Agilent LCMS products and the application strategies in veterinary drug analysis

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1 Agilent LCMS products and the application strategies in veterinary drug analysis Experience A New Level of Confidence Ma Xin 马 鑫 LCMS Application Engineer Internal Sales Training - The new 6495 QQQ LC/MS 9/1/2014 1

2 Two Great Companies from One 两 个 全 新 公 司 的 诞 生 Agilent Technologies and Keysight Technologies Separation expected in early November 2014, subject to local legal procedures. Agilent has evolved into two distinct investment and business opportunities. As two separate companies, each will be able to direct resources to its own business and customer needs. Each company has the operational and financial scale to stand on its own. The companies can concentrate solely on distinct markets and growth drivers. Both companies are well positioned to maximize growth. BCEAI 2013 Agilent Restricted October

3 Agilent China at a Glance (since 1985) 安 捷 伦 中 国 概 略 : 在 中 国 从 1985 年 至 今 21 Operations in14 cities 1800 employees 500 staff in R&D team 6 Legal Entities - Agilent Technology Holding Co. - Agilent Technology (China) Co. Ltd. - Agilent Technologies (Shanghai) Co. Ltd. - Agilent Technology Software Co. - Agilent Technology Chengdu Co. Ltd. - Agilent Technology Trading Co. Ltd. Chengdu Beijing Host one of the largest R&D team, as well as one of the major labs in Agilent Shanghai Full functions covered Full functions covered Agilent Profile September 1,

4 Agilent in Chemical Analysis 在 化 学 分 析 主 要 应 用 领 域 No. 1 in our core markets Food 食 品 Environment 环 境 Chemical & energy 化 学 化 工 能 源 Forensic & drug testing 法 医 毒 物 CORE TECHNOLOGY PLATFORMS Gas chromatography Mass spectrometry Spectroscopy Chemistries November Agilent 28, Profile

5 Continually Expanding Portfolio 持 续 拓 展 的 产 品 技 术 平 台 A few of our recent additions this week 7900 & 4200 Q-TOF LC/MS ICP-MS QQQ WINNER: 2013 R&D 100 Product of the Year and Product of the Show at ASMS GC/MS QQQ WINNER: Industrial Business Outlook 2012 Gold Award WINNER: 2012 R&D 100 Product of the Year GC/MSD MP-AES FTIR Spectrometer BCEAI 2013 Agilent Restricted October

6 Customers of Third-party Testing Laborotory 第 三 方 检 测 客 户 群

7 Analysis Strategy of Food Safety 食 品 安 全 分 析 策 略 Thermally stable & Nonpolar 热 稳 定 & 非 极 性 组 分 Volatility Thermally stable Volatility Thermally stable 7200 GC-QTOF 7000C GC-QQQ Unknown Analysis 未 知 物 筛 查 非 靶 标 化 合 物 分 析 Unknown Screening what s in the sample? 靶 标 化 合 物 分 析 Target Screening are specific compounds present & how much? Target Analysis 已 知 物 分 析 Nonvolatile Thermal unstable Non-Selective Selective Nonvolatile Thermal unstable 6500 LC-QTOF PCDL MSC All ion MSMS Thermal unstable & Polarity 热 不 稳 定 & 极 性 组 分 6400 LC-QQQ Dynamic MRM tmrm Introduction of the new 6495 QQQ LC/MS 9/1/2014 7

8 Proven ifunnel Technology Agilent Jet Stream Hexabore Capillary Dual Ion Funnel Thermal gradient focusing Efficient desolvation Creates an ion rich zone Six capillary inlets Samples x10 times more ion rich gas Removes the gas but captures the ions Removes neutral noise Agilent 6495 QQQ LC/MS System 9/1/2014 8

9 6495 QQQ Technologies Continued Development 2 Proven ifunnel Technology Agilent Jet Stream Hexabore Capillary Dual Ion Funnel Increased ion generation Enhanced ion sampling New Enhanced Q1 Ion Optics Improved ion transmission 2 3 New Tapered Hexapole Collision Cell Effective ion collection and transmission New Detector with High Energy Conversion Dynode Improved NEG ion detection with low noise Quantitative Applications Enhanced peak area response Improved peak area %RSD More sensitive and precise Lower Limits of Detection (IDL)and Quantitation (LLOQ) More reliable and robust Agilent 6495 QQQ LC/MS System 9/1/2014 9

