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Charles Lévesque, M.Sc. Product Manager-Analytical Chemistry Vincent Bédard, B.Sc. HPLC Specialist François Béland, Ph.D. Vice-President of R&D New Generic.

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Presentation on theme: "Charles Lévesque, M.Sc. Product Manager-Analytical Chemistry Vincent Bédard, B.Sc. HPLC Specialist François Béland, Ph.D. Vice-President of R&D New Generic."— Presentation transcript:

1 Charles Lévesque, M.Sc. Product Manager-Analytical Chemistry Vincent Bédard, B.Sc. HPLC Specialist François Béland, Ph.D. Vice-President of R&D New Generic Approach for HPLC analysis: The Modern Silica-Based HPLC Columns; SiliaChrom ® Series

2 Outlines HPLC Listing of all C18 SiliaChrom Strategy SiliaChromChemistry Review New Techologies-SB,AQ,XT Evaluation of SiliaChrom Recent SiliaChrom Applications Generic Approach for HPLC applications SiliaChrom Focus

3 Alltima C18 Alltima C18 LL Alltima HP C18 Alltima HP C18 EPS Alltima HP C18 Hi-Load Alltima HP C18 AQ Alltima HP C18 Amide Alltima WP C18 AlphaBond C18 Amethyst C18 AminoQuant APEX ODS Apollo C18 AQUA C18 Aquapore OD-300 C18 AQUASIL C18 Ascentis C18 Ascentis Express C18 ASI C18 ASI microC18 Atlantis dC18 Atlantis T3 Axxi-Chrom C18, ODS Bakerbond C18 BetaBasic 18 BetaMax Neutral BETASIL C18 Bio-C18 ACE 300 C18 ACE AQ ACE C18 ACE C18-AR AccQTag Ultra ACQUITY UPLC BEH130 ACQUITY UPLC BEH 300 ACQUITY UPLC BEH C18 ACQUITY UPLC BEH Shield RP18 ACQUITY UPLC HSS C18 ACQUITY UPLC HSS C18 SB ACQUITY UPLC HSS T3 ACQUITY UPLC OST C18 Adsorbosil C18 Adsorbosphere C18 Adsorbosphere HS C18 Adsorbosphere UHS C18 Adsorbosphere XL 300 C18 Adsorbosphere XL C18 Adsorbosphere XL C18-B Allsphere ODS-1 Allsphere ODS-2 Allure C18 Allure Aqueous C18 BioBasic 18 Bio-Sil ODS Bio-Sil ODS-10 Bondclone C18 BR-C18 Burdick & Jackson C18 Cadenza CD-C18 Cadenza 5CD-C18 Cadenza CL-C18 Cadenza 5CL-C18 Cadenza CW-C18 Cadenza 5CW-C18 Cadenza HS-C18 Capcell C18 AG 120 Capcell C18 SG 120, 300 Capcell C18 UG 80, 120, 300 Capcell C18 MG Capcell C18 MGII Capcell C18 MGIII Capcell C18 ACR Capcell C18 AQ Capcell C18 IF Chemcosorb 5-ODS-H Chromegabond C18 Chromegabond WR-C18 Chromolith RP-18e Clarity Oligo-RP Clipeus C18 Cogent HPS C18 Cogent hQ C18 Cogent Simulare C18 Cogent MicroBee C18 Cogent Bidentate C18 Cogent e-Column C18 Cosmosil C18 MS-II Cosmosil C18 AR-II Cosmosil C18 AR-300 Cosmosil C18 PAQ Delta-Pak C18 Denali Develosil ODS Diazem 2060 C18 60A Diazem 300A C Diazem C18 Diazem C18 Diazem C18 Diazem C18EC Diazem C18 Diazem C Diazem C Diazem C18 120A Diazem C18 120A series 43,44,45 and 46 Diazem C18 series 200A, 300A Diazem C18 series 2000 Diazem C18 series 3000 Diazem C18 series 4000 Diazem C18 60A HPLC C18 column listing (L1-USP)

