Lassen SQ_Customer talk.ppt

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褒耗晏
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1 Liquid Chromatography/ Mass Spectrometry Fundamental Jimmy Chan The Advantage of LC-MS A general detector It can connect with HPLC to separate and get more structure informations It can use in quantitative and qualitative analysis 1

2 The Limit of LC-MS Sample must be ionization Ionic compound can motion transfer and analysis MS must vacuum 离子飞行的平均自由程需最大化理论上, 真空度越高越好 MS must analysis with M/Z The Solvent of API-MS Suitable for both ES and APCI Methanol Acetonitrile Water Ethanol Propanol Isopropanol Butanol DMF (1) DMSO (1) Acetone CH 2 Cl 2 CHCl 3 Suitable for only APCI Toluene Benzene (2) Hydrocarbons (e.g., Hexane) Styrene (2) CCl 4 CS 2 Cyclic Hydrocarbons (e.g., Cyclohexane) (1) At lower solvent percentages (~10 % or less) under ES conditions (2) These solvents not often used due to environmental health and safety issues 2

3 ph The Selection of API Addtive 乙酸, 甲酸,TFA, 氢氧化铵一般的缓冲液 / 离子配对试剂乙酸铵 / 甲酸铵三氟乙酸 (TFA) 七氟丁酸 (HFBA) 四乙基或四丁基氢氧化铵 (TBAH) 阳离子化试剂 µm 的乙酸钠或乙酸钾一般考虑挥发性 ( 污染喷雾室, 堵塞喷嘴 ) 导电性 ( 对离子气化过程减少小液滴的形成 ) 离子配对 ( 进入气相时中和预带电离子 ) 不能使用在 HPLC 中常使用的磷酸盐, 硫酸盐或硼酸盐! The Application of wide-range Small Molecular: Medical drug Analysis Drug abuse Analysis Antibacterial drug Analysis Steroid Analysis Structure comformation Proteomics: Peptide and Protein molecular Weight Peptide Sequence Peptide and Protein Purity Analysis Protein identification 3

4 The LC method transfer to LC/API-MS method 非挥发性的缓冲盐挥发性缓冲盐浓度 <10 mm ( 对于 ES ) 或 <100 mm ( 对于 APCI) 用醋酸铵或甲酸三氟乙酸 (TFA) 七氟丁酸(TFBA) 羟基四丁基胺取代磷酸盐硫酸盐和硼酸盐如果必须使用非挥发性盐,, 使用仅仅阳离子或阴离子部分不挥发的缓冲盐, 例如, 用醋酸钠而不用磷酸钠 Formic acid, acetic acid, TFA, ammonium hydroxide 挥发性离子对试剂可以使用 API-Electronspray Ionization 溶剂喷雾雾化气 ( 气体用红色显示 ) 电喷雾离子 V 加热的氮气干燥气双电子毛细管入口蒸发 Rayleigh Coulomb 极限爆裂蒸发被分析离子 4

API-Chemical Ionization 含有气溶胶的分析物溶剂在蒸发器中蒸发蒸汽通过电晕针放电形成带电荷的反应剂离子流动相分析物溶剂 --------->[ 溶剂 H] [ 溶剂 H] M---------> 溶剂 [M] 电荷转移至分析物分子分析物离子 The Characteristic Of API ESI APCI Compound with volitility

5 API-Chemical Ionization 含有气溶胶的分析物溶剂在蒸发器中蒸发蒸汽通过电晕针放电形成带电荷的反应剂离子流动相分析物溶剂 >[ 溶剂 H] [ 溶剂 H] M > 溶剂 [M] 电荷转移至分析物分子分析物离子 The Characteristic Of API ESI APCI Compound with volitility Compound with thermo unstabilite Ionization form in liquid Besides singal charge, it can form mutiple charges Compound with volitility Compound polarity lower than ESI Compound with thermo stabilite Ionization form in gas Only singal charge 5

6 The Relative Application of LC/MS What is LC/MS?? 6

7 Which mode do you get the singal charge?? Abundance [MH] MW = 308 基峰 : 谱图中最强的离子 m/z--> 苯基丁氮酮 (Phenylbutazone) 的 API-ES 谱图 Which mode do you get the singal charge?? Ginsenoside 7

8 Which mode do you get the mutiple-charge?? Abundance m/z--> Abundance m/z--> A22 A A A A A A A A A A A A Assume adduct is a proton: M1=(An)/n M2=(An1)/(n1) Solve for A 肌红蛋白 (Myoglobin) 的 API-ES 谱图 Mass Spectrum Concept Abundance 3D-Data(R.T m/z Intensity) m/z Time COOH OH COOH OCOCH 3 CF 3 CF 3 TIC 反应产物的联机分析结果 a 色谱图 ;b 总离子流图 ;d 质谱棒图 8

9 Raw, Filtered Full, and Condensed Data Abund Raw Data 原始数据 m/z Abund threshold m/z Filtered Full Data 过滤后的数据 Profile Abund m/z Condensed Data 精简数据 line spectra Mass Spectrum 9