10 6495 QQQ - Premium Performance Improved sensitivity and lower LLOQs Average 3x in S/N specifications and applications Improved precision and excellent accuracy at the lowest levels 3x in IDL specifications Proven 6 orders of linear dynamic range Proven robustness in complex matrix Food matrix and biological matrix (plasma) Improved mass range, fast scan speed and MRM acquisition rate Agilent 6495 QQQ LC/MS System 9/1/

11 Improved Sensitivity and Precision 6495 QQQ IDL IDL for the Agilent 6495 QQQ LC/MS System 6495 QQQ IDL Amount measured Replicates Area %RSD t (99%) IDL Reserpine (+) 1 fg 5 fg n = fg 1.02 fg Chloramphenicol (-) 1 fg 5 fg n = fg 1.25 fg 1 fg of reserpine used to measure IDL (+) 1 fg of chloramphenicol used to measure IDL (-) 1 x IDL = t x (%RSD / 100) x Amount measured = x (7.2 / 100) x 1 fg = 0.20 fg x IDL = t x (%RSD / 100) x Amount measured = x (9.7 / 100) x 1 fg = 0.27 fg Acquisition Time (min) Acquisition Time (min) Improved precision (%RSD) and Instrument Detection Limit (IDL) achieved on the 6495 vs. the 6490 Agilent 6495 QQQ LC/MS System 9/1/

12 Wide Linear Dynamic Range, Pesticide Fluazinam Responses Fluazinam, 1 fg pg (0.5 pg/ml 100 ng/ml) 5.5 Orders of Linear Dynamic Range R 2 = Precision Area %RSD < 20 %Accuracy Zoom-in ppt Level (ng/ml) ,000 5,000 10,000 50, ,000 %RSD (n = 7) %Accuracy Wide (5.5 orders) linear dynamic range, excellent assay accuracy and precision at all levels Internal Sales Training - The new 6495 QQQ LC/MS 9/1/

13 Proven System Robustness in Complex Food Matrix: Multi-Residue Pesticide Analysis in Black Tea Overlaid six data files / MRM chromatograms QC injections Day 1 QC injections Day 2 QC injections Day 3 Pesticide standards analyzed before, in the middle, and after matrix injections No signal decrease was observed after 3 days of operation running with heavy food matrix (black tea) %RSD < 6 (n = 6 data files) Average results of > 250 pesticides analyzed Internal Sales Training - The new 6495 QQQ LC/MS 9/1/

14 QQQ---Method development workflow of target compounds 目 标 化 合 物 监 测 方 法 开 发 流 程 Auto-optimizer MS parameters Optimizer Chromatographic Acquisition by dmrm/tmrm Generate reports Optimizer Database Search Report By MRM 1. Optimizer 自 动 优 化 软 件 自 动 优 化 化 合 物 子 离 子, Fragmentor 和 碰 撞 能 量 2. 优 化 色 谱 分 离 条 件 或 利 用 iset 技 术 转 化 原 有 色 谱 条 件 3. 对 于 多 农 残 或 多 兽 残 分 析 可 以 升 级 为 动 态 或 触 发 式 采 集 模 式, 兼 顾 灵 敏 度 及 采 集 速 率 4. 流 程 式 定 量 报 告 和 快 速 预 览 界 面, 灵 活 可 编 辑 报 告 格 式

15 Sensitive Quantitation of Estrogens in Cow Milk Eight estrogens spiked in cow milk at 0.1 µg/kg µg/kg estrogens spiked in cow milk --- Milk blank 1 Estriol ,8 ID Compounds LLOQ (pg/ml) %RSD n = 7 IDL (pg/ml) 1 Estriol α-Estradiol β-Estradiol Ethynyl estradiol Estrone Diethylstilbestrol Hexestrol Dienestrol Estrogen levels in milk products are regulated by government agencies down to 1 µg/kg Highly sensitive and precise quantitation of estrogens was achieved using the new 6495 with a simple QuEChERS procedure without derivatization or enrichment. LLOQs and IDLs are at low pg/ml level (low-fg on-column), far below the regulatory requirement Introduction of the new 6495 QQQ LC/MS 9/1/

16 LLOQs and IDLs at pg/ml (fg) level Inj 1 Estriol LLOQ level of 5 pg/ml, 25 fg on-column Inj 5 Estriol µg/kg R 2 = Inj 2 Inj 6 Inj 3 Inj 4 Inj 7 Levels (µg/kg) %RSD (n = 7) % Accuracy Replicates %RSD Estriol IDL n = 7 injections pg/ml, 8.9 fg Excellent precision and accuracy are observed at the lowest levels (LLOQs) Excellent linearity in quantitation range Introduction of the new 6495 QQQ LC/MS 9/1/