4 HyPURITY AQUASTAR HyPURITY C18 IBM Octadecyl IB-SIL C18 IB-SIL C18 BD Inertsil ODS Inertsil ODS series 1,2,3 ISCO C-18 JSphere ODS-H80 JSphere ODS-L80 JSphere ODS-M80 Jupiter C18 Kinetex C18 Kromasil C18 LiChrospher PAH LiChrosorb RP-18 LiChrospher 100 RP-18 LiChrospher RP18e Luna C18 Luna C18(2) Diazem C18 60A series 1,2,3 Diazem C18 series EC Diazem C18 Dual-Zone Discovery C18 Discovery HS C18 Discovery Bo Wide Pore C18 Econosil C18 Econosphere C18 Everest C18 Fortis C18 Fortis H2o Gammabond C18 Gemini C18 Gemini-NX Genesis C18 GP-C18 GraceSmart C18 Haisil 100, 300 C18 Haisil HL C18 HALO C18 Hi Chrom Reversible ODS Hibar II C18 Hibar ODS HP-C18 HxSil C18 HydroBond PS-C18 High Strength Silica (HSS) HPLC C18 High Strength Silica (HSS) HPLC C18 SB High Strength Silica (HSS) HPLC T3 Hyperprep HS C18 Hypersil 100 C18 Hypersil BDS C18 Hypersil GOLD Hypersil GOLD aQ Hypersil-ODS Hypersil ODS-2 Macrosphere 300 C18 MetaSil AQ C18 MetaSil ODS MetroSep C18H MetroSep C18AQ MetroSep C18 AQ+ MetroSep C18SH MetroSep C18EPS MPLC RP-18 µBondapak C18 µBondapak C18 Radial-Pak Microsorb C18 MicroSpher C18 MonoChrom C18 nanoAcquity BEH130 nanoAcquity BEH300 Nova-Pak C18 NUCLEODUR C18 Gravity NUCLEODUR C18 Isis NUCLEODUR C18 Pyramid NUCLEODUR 100 C18 ec NUCLEODUR Sphinx RP Nucleosil 100 C18 Nucleosil C18 Nucleosil C18 AB Nucleosil C18 HD Nucleosil C18 Nautilus Nucleosil C18 MPN Nucleosil C18 PAH Nucleosil C18 PPN ODS Ultrapak OmniSpher C18 Onyx C18 Partisil ODS Partisil ODS2 Partisil ODS3 Partisphere C18 Peco HCODS Pecosil C18 Pecosphere 3x8C Pecosphere 3x8CR C18 Pecosphere 5x15CRT C18 Pecosphere-3C C18 Pecosphere-3CR C18 Pecosphere-3x3 C18 Peptica C18 serresA,B,C Pfarma C18 series A,B,C Pinnacle II C18 Pinnacle DB C18 Pinnacle DB Aqueous C18 Platinum C18 Platinum EPS C18 Platinum 300 C18 Polaris C18-A Polaris C18-Ether Polygosil 60-7 C18 Polygosil C18 HPLC C18 column listing (L1-USP)