10 LCDEMO02.D: EIC 285? ll MS, Smoothed (2.4,3, SG) LCDEMO02.D: BPC ? ll MS, Smoothed (2.4,3, SG) Time [min] Total Ion Chromatography (TIC) UV TIC Extract Ion Chromatography (EIC) x10 6 Intens. EIC 285 Increase S/N Subtract background Separate the co-elution x10 7 TIC What the purpose of EIC?? Quantitative!! 10

11 The Explain of API Mass Spectrum!! ESI and APCI can supply the compound messgae The ion depend on additive and condition Positive [MH] 酸性条件 [MNa], [MK] ( 有盐时 ) [MNH 4 ] 有铵盐缓冲溶液 [MX] X = 溶剂或缓冲溶液中的阳离子 [2MH] 在高浓度时形成的二聚体 [MHS] 溶剂添加剂 Negative [M-H] - 碱性条件 [MX] - X = 溶剂或缓冲溶液中的阴离子 [M-HS] - 溶剂添加剂 What is the Collision-Induced Dissonciation(CID)? 通过与中性分子碰撞将能量传递给离子的过程能量传递足以导致键的开裂和所选离子的重排 11

The CID in API Transition region 这是什么? 利用 API-MS 的传输区的电场在中压区 (~0.

12 The CID in API Transition region 这是什么? 利用 API-MS 的传输区的电场在中压区 (~ torr) 碰撞激活离子来产生 CID 质谱图硬件 : ION TRAP The Process of HED and EM EM 0 to e - SQ 负离子采样 EM 0 to Dynode 7 kv 正离子采样 Lens e - SQ Dynode 7 kv Lens 12

Introducing the Agilent 6130 Singal Quadrupole Mass Spectrometer New Agilent 6000 LC/MS Series A new and expanded portfolio for applications from life science to chemical analysis, combining

13 Introducing the Agilent 6130 Singal Quadrupole Mass Spectrometer New Agilent 6000 LC/MS Series A new and expanded portfolio for applications from life science to chemical analysis, combining exceptional performance and legendary Agilent reliability New QQQ Updated Ion Trap Family 6300 Series Ion Trap LC/MS 6410 Triple Quadrupole LC/MS New QTOF 6510 Quadrupole Time-of-Flight LC/MS Updated TOF 6210 Time-of-Flight LC/MS New Single Quads 6100 Series Single Quad LC/MS 13

14 Lassen LC/MS Singal Quadrupole system introduction Atmosphere Pressure Ionization -Mass Spectrometry (API-MS) API-MS is usde for analysis liquid samples that introduce by HPLC FIA and Infusion. Application in wide mass range. 1pg sensitiveity limit. 14

15 6100 Series SQ: and Entry Level (10 to 1500amu) p/n G6110AA 2. Mid-range (POS/NEG, 10 to 1500amu) p/n G6120AA 3. Research (2 to 3000amu) p/n G6130AA 4. Research (fast scan, 10 to 1350amu) p/ng6140aa 6120 SIM Scan ( 10pg; S/N>20:1 ) Mass range: 10~1500 m/z Pos/Neg Switching and Muti Signal Acquisition Function Block Diagram Inlets Ion Source Ion Optics Mass Filter Detecto r Vacuum System Electronics 15

16 Agilent LC-6320 Ion Trap/MS Liquid introduce Nubalizer gas Nubalizer Neutral Molecules Analyte Ions Clusters Salts Shield hole Focus HED/EM Len capillary Octopole Quadrupole Heat N 2 碎片碰撞电压 (CID) Mutimod Source ( ESIAPCI ) HPLC inlet Nebulizer ESI Zone APCI Zone Charging electrode Reversing electrode Thermal container APCI counterelectrode Corona needle IR emitters Drying gas Capillary Sensor for vapor temperature 16

17 The G1978A Multimode Source Nebulizer APCI Counterelectrode IR emitters APCI corona needle Sensor for vapor temperature Field-shaping electrodes Desolvation (Dry Gas with Heat) 冷却的毛细管加热的干燥气直接吹向喷雾的液滴 17

18 Desolvation and Capillary Skimmer Octopole and Lens 18

19 Octopole RF Ion Introduction Lenses 八极杆 RF 离子导引提高了所有质谱离子的传输率更高的灵敏度更强的分辨率减少离子能量的色散 Skimmer assembly Octopole RF Ion Guide Octopole 19

20 Agilent Singal Quadrupole Mass Spectrometry 20

21 Discussion and Question?? 21

材料与方法仪器 ACQUITY UPLC Xevo TQ-S 三重四级杆液质联用仪 ( 美国 Waters 公司 ); 电喷雾离子源 (ESI), Masslynx 4.1 数据处理系统 (Waters);Thermo 低温离心机 ;KQ

材料与方法仪器 ACQUITY UPLC Xevo TQ-S 三重四级杆液质联用仪 ( 美国 Waters 公司 ); 电喷雾离子源 (ESI), Masslynx 4.1 数据处理系统 (Waters);Thermo 低温离心机 ;KQ [ 应用纪要 ] Rapid identification of 5 illegal additives in slimming health food by Xevo TQ-S 郝刚苏州市食品药品检验所, 苏州 15104 关键词超高效液相色谱 - 质谱, 多反应监测, 减肥类保健食品非法添加, 快速定性定量