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Seeing is Believing: The Ultrafast LCMS-2020 is the Fastest MS Detector in the World! Analytical & Measuring Instruments Division, Shimadzu Corporation.

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Presentation on theme: "Seeing is Believing: The Ultrafast LCMS-2020 is the Fastest MS Detector in the World! Analytical & Measuring Instruments Division, Shimadzu Corporation."— Presentation transcript:

1 Seeing is Believing: The Ultrafast LCMS-2020 is the Fastest MS Detector in the World! Analytical & Measuring Instruments Division, Shimadzu Corporation Tel: Parto Zist Beboud

2 LAAN-E-LM003 2 Speed is Power UF scanning Ultrahigh scan speed of 15,000 µ/sec UF switching High-speed positive/negative polarity switching at 15 msec UF sensitivity High sensitivity even in high-speed analysis LCMS-2020 Seeing is Believing

3 LAAN-E-LM Overview of LC/MS

4 Electrospray ionization LAAN-E-LM003 4

5 Atmospheric pressure LAAN-E-LM003 5

6 6 Principle of a Mass Spectrometer (MS) Mass Spectrometer  A high voltage is applied to the column eluate, and it is atomized by nitrogen gas.  The charged droplets gradually become smaller, and ion evaporation occurs.  As the ions pass through the mass separator, they are separated according to mass.  In the detector, the quantity of ions is detected as a current value. Positive ion (protonated molecule) Proton added Proton removed Negative ion (deprotonated molecule) Proton (hydrogen ion) イオン化したい化合物の分子 Separated in a column Mixed sample injected Introduced into MS in order from component with lowest retention Mobile phase Column eluate Ion focusing unit Mass separator Detector Ion source Atmospheric pressure region Vacuum region

7 LAAN-E-LM003 7

8 8 LC-MS: System Configuration and Ionization Method This is an extremely soft ionization method, and is suited to the ionization of high- polarity compounds. This method is suited to the ionization of medium- and low-polarity compounds. Solvent delivery pump (gradient) Mobile phase Mixer Autosampler Controller LC detector Autosampler cleaning liquid Column oven Column Degasser Reservoir tray Rotary pump Nitrogen gas generator ESI: Electrospray Ionization APCI: Atmospheric Pressure Chemical Ionization Mass spectrometer Glass capillary Charged droplets are formed. Liquid sample Nitrogen gas High voltage (±3 to 5 kV) +: Positive ions are created. -: Negative ions are created. Ion evaporation Liquid sample Heater Nitrogen gas Sample molecule Corona needle Solvent molecule High voltage (  3 to 5 kV) +: Positive ions are created. -: Negative ions are created. Note: The LC detector may be removed if it is not required.

9 LAAN-E-LM003 9 Means of identifying sample  Retention time Column Injector Time SPD-20A/20AV SPD-M20A PDA detector  Retention time + Mass information LCMS-2020 The molecular weight of the sample can be ascertained with an MS spectrum. Provides greater selectivity than a UV spectrum nm mAU nm mAU UV spectra Information Obtained from LC and MS

10 LAAN-E-LM LC-MS: Analytical Data  MS m/z time (1) (2) (3) (4) (5) m/z (5) (4) time (3) m/z 193 (2) m/z 582 (1) TIC Mass Chromatogram Mass Spectrum  PDA (Photodiode Array) time (3) 210 nm (2) 580 nm LC Chromatogram nm time (2) (3) (4) (5) int. UV Spectrum nm (5) (4) AU

11 LAAN-E-LM If peak elution is late: If an impurity coincides with the target component: Mobile phase preparation errors Fluctuations in peak retention times Peak misidentification AB AB If an impurity coincides with the target component: Changes in area value Incorrect quantitation Risks Associated with LC Detectors

12 LAAN-E-LM The greatest merit in using an MS instrument as an LC detector: In addition to retention times, mass information for each peak can be obtained simply at the same time. m/z 264 m/z 278 m/z 267 m/z 281 The peaks (including those that cannot be separated by time) can be separated using mass information.  This reduces the risk of qualitative and quantitative errors. Merits of MS Detectors Mass information is a powerful tool for reducing the risks associated with LC analysis, such as the following: Peak identification (i.e., qualitative) errors Quantitative errors due to the elution of unpredicted impurities