5 Shodex ODSpak F-411 Shodex Silica C18M Shodex Silica C18P Spectraphysics C18 Spheri-(5 and 10) RP-18 Spheri-(5 and 10) ODS C18 Spherisorb ODS1 Spherisorb ODS2 Spherisorb ODS-B STR ODS-II STR ODS-M Sunfire C18 Supelcosil LC-18 Supelcosil LC-18-DB Supelcosil LC-318 Superiorex Superspher 100 RP-18 Symmetry C18 SymmetryPrep C18 Symmetry300 C18 SymmetryShield RP18 SynChropak RPP Polygosil C18 60ª Premier C18 Prep Nova-Pak HR C18 Prevail C18 Prevail OA Prevail Select C18 PrimeSphere C18-HC Prodigy ODS(3) ProntoSIL C18 H ProntoSIL C18 SH ProntoSIL C18 AQ ProntoSIL C18 AQ Plus ProntoSIL C18 ace-EPS ProSphere 300 C18 ProSphere 300 PAH ProSphere HP C18 ProSphere HP C18 AQ ProSphere HP C18 ZAP! ProteCol-G C18 HQ203 ProteCol-P C18 HQ105 ProteCol-P C18 HPH125 ProteCol-P C18 GP125 Purospher RP-18 Purospher RP18e Purospher STAR RP18e Pursuit C18 Pursuit XRs C18 Radial-Pak C18 Reduced Activity C18 Regis ODS Resolve C18 Resolvex C18 Sarasep P C18 Scherzo SM-C18 SelectaPore 90M, 300P, 300M Wakosil C18 RS 3 and 5 Wakosil 5C18 HG and AR Wakopak Wakosil-II 3C18 HG Wakopak Wakosil-II 3C18 AR Wakopak Wakosil-II 3C18 RS Wakopak Wakosil-II 5C18 Wakopak Wakosil-II 5C Wakopak Wakosil-II 5C18 HG Wakopak Wakosil-II 5C18 AR Wakopak Wakosil-II 5C18 RS Wakopak Wakosil-II 5C18 HG Prep Wakopak Wakosil-II 5C18 AR Prep Wakopak Wakosil-II 5C18 RS Prep Wakopak Handy ODS Wakopak Wakosil 5C18 Wakopak Wakosil 5C18T Wakopak Wakosil 5C18N SynChropak RPP-1000 SynChropak RPP-4000 SynChropak RPP-100 SynChropak RPS-100 Synergi Fusion-RP Synergi Hydro-RP Targa C18 TSK-GEL ODS-100V TSK-GEL ODS-100Z TSK-GEL ODS-100S TSK-GEL Super-ODS TSK-GEL ODS-80Tm TSK-GEL ODS-80Ts TSK-GEL ODS-120A TSK-GEL ODS-120T TSK-GEL ODS-140HTP TSK-GEL OligoDNA-RP Ultra C18 Ultra Aqueous C18 Ultra II C18 Ultra II Aqueous C18 Ultracarb ODS Ultrasphere ODS Ultrasphere C-18 Ultrasphere C-18-IP Ultron VX-ODS Unison UK-C18 Unison US-C18 Velosep RP-18 VisionHT C18 VisionHT C18-B VisionHT C18HL VisionHT C18-P Viva C18 Vydac 201TP Vydac 202TP Vydac 218MS Vydac 218TP Vydac 238MS Vydac C18 HPLC C18 column listing (L1-USP)

6 YMC-Pack Hydrosphere C18 YMC-Pack ODS-A YMC-Pack ODS-AL YMC-Pack ODS-AM YMC-Pack ODS-AQ YMC-Pack Pro C18 YMC-Pack Pro C18 RS Zodiac 120 C18H Zodiac 120 C18AQ Zodiac 120 C18 Eurobond Zodiac 120 AC EPS Zodiac 120 C18SH Zodiac ODS Zodiac ODS H ZORBAX ODS C18 ZORBAX ODS Classic ZORBAX Eclipse Plus C18 ZORBAX Eclipse PAH ZORBAX Eclipse XDB-C18 ZORBAX Extend-C18 ZORBAX Rx C18 ZORBAX StableBond C18 ZORBAX 300 StableBond C18 Wakopak Wakosil 10C18 Wakopak Wakosil 5C18 AR Wakopak Wakosil 5C Wakopak Wakosil 5C18-200T Wakopak Wakosil 5C18-200N Wakopak Wakosil 10C Wakopak MS-5C18 GT Wakopak Navi C18-5 Wakopak Combi ODS Wakopak Combi ODS Fast XBridge BEH130 XBridge BEH300 XBridge C18 XBridge OST C18 XBridge Shield RP18 XTerra MS C18 XTerra RP HPLC column as C18 in 2010 Which is the best one! Do I need to buy all! What is my C18 strategy?...etc... HPLC C18 column listing (L1-USP) $$$

