo coupling reaction, sulfamic acid was used as diazo component, and His was used as a diazo component, respectively. Tyr was coupled with thyrotropin releasing hormone (TSH) to form azo molecules with SERRS activity. Quantitative analysis. Using this method to achieve the normal and hyperthyroidism patients serum TRH detection, this method is simple, rapid, hypersensitivity. Selectivity and practicability and reliability are based on the classical total amino acid determination method-ninhydrin colorimetry. The reaction solution loaded with excess ninhydrin reagent and mixed amino acid sample was tested by SERS on silver nanoparticles Ag NPs self-assembled film. The SERS intensity change of excess ninhydrin reflects the change of total amino acid content. The chemical derivation method is mature and reliable and easy to operate. The application of SERS technology greatly improved its detection sensitivity. 4) based on the semiconductor nano-material TIO _ 2, the specific capture ability of phosphate group was greatly improved. A novel bifunctional Ag@Ti O2 core-shell SERS active substrate was prepared. The nanomaterials can not only enrich phosphorylated amino acids or peptides, but also enhance the Raman signal of the molecules to be tested. This method provides a new idea and hand for the detection of phosphorylated peptides in actual samples and the exploration of protein phosphorylation. Paragraph
【學(xué)位授予單位】：吉林大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2017
【分類號(hào)】：O657.37

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