17 Multi- Veterinary Drugs Analysis by 6490QQQ 54 Antibiotics,15min 100ppt 磺 胺 间 甲 氧 嘧 啶 磺 胺 氯 哒 嗪 恩 诺 沙 星 奥 比 沙 星 地 塞 米 松 群 勃 龙 醋 酸 酯 磺 胺 嘧 啶 磺 胺 异 恶 唑 诺 氟 沙 星 培 氟 沙 星 氢 化 可 的 松 喹 乙 醇 磺 胺 甲 氧 嘧 啶 磺 胺 二 甲 嘧 啶 沙 拉 沙 星 萘 啶 酸 泼 尼 松 龙 曲 吡 那 敏 磺 胺 甲 噻 二 唑 苯 甲 酰 磺 胺 环 丙 沙 星 氟 甲 喹 敌 菌 净 氟 尼 辛 磺 胺 噻 唑 磺 胺 甲 氧 哒 嗪 麻 保 沙 星 恶 喹 酸 乙 胺 嘧 啶 吡 喹 酮 磺 胺 二 甲 氧 嘧 啶 磺 胺 喹 恶 啉 二 氟 沙 星 仲 丁 威 氯 丙 嗪 氨 基 比 林 磺 胺 醋 酰 磺 胺 吡 啶 氧 氟 沙 星 泰 勒 菌 素 奥 美 普 林 安 替 比 林 磺 胺 甲 恶 唑 甲 氧 苄 啶 洛 美 沙 星 林 可 霉 素 克 球 酚 左 旋 咪 唑 磺 胺 甲 基 嘧 啶 磺 胺 邻 二 甲 氧 嘧 啶 丹 诺 沙 星 替 米 考 星 乙 氧 酰 胺 苯 甲 酯 氯 舒 龙

18 β- Agonists Analysis by 6490QQQ 18 β- Agonists,10min 10ppt Clenbuterol 10ppt S/N=22 Salbutamol 10ppt S/N=21 Ractopamine 10ppt S/N=40

19 Nitrofurnans Analysis by 6490QQQ 4 derivatives of nitrofurans metabolites,6min 50ppt AMOZ SEM AHD AOZ SEM 50ppt S/N=10 AHD 50ppt S/N=19

20 Triggered MRM (tmrm)- 触 发 多 反 应 通 道 监 测 Triggered MRM, tmrm Database and Library Trigger Threshold Apex tmrm Product Ion Spectrum Match Score 95.1 tmrm database accelerates acquisition method setup tmrm library Match Score for confirmation tmrm is Fast and Sensitive UHPLC compatible Simultaneous Quantitation, Screening and Confirmation Enhanced Optimizer Workflow in MassHunter B.07 (to Better Support tmrm) MassHunter Optimizer B.07 Find 10 MRM transitions in Optimizer Injection volume flexibility Improved method editing Agilent 6495 QQQ LC/MS System 9/1/

21 Quantitation of antiviral drugs in chickens by UHPLC-QQQ with triggered MRM for unprecedented confirmation Sample Preparations

22 Improve the ability of the confirmation false positive results Sample disruptors Standard Spike sample Spike High matching score Spectrum False Positives Low matching score

23 Targeted Screening & Confirmation with QQQ Pesticides Method Test Mix: 254 compounds DB: 700+ compounds Library: 200+ compounds Veterinary Drugs tmrm Databas e & Library LC/MS Applicatio n Kits Standards Test Mix: 146 compounds DB: 500+ compounds LC Column On Site Training Library: 100+ compounds Forensic Toxicology Test Mix: 139 compounds DB: compounds Library: 100+ compounds Agilent unique data dependent acquisition for fast and sensitive compound screening, quantitation and confirmation. 安 捷 伦 科 技 2014 年 质 谱 新 产 品 发 布 及 研 讨 会 9/1/

24 Analysis Strategy of Food Safety 食 品 安 全 分 析 策 略 Thermally stable & Nonpolar 热 稳 定 & 非 极 性 组 分 Volatility Thermally stable Volatility Thermally stable 7200 GC-QTOF 7000C GC-QQQ Unknown Analysis 未 知 物 筛 查 非 靶 标 化 合 物 分 析 Unknown Screening what s in the sample? 靶 标 化 合 物 分 析 Target Screening are specific compounds present & how much? Target Analysis 已 知 物 分 析 Nonvolatile Thermal unstable Non-Selective Selective Nonvolatile Thermal unstable 6500 LC-QTOF PCDL MSC All ion MSMS Thermal unstable & Polarity 热 不 稳 定 & 极 性 组 分 6400 LC-QQQ Dynamic MRM tmrm Introduction of the new 6495 QQQ LC/MS 9/1/