13 LAAN-E-LM Features of the LCMS-2020

14 LAAN-E-LM With the introduction of the Prominence Ultrafast LC (UFLC), which offers the ultimate level of speed, superior reproducibility, and comprehensive expandability, ultrafast LC analysis is no utilized in a variety of new fields min. HPLC min.2.0 UFLC Shimadzu's Ultrafast LC Lineup Prominence UFLC: Greatly reduces analysis time without sacrificing separation. Prominence UFLCXR:Achieves greater separation while maintaining ultrahigh speed. From HPLC to UFLC, Then to UFLC-MS

15 LAAN-E-LM A mass spectrometer responding to the performance of UFLC: LCMS-2020 What makes an MS instrument suitable for UFLC? The ability to acquire data at high speed without sacrificing data quality is required! The three things that enable ultrafast analysis: The ability to perform scan measurement at high speed UFscanning The ability to switch between positive and negative ion measurement at high speed UFswitching High sensitivity in high-speed measurement UFsensitivity UFLC-MS

16 LAAN-E-LM points10 points 4 to 5 points Influence of Data Sampling Points on Peak Form 1.0 min. UFLC Data If the number of data points decreases, the sensitivity also decreases. This adversely affects the reproducibility. Influence of Data Sampling Points

17 Increase the scan speed. With conventional instruments, the sensitivity decreases. Decrease the scan speed. It is difficult to handle high- speed analysis. LAAN-E-LM LC-MS: Scan Speed Scan: Data is acquired in the desired m/z range. m/z 100 1,100 Scan speed (scan cycle) t(s) Scan speed (scan cycle) m/z 100 1,100 t(s) Quadrupole rod With the LCMS-2020, it has become possible to maintain sensitivity when the scan speed is increased.

18 LAAN-E-LM UFscanning   With measurement at 15,000 u/sec, the sharp peaks of UFLC are captured reliably. Samples: Seeing is Believing.

19 LAAN-E-LM  With measurement at 15,000 u/sec, the sharp peaks of UFLC are captured reliably.  Ultrafast analysis of 16 drugs UFscanning  1: Famotidine 2: Cimetidine 3: Atenolol 4: Lidocaine 5: Atropine 6: Metoprolol 7: Yohimbine 8: Noscapine 9: Bupivacaine 10: Alprenolol 11: Tetracaine 12: Diphenhydramine 13: Erythromycin 14: Dibucaine 15: Isopropylantipyrine 16: Warfarin Seeing is Believing.

20 LAAN-E-LM LC-MS: Positive/Negative Ion Measurement  Simultaneous Measurement of Positive and Negative Ions  The ease with which positive/negative ions are created depends greatly on the compound characteristics.  With positive/negative polarity switching, both positive and negative ions are measured at the same time.  In simultaneous measurement, the number of sampling points is important. Peak that readily becomes negative ions Peak that readily becomes positive ions Ionization in negative ion mode Ionization in positive ion mode Chromatograms for both positive and negative ions obtained in a single analysis. 20 points Number of Sampling Points 10 points 4 to 5 points

21 LAAN-E-LM UFswitching   With 15-msec switching during measurement, the sharp peaks of UFLC are captured reliably. Samples: Polarity switching time: Positive ion measurement SIM2CH Negative ion measurement SIM 2CH Positive ion measurement SIM 2CH

22 LAAN-E-LM UFscanning  + UFswitching   With measurement at 15,000 u/sec and switching at 15 msec, the sharp peaks of UFLC are captured reliably. Polarity switching time: Positive ion measurement 15,000 u/sec Negative ion measurement 15,000 u/sec Positive ion measurement 15,000 u/sec Mass Spectra of Dymuron Mass Spectra of Carpropamid Mass Spectra of Bentazone

23 LAAN-E-LM  Compounds That Give Both Positive and Negative Ions: Catechins Negative ions Positive ions m/z 307 m/z 291 m/z 459 m/z 443 m/z 473 m/z 305 m/z 289 m/z 457 m/z 441 m/z 471 Example of the Analysis of Catechins in Tea (Filtered Sample, 2 µL) 1: (-)-gallocatechin, 2: (-)-epigallocatechin, 3: (+)-catechin, 4: (-)-epicatechin, 5: (-)-epigallocatechin gallate, 6: (-)-gallocatechin gallate, 7: (-)-epicatechin gallate, 8: (-)-catechin gallate 9, 10: methylated catechins (-)-epicatechin High-Speed Positive/Negative Switching Analysis of Catechins Seeing is Believing.