7 A new modern strategy is absolutely necessary in 2010 for HPLC end-users New Generic Approach to Find Appropriate HPLC column (Total Solution for HPLC end-users) Better matrix (silica-gel) -Stationary Phase Specification (manufacturers) Phase according to Polarity (acidic and basic analytes) Phase according to Hydrophobicity (% carbon loading) Phase according to Low Metal Content and Silianol Activity Phase compatible with new LC-MS technology Phase for very aqueous mobile phase (100% buffer or water) Etc... Note: Every Pittcon and Analytica Events present new C18, more and more HPLC C18/year

8 New Modern HPLC Strategy How to improve today silica-based HPLC columns? Better bonding chemistry/ improved control for better reproducibility. What about the matrix? (supportsilica) High Quality Level of the Support (Silica) narrowest particule size distribution perfectly spherical high level of purity High surface area (m 2 /g) Efficiency pore size (Å) No fines SiliaSphere (SiliCycle) Low metal content High mechanical stability Reproducible loading (critical in reversed-phase) X X Supplier A (top) Supplier B (bottom) (both are real HPLC supports)

9 Generic Approach/Most popular applications Now better support (Silica) Similar chemistries than other HPLC suppliers Improved quality and efficiency HPLC column for low pH (not hybrid, pure silica based) HPLC column for high pH (pseudo-hybrid) HPLC column for 100% aqueous mobile phase (or buffer) HPLC column for LC-MS (no bleeding) HPLC column for QC/QA validation and method development (easy method) No tailling, better sensitivity and peak shape for quaternary ammonium compounds) HPLC column for HILIC technology (extremely polar compounds)

10 Portofolio of SiliaChrom Columns A column for every need: ONLY 7 Chemistries! SiliaChrom SB C18 and SB C8 SiliaChrom XDB1 C18,C8, C4, CN, AMINO, PHENYL, DIOL, Silica… SiliaChrom XDB and XDB2 (C8&C18) SiliaChrom AQ C18 (dtC18) SiliaChrom HILIC SiliaChrom XT C18 (XT Fidelity C18) NO MORE CHEMISTRIES

11 SiliaChrom-Chemistries Review Silica-Based HPLC columns for Extremely Acidic Compounds H3O+H3O+ Steric Effect Note: Not Endcapped silica gel H3O+H3O+ R=C8 or C18, pH<2.0 New Technology

12 SiliaChrom-Chemistries Review Silica-Based HPLC columns for all type of compounds (pH=2-9) Note: Partially End capped silica gel TMS endcapped Free silianol groups (OH) R=C18 Ratio of TMS-OH-C18 is well-balanced New Technology Polar Surface

13 SiliaChrom-Chemistries Review Silica-Based HPLC column for EXTREMELY POLAR compounds Note: Not Endcapped silica gel SiliaChrom HILIC Normal-Reversed Phases Conditions: Only SiliaChrom HILIC

14 SiliaChrom-Chemistries Review Silica-based HPLC columns for low to high hydrophobic compounds pH= Note: Double Endcapped silica gel SiliaChrom XDB C8 (8%) and C18 (15%) SiliaChrom XDB1 C8 (14%) and C18 (22%) SiliaChrom XDB2 C18 (18%) 1) Different carbon loadings (8-22%C) 2) XDB2=Luna C18(2) 3) XDB1= Phenyl, NH 2, CN, 300A, etc…

15 SiliaChrom-Chemistries Review Silica-Based HPLC columns for EXTREMELY Basic compounds pH= SiliaChrom XT C18 Series Replace OH by CH 3 (Pseudo-Hybrid tech.) No bridge (No Backpressure issue as XBridge (Waters)) New Technology

16 SiliaChrom-Chemistries Review

17 SiliaChrom Series HPLC Classification-USP/NIST Approaches All SiliaChrom HPLC columns are evaluated by USP and NIST approaches: Uracil: void volume marker Toluene: hydrophobic retention Ethylbenzene: methylene selectivity and plate count determination Amitriptyline: activity towards bases (silianol activity evaluation) (A) Quinizarin: activity towards chelating reagents (metal content evaluation) (B) ( Pharmacopeial Forum, Vol. 31(2) March-Apr. 2005, p.637) (NIST, SRM 870, Certificat Issue Date:December 19th 2002, Valid until September 30th 2010) (B)(A)