25 6550 Instrument Technologies Comprehensive Performance Enhancements Mass Resolution >40, spectra /sec MS and 33 spectra/sec MS/MS 5 orders of linear dynamic range <1 ppm MS mass accuracy; <2 ppm MS/MS Unrivalled sensitivity Longer Invar flight tube (resolution + stable mass accuracy) Dual Agilent JetStream orthogonal spray (robust and stable mass calibration) New- Ion Beam Compression and Shaping (resolution + sensitivity) New embedded processor (50 spectra/sec) New- ifunnel (10X sensitivity gain) Hexapole axial focusing collision cell (faster MS/MS spectra) 4 GHz digitizer + ADC detector (resolution, mass accuracy, dynamic range) 25 Nov 2013

26 QTOF---- Non target compounds screening identification workflow 非 目 标 化 合 物 筛 查 鉴 定 流 程 Data Acquisition Extract Compound Identify Compound Generate reports MassHunter Acquisition Find cpd.by Formula: MSC Compound search by MS/MS --Source: Database --Match: Mass&RT MFG Molecular Formula Genaration Targeted No Yes Untargeted Match? Find compounds By Molecular Feature Agilent Only Database Search By Mass&RT&MS/MS Report 1. Auto MS/MS 自 动 二 级 质 谱 扫 描 模 式 同 时 完 成 一 级 二 级 质 谱 数 据 采 集 2. 依 据 农 残 数 据 库, 按 精 确 分 子 量 同 位 素 分 布 和 保 留 时 间 提 取 化 合 物 3. 二 级 谱 图 确 认, 如 谱 库 中 无 标 准 谱 图, 可 以 使 用 MSC 软 件 进 行 结 构 推 测 与 确 认 4. 生 成 报 告

27 Identify with Confidence: Personal Compound Database and Library (PCDL) Agilent is the first vendor to sell accurate mass MS/MS libraries - Higher confidence identifications - Run library searches from Qual or PCDL Manager - Display structures and difference spectra of hits in Qual - Customers can create their own libraries or append to a purchased library

28 Identify with Confidence: Molecular Structure Correlator (MSC) Use MS/MS spectra to go from formula to structures - Match substructures to specific fragments for positive identifications - Access formula and structures from Agilent PCD/PCDLs or the ChemSpider database (26 million compounds) to provide the most likely structures (only Agilent offers this capability!) C23H31NO2 MSC Proadifen

29 Veterinary drug screening project UHPLC-(Q)TOF screening Pig muscle extract spiked with 50 compounds Type A compounds: not authorized or forbidden; 20 ng/ml Type B compounds: listed in 37/2010/EC table 1; 200 ng/ml TIC

30 Veterinary Drugs Nemadectin - Sensitivity 0.5 µg/kg

31 Screening results review Structure EIC MS Sample Spectrum Mirror MS/MS Library

32 All Ions MS/MS Create MS/MS Spectra in TOF with High Fragmentor Voltage Great solution for screening in food, environmental and forensic/toxicology markets Unique to Agilent: MS/MS spectral libraries to identify fragments coelution score to confirm fragments belong to the precursor ion Re-interrogate data for unknowns without reacquiring Quickly add unknowns to the MS/MS library and include screening Rapidly set up a quantitative method for multiple compound screening 32 Nov 2013

33 Use of MSMS All Ions Browsing for High Confidence Data Review Quick browsing compound details view to quickly inspect All Ions MSMS results

34 Untargeted Screening with TOF or QTOF Pesticide Compounds Accurate Mass MS/MS Library LC Column Method LC/MS Application Kits On Site Training Standards Test Mix: >250 (new) Database: >1600 Library: >500 (new) Veterinary Drugs - new Test Mix: >200 Database: >1000 Library: >500 Forensic Toxicology Test Mix: >100 (new) Database: >9000 Library: >2500

35 Import and build Qual/Quant Method from Qual MH Quant MH Qual Compound results.cef file Reference pattern library Adduct pattern Autodynamic Quant method creation - Accurate mass EICs for quantifier and qualifier - Relative responses - Retention times ASMS Vancouver, May 2012

36 Introduction of the new 6495 QQQ LC/MS 9/1/

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