24 LAAN-E-LM LCMS-2020 Measurement time + voltage switching time in each mode: 150 msec Measurement time + voltage switching time in each mode: 700 msec Glycyrrhizin Positive Negative Positive Negative High-Speed Positive/Negative Switching Analysis of Glycyrrhizin  With measurement at 15,000 u/sec and switching at 15 msec, the sharp peaks of UFLC are captured reliably. Seeing is Believing.

25 LAAN-E-LM Configuration of the LCMS-2020 Ionization probe Bottle for standard sample Dual inlet turbomolecular pump Mass spectrometric detector Ion optical system 350 mm 726 mm 553 mm

26 LAAN-E-LM  High Sensitivity Even in High-Speed Analysis Qarray Skimmer Octopole Entrance lens To quadrupole rod From desolvation line UFsensitivity 

27 LAAN-E-LM  UFsensitivity High sensitivity even in high-speed analysis Reserpine, 1 pg: S/N > 150 (RMS) SIM chromatogram of reserpine (1 pg) (x1,000) (1.00) Seeing is Believing. UFsensitivity 

28 LAAN-E-LM LC-MS: Ion Source CID Normal CID Normal CID Normal CID  Ultrafast Analysis of Erythromycin Impurities Set quite a high lens voltage, and produce fragment ions.

29 LAAN-E-LM A sample was added to blood plasma, acetonitrile was added, centrifugal deproteinization was performed, and the sample was continuously injected in quantities of 1  L. A reproducibility of 2.26% was attained over 10 days, indicating superior durability as well as reproducibility. Ion Source After Continuous Analysis Plasma Sample Injection Number Peak Area of Nortriptyline %RSD: 2.26 Internal standard Analysis time: 6 min 2,500 injections, 10 days Long-Term Stability Features Supporting the Three UFs

30 LAAN-E-LM Easy Maintenance Simple capillary replacement Large source window Sample bottle for auto-tuning The desolvation line can be replaced without stopping the vacuum. Features Supporting the Three UFs The ionization probe can be attached and detached with a single operation.

31 LAAN-E-LM A Variety of Ionization Options Molecular weight 10,000 1, No polarity Medium polarity High polarity ESI DUIS APCI Features Supporting the Three UFs ESI probe Introduction of sample Corona needle

32 LAAN-E-LM A Variety of Ionization Options Features Supporting the Three UFs MS Chromatograms MS Spectra Obtained with DUIS Measurement Thiamine Riboflavin Calciferol 3 Types of Water-Soluble/Lipid-Soluble Vitamins (Mixed Sample) 1. Thiamine:m/z 265:Cations, water-soluble vitamin created by dissociation 2. Riboflavin:m/z 377:Protonated molecules, water-soluble vitamin 3. Calciferol:m/z 397:Protonated molecules, lipid-soluble vitamin

33 LAAN-E-LM LCMSsolution ver.5 The popular aspects of LCsolution and LCMSsolution Ver. 3 have been retained, and an easier-to-use interface has been added. LCMSsolution Ver. 5

34 LAAN-E-LM Comparison of PDA and MS chromatograms Comparison of chromatograms in different data files Switching between parallel display and overlap display New Functions of Data Browser

35 LAAN-E-LM Searches for the optimum lens voltage for each compound Sets optimum values in method files Automatic Optimization of MS Parameters

36 LAAN-E-LM Electrospray VS MALDY

37 LAAN-E-LM003 37

38 LAAN-E-LM003 38

39 LAAN-E-LM003 39

40 LAAN-E-LM003 40

41 LAAN-E-LM003 41

42 LAAN-E-LM003 42

43 LAAN-E-LM LCMS-2020: Influence of Mobile Phase Solvent

44 LAAN-E-LM Xanthine Derivatives 0.1% Aqueous Formic Acid/MeOH = 80/20 Shim-pack VP-ODS 2  50mm, 5µm 0.2mL/min Y = ( )X + ( e+006) R2 = Y = ( )X + (880292) R2 = Y = ( )X + (612820) R2 = Caffeine Theophylline Theobromine