18 SiliaChrom Evaluation HPLC System: Thermo Surveyor with PDA (ChromQuest-Data handling version 5.0) Mobile Phase: MeOH/buffer (80/20, v/v) Buffer: 20mM of phosphate buffer adjusted at pH=7.0 Flow rate: 1.0 mL/min Detector: UV scan (PDA), Total scan nm Temperature: 23 C All HPLC columns: 4.6x150mm, 5µm

19 Time (min) uAU Phenomenex Luna C18(2) Uracil-Toluene-Ethylbenzene-Quinizarin-Amitriptyline Low Plate Count (see Results Table)

20 Time (min) uAU Agilent Zorbax SB C18 Uracil-Toluene-Ethylbenzene-Quinizarin-Amitriptyline Important Tailling

21 Time (min) uAU Waters Sunfire C18 Uracil-Toluene-Ethylbenzene-Quinizarin-Amitriptyline Asymetry Metal Contamination

22 Time (min) uAU SiliaChrom XDB2 C18 Uracil-Toluene-Ethylbenzene-Quinizarin-Amitriptyline Very good peak shapes

23 HPLC Column K Ethylbenzene Ethyl/Tol N Ethylbenzene RA S (Quinizarine) Tf (Amitriptyline) SiliaChrom XDB2 C Luna C18(2) Zorbax SB C Sunfire C SiliaChrom XDB1 C Summary Results-Case Study Basic analyte: SiliaChrom XDB2 Less polar analyte: SiliaChrom XDB1 Basic analyte: SiliaChrom XDB2 Less polar analyte: SiliaChrom XDB1

24 Recent SiliaChrom Applications Acidic and Basic Compounds (8) with one SiliaChrom HPLC column (AQ C18) Fluoroquinolones for Veterinary use by HPLC-Marbofloxacin and Sarafloxacin Halogenated Herbicides by HPLC-Dicamba, Atrazine and 2,4-D Catecholamines by HPLC- Polar Compounds Faster HPLC with SiliaChrom-Polar and mid-polar Compounds Isomer separation for Chlorophenol series by simple HPLC method HPLC study for Naloxone-pH=2.5, 7.0 and 10.5 HPLC Chromatogram of Ganciclovir– Polar Compound (diol-amine) HPLC Chromtogram of CefotaximeExtremely Polar Compound (acid-amine-sulfur) And more….

25 Recent SiliaChrom Applications Acidic and Basic Compounds (8) with same SiliaChrom HPLC column 1-Maleic Acid 2-Acetaminophen 3-Pseudoephedrine 4-Chlorpheniramine 5-Benzoic Acid 6-Salicylic Acid 7-Dichlorphen 8-Ibuprofen

26 Recent SiliaChrom Applications Acidic and Basic Compounds (8) with same SiliaChrom HPLC column Phenomenex Luna C18(2) 5µm 4.6x150mm nm 210 nm 230 nm

27 Recent SiliaChrom Applications Acidic and Basic Compounds (8) with same SiliaChrom HPLC column (same HPLC conditions than Luna C18(2)) SiliaChrom AQ C18 5µm 4.6x150mm nm 230 nm

28 Recent SiliaChrom Applications Fluoroquinolones for Veterinary use by HPLC-Marbofloxacin and Sarafloxacin SiliaChrom AQ C18 5µm 4.6x150mm Waters Atlantis dC18 5µm 4.6x150mm Asymmetry SiliaChrom AQ C18 Atlantis dC Zwitterion Molecules Note: SiliCycle has the perfect equivalent (better) with SiliaChrom AQ C18 (or dtC18) than Atlantis dC18 (Waters)

