液質(zhì)聯(lián)用中質(zhì)譜條件的優(yōu)化策略(參考版)
2025-02-27 09:03本頁(yè)面
【正文】 ? 組合所有的分離要素，建立分析方法，實(shí)施有關(guān)質(zhì)量控制技術(shù)規(guī)范要求。? 研究目標(biāo)化合物的色譜保留行為，原則上采用 50mm柱，保留時(shí)間在 13分鐘較為理想。? 對(duì)所獲得的 scan、 daughters圖可能的譜圖結(jié)構(gòu)表現(xiàn)、碎片離子的裂解規(guī)律進(jìn)行解釋。? 確定目標(biāo)化合物的檢測(cè)濃度要求，確定液質(zhì)聯(lián)用的可行性。? 在樣品基質(zhì)對(duì)于電離有明顯抑制效應(yīng)時(shí)，考慮采用空白樣品提取溶液作為標(biāo)準(zhǔn)系列的基質(zhì)溶劑。? 對(duì)于以高靈敏度定量為主的分析，樣品要求統(tǒng)一于一般液相色譜的殘留分析要求。定性優(yōu)先考慮色譜分離的好壞，兼顧考慮質(zhì)譜要求；定量?jī)?yōu)先考慮質(zhì)譜結(jié)果的可靠性，而無(wú)需考慮色譜行為的科學(xué)性。? 研究 scan、 daughter譜圖，尋找特征具有結(jié)構(gòu)特征的碎片離子，一般而言碎片越穩(wěn)定、豐度越大而得到的靈敏度越高。? 檢測(cè)器的倍增電壓，有選擇性的調(diào)節(jié)。? 合適的碰撞能量和碰撞室的聚焦電壓。A max.)? Sheath Gas Flow Rate (arb): 3580? Auxiliary Gas: 0? Capillary Temp (176。C): 400550 (600 max)? Discharge Current (181。 change column chemistry before changing solvent mix or position? column or lower, flow rates of 200400 uL/min? Peak widths for quantification not greater than 810 secs? Dissolve sample in start mobile phase solvent (weakest solvent possible)02383Solvents Compatible With ESIMethanolAcetonitrilePropanolIsopropanolButanol2methoxy ethanolAceticFormicTFAAmm. AcetateAmm. FormateHFBATEAAmm. HydroxideTetrabutyl amm. hydroxideHexafluorobutanolSamples can be dissolved in any HPLC patible solventModifiers Between 1% and 5 50 mM02382Electrospray Summary? Analyte type:– preformed ions (acids and bases)– polar neutrals– multiply charged ions of biopolymers– 100 da up to 200 000 da? Typical flow rates: low nL ml/min? Promote ionization:– correct pH– favorable HPLC solvent position– Postcolumn addition of reagents? Soft ionization technique? Typical applications : Drugs, Sugars, Peptides, Proteins, Oligonucleotides01328Steps for APCI Optimization? If analyte’s pKa is unknown, evaluate 3 pH regions in positive and negative ion modes.? Acids – Negative Ion detection, adjust pH 2 units below pKa– Decrease pH use formic acid , acetic acid, TFA? Bases – Positive Ion detection, adjust pH 2 units above pKa– Increase pH with NH4OH, TEA, TMA? Adjust corona discharge voltage? Adjust nebulization temperature– Consider possible thermal deposition of analyte01565APCI – Typical Operating Conditions? Flow Rates: 50181。216。C 2023006000001000000AbundanceWithout TFAFix 2023006000001000000minAbundanceWith TFAFix1:2 postcolumn addition of 75% propionic acid in IPATryptic Digest Map by ESLC/MS1 nmol Chicken LysozymeSignal Suppression due to Additives? Ion pairing with analyte, surface tension effect? Solut
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