45 45 LAAN-E-LM003 Xanthine Derivatives XR-ODS 2  75mm, 2.2 µm 0.4 mL/min  Prioritizing maintenance of the separated state Theobromine in Green Tea Elution Time 9 min  2.2 min Reduced by 6.8 min Solvent Volume 1.2 mL  0.88 mL 51% OFF 0.1% Aqueous Formic Acid/MeOH = 80/20 Area Concentration Theobromine

46 LAAN-E-LM Xanthine Derivatives XR-ODS 2  50 mm, 2.2 µm 0.4m L/min  Specifying the column size Theobromine in Green Tea Elution Time 9 min  1.6 min Reduced by 7.4 min Solvent Volume 1.8 mL  0.64 mL 64% OFF 0.1% Aqueous Formic Acid/MeOH = 80/20 Theobromine Area Concentration

47 LAAN-E-LM Catechins: ESI(-) 0.5% Aqueous Formic Acid/ACN 2% (0 min) - 25% (3 min) - 2% ( min) Gradient XR-ODS 2  50 mm, 2.2 µm 0.5 mL/min 0.5% Aqueous Formic Acid/ACN/THF MAX 19.2 MPa 1: Gallic acid 2: (-)-gallocatechin, 3: (-)-epigallocatechin, 4: (+)-catechin, 5: (-)-epicatechin, 6: (-)-epigallocatechin gallate, 7: (-)-gallocatechin gallate, 8: (-)-epicatechin gallate, 9: (-)-catechin gallate *Adding THF improves epimer separation. *Peaks 5 and 6 are also separated well. 2% (0 min) - 25% (3 min) - 2% ( min) Gradient

48 LAAN-E-LM Catechins: ESI(-) 0.5% Aqueous Formic Acid/MeOH MAX 27.5 MPa % Aqueous Formic Acid/ACN Same analysis time Mobile phase B Acetonitrile  Methanol *Epimer separation is better. *Sensitivity is also good. *There is insufficient separation between peaks 3 and 4 although this is not a problem with LCMS. MAX 19.2 MPa XR-ODS 2  50 mm, 2.2 µm 0.5 mL/min 2% (0 min) - 25% (3 min) - 2% ( min) Gradient 3% (0 min) - 50% (3 min) - 3% ( min) Gradient

49 LAAN-E-LM Carbamate Pesticides [M+H] + : m/z 207[M+NH 4 ] + : m/z 240[M+H] + : m/z 237[M+H] + : m/z 163[M+H] + : m/z 242 [M+H-H 2 O] + : m/z 220 [M+H] + : m/z 258[M+NH 4 ] + : m/z 208[M+H] + : m/z 224[M+H] + : m/z 222 [M+H] + : m/z 202[M+H] + : m/z 355[M+H] + : m/z 226[M+H] + : m/z 180 (1) Aldicarb Sulfoxide(2) Aldicarb Sulfone(3) Oxamyl(4) Methomyl(5) Methiocarb Sulfoxide (6) 3-OH Carbofuran(7) Methiocarb Sulfone(8) Aldicarb(9) Bendiocarb(10) Carbofuran (11) Carbaryl(12) Thiodicarb(13) Ethiofencarb(14) XMC [M+H] + : m/z 239 [M+H] + : m/z 194 [M+H] + : m/z 208[M+H] + : m/z 226[M+H] + : m/z 411 (15) Pirimicarb (16) Isoprocarb(17) Trimethacarb(18) Fenobcarb(19) Methiocarb(20) Benfuracarb

50 LAAN-E-LM Carbamate Pesticides 10-mmol/L Aqueous Ammonium Acetate/ACN 5% (0 min) - 95% (20-25 min) - 5% ( min) Gradient 10-mmol/L Aqueous Ammonium Acetate/MeOH VP-ODS 2  150 mm, 5 µm, 0.2 mL/min *It is difficult to separate peaks 16 and 17 with acetonitrile, so use of methanol is recommended for this analysis. *Sensitivity is also better with methanol. 5% (0 min) - 95% (20-25 min) - 5% ( min) Gradient