29 Recent SiliaChrom Applications Halogenated Herbicides by HPLC-Dicamba, Atrazine and 2,4-D SiliaChrom dtC18 5µm 4.6x150mm

30 Recent SiliaChrom Applications Halogenated Herbicides by HPLC-Dicamba, Atrazine and 2,4-D Supelco Ascentis C18 5µm 4.6x150mm Asymmetry SiliaChrom dtC18Ascentis C

31 Recent SiliaChrom Applications Catecholamines by HPLC-Polar Compounds SiliaChrom AQ C18 5µm 4.6x150mm Mobile Phase: Water 1.0% AcOH only! Epinephrine Norepinephrine Dopamine Tyrosine Levodopa

32 Recent SiliaChrom Applications Faster HPLC with SiliaChrom-Polar and mid-polar Compounds SiliaChrom AQ C18 5µm 3.0x50mm Caffeine Chlorthalidone Albendazole Warfarin Tolnaftate Dioctyl Phatalate 95/5 0.1%TFA in H 2 O/ACN – 5/95 0.1% TFA in H 2 O/ACN in 2.25 min mL/min, 23°C, 254 nm (LC-MS conditions from AstraZeneca Canada) Test Mix to evaluate HPLC column from AstraZeneca (Canada)

33 Recent SiliaChrom Applications SiliaChrom XDB1 PHENYL 5µm 4.6x150mm 60/40, H 2 O/MeOH, 1.00 mL/min, 23°C, 254nm : Easy Method!

34 Recent SiliaChrom Applications HPLC study for Naloxone-pH=2.5, 7.0 and 10.5 SiliaChrom XT C18, 5µm, 4.6x150mm 80/20, Phosphate Buffer/MeOH, 1.00 mL/min, 23°C, 254nm pH=2.5 50/50, Phosphate Buffer/MeOH, 1.00 mL/min, 23°C, 254nm pH=7.0 Fronting ? Assymetry = 0.89 Assymetry = 0.85

35 Recent SiliaChrom Applications HPLC study for Naloxone-pH=2.5, 7.0 and 10.5 SiliaChrom XT C18, 5µm, 4.6x150mm 50/50, H 2 O/MeOH 0.1% NH 4,OH, 1.00 mL/min, 23°C, 254nm pH=10.5 Assymetry = 0.99 Remember, pH range for XT is Good Chromatography Tool

36 SiliaChrom AQ C18 (4.6x150mm, 5µm) MeOH:Water, 5:95 UV 252nm 1mL/min SiliaChrom AQ C18 (4.6x150mm, 5µm) MeOH:Water, 5:95 UV 252nm 1mL/min HPLC Chromatogram Ganciclovir – Polar Compounds (diol-amine) Recent SiliaChrom Applications

37 SiliaChrom AQ C18 (4.6x150mm, 5µm) 0.4 mM KH2PO4 + 8mM K2HPO4: MeOH, 89:11 UV 254nm 1mL/min SiliaChrom AQ C18 (4.6x150mm, 5µm) 0.4 mM KH2PO4 + 8mM K2HPO4: MeOH, 89:11 UV 254nm 1mL/min HPLC Chromatogram of CefotaximeExtremely Polar compound (acid-amine-sulfur) Recent SiliaChrom Applications

38 SiliaChrom XDB1 C18 (4.6x150mm, 5µm) ACN:Water, 70:30 UV 254nm 1mL/min SiliaChrom XDB1 C18 (4.6x150mm, 5µm) ACN:Water, 70:30 UV 254nm 1mL/min HPLC for mixture of Fenitrothion-Pyriproxyfen Different Level of Hydrophobicity Recent SiliaChrom Applications

39 SiliaChrom XDB2 C18 (4.6x150mm, 5µm) SiliaChrom XDB2 C18 (4.6x150mm, 5µm) HPLC Determination for Ephedrine (from Ephedra Sample) Zorbax SB C18 (4.6x150mm, 5 µm) Zorbax SB C18 (4.6x150mm, 5 µm) Recent SiliaChrom Applications

40 HPLC chromatogram of Berberine-Metal Content Effect SiliaChrom dtC18 (4.6x250mm, 5µm) ACN: 1%TFA, 35:65 UV 346nm 1 mL/min SiliaChrom dtC18 (4.6x250mm, 5µm) ACN: 1%TFA, 35:65 UV 346nm 1 mL/min Sunfire C18 (4.6x250mm 5µm) Recent SiliaChrom Applications

41 SiliaChrom SB C18 (4.6x150mm, 5µm) 59:39:2 ACN:Water:TFA (pH = 1.00) UV 270 nm, 1 mL/min SiliaChrom SB C18 (4.6x150mm, 5µm) 59:39:2 ACN:Water:TFA (pH = 1.00) UV 270 nm, 1 mL/min Durability Testing at pH-1.00 SiliaChrom SB C18 ( (Uracil, Phenol, Toluene and Ethylbenzene) Time/h Retention time (Ethylbenzene) (min) Tailing Factor (Ethylbenzene) (USP) Theoritical Plates # (Ethylbenzene) (USP) SD Mean RSD (%) hr 120 hrs Poster presented at Pittcon 2010, Orlando FL, USA LC-GC Application NoteSeptember 2009

42 Time/hTailing Factor Retention time (Naphthalene) (min) hr 400 hrs SiliaChrom XT C18 (4.6x150mm, 5µm) MeOH: Water/TEA, pH=11.5, 40:60 UV 215nm, 1 mL/min SiliaChrom XT C18 (4.6x150mm, 5µm) MeOH: Water/TEA, pH=11.5, 40:60 UV 215nm, 1 mL/min Durability Testing at pH-11.5 SiliaChrom XT C18 Naphthalene Uracil, Phenol, Nitrobenzene and Naphtalene

43 SiliaChrom XT C18 (4.6x150mm, 5µm) (ACN-Water-TEA) 90 C, UV 270 nm 1 mL/min SiliaChrom XT C18 (4.6x150mm, 5µm) (ACN-Water-TEA) 90 C, UV 270 nm 1 mL/min Durability Testing at pH=11.50, 90 C SiliaChrom XT C18 Fidelity ( (Uracil, Phenol, Toluene and Ethylbenzene) 0 hr 120 hrs

44 SiliaChrom HILIC (4.6x150mm, 5µm) ACN 0.1% TFA: Water 0.1%TFA, 10:90 UV 280nm 1 mL/min SiliaChrom HILIC (4.6x150mm, 5µm) ACN 0.1% TFA: Water 0.1%TFA, 10:90 UV 280nm 1 mL/min Waters HILIC Silica HPLC column (4.6x150mm, 5 µ m) Waters HILIC Silica HPLC column (4.6x150mm, 5 µ m) Vitamin B1, Vitamin B6, Vitamin C and Vitamin B2 SiliaChrom HILIC, Vitamins B1 B6 C B2

45 Generic Approach with SiliaChrom Series Final StepPolarityResolution Acidic, Basic or Neutral Compounds SiliaChrom AQ (MeOH/H2O) or (ACN/H2O), pH=2-9 Not enough retention Acidic SiliaChrom SB pH=1-2 or HILIC Basic SiliaChrom XT pH=9-12 Series or HILIC SiliaChrom XDB, XDB1 or XDB2 Good Retention, but not enough resolution SiliaChrom SB, XT or XT Fidelity (low and/or high pH) Neutral Note: SiliaChrom AQ is an universal HPLC column (see how particle is designed : TMS-OH-C18) 87% of HPLC success in my Lab !

46 Conclusion Low pHHigh pHHigh level of WaterHydrophobicityLow MetalPolarExtemely ( )(9-12)Mobile Phase(different Level)ContentAnalytePolar Analyte SiliaChrom SBXTAQ XDB, XDB1 XDB2dtAQHILIC ONLY 8 Chemistries to cover more than 85% of Regular HPLC Application using Silica-Based as support Be aware for your HPLC column strategy, Marketing is not real SCIENCE!

47 Thank you!


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