51 LAAN-E-LM Carbamate Pesticides 10-mmol/L Aqueous Ammonium Acetate/MeOH 5% (0 min) - 95% (3-4 min) - 5% ( min) Gradient XR-ODS 2  50 mm, 2.2 µm, 0.4 mL/min MAX 25.5 MPa Analysis Time 40 min  8 min Reduced by 32 min Solvent (Methanol) Volume 3.05 mL  1.06 mL 65% OFF

52 LAAN-E-LM Drugs [M+H] + : m/z 338[M+H] + : m/z 253[M+H] + : m/z 267[M+H] + : m/z 315 [M+H] + : m/z 237[M+H] + : m/z 235[M+H] + : m/z 355[M+H] + : m/z 250[M+H] + : m/z 280 [M+H] + : m/z 281[M+H] + : m/z 278[M-H] - : m/z 329[M+H] + : m/z 231[M+H] + : m/z 309 [M-H] - : m/z 307 (1) famotidine (2) cimetidine(3) atenolol(4) ranitidine (5) procaine(6) lidocaine(7) yohimbine(8) alprenolol(9) doxepin (10) imipramine(11) amitriptyline(12) furosemide(13) isopropylantipyrine(14) warfarin

53 LAAN-E-LM Drugs 0.1% Aqueous Formic Acid/ACN XR-ODS 2  50 mm, 2.2 µm 0.4 mL/min Analysis Time 6.5 min  4 min Reduced by 1.5 min Solvent Volume 1.06 mL  0.36 mL 66% OFF *Analysis can be performed in the same way even if the gradient conditions are changed; as a result, the analysis time can be reduced. 5% (0 min) - 80% (3-4 min) - 5% ( min) Gradient 5% (0 min) - 80% (1-1.5 min) - 5% ( min) Gradient

54 LAAN-E-LM Drugs 0.1% Aqueous Formic Acid/MeOH 0.1% Aqueous Formic Acid/ACN Same analysis time Mobile phase B Acetonitrile  Methanol *It is difficult to separate peaks 1 to 5. XR-ODS 2  50 mm, 2.2 µm 0.4 mL/min 5% (0 min) - 80% (3-4 min) - 5% ( min) Gradient 5% (0 min) - 85% (3-4 min) - 5% ( min) Gradient

55 LAAN-E-LM Drugs mmol/L Aqueous Ammonium Acetate/MeOH Same analysis time Mobile phase B Acetonitrile  Methanol Mobile phase A 0.1% aqueous formic acid  10-mmol/L aqueous ammonium acetate 0.1% Aqueous Formic Acid/ACN *Peaks 1 to 5 are separated well. *Sensitivity is at least as good. *The elution order changes but, with LC/MS, identification is simple. XR-ODS 2  50 mm, 2.2 µm 0.4 mL/min 5% (0 min) - 85% (3-4 min) - 5% ( min) Gradient 5% (0 min) - 80% (3-4 min) - 5% ( min) Gradient

56 LAAN-E-LM Drugs 10-mmol/L Aqueous Ammonium Acetate/ACN 10-mmol/L Aqueous Ammonium Acetate/MeOH *The elution order changes but, with LC/MS, identification is simple. XR-ODS 2  50 mm, 2.2 µm 0.4 mL/min 5% (0 min) - 85% (1-1.5 min) - 5% ( min) Gradient 5% (0 min) - 65% (1-1.5 min) - 5% ( min) Gradient

57 LAAN-E-LM A mass spectrometer responding to the performance of UFLC: LCMS-2020 With the LCMS-2020, the peaks output from the Prominence UFLC at high speed can be captured. High sensitivity can be attained even in high-speed measurement. It is easy to change from acetonitrile to methanol. - Sensitivity - Pressure - Changes in separation patterns Summary

58 LAAN-E-LM Technical Reports on Ways of Saving Solvent No. 24 Reduced Solvent Consumption and Operational Costs with Ultra High Speed Analysis No. 25 Reduced Solvent Consumption and Operational Costs with Existing LC Systems Related Technical Reports


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