BÜCHI Labortechnik AG Purification
BUCHI Chromatography History 1977: BUCHI Fraction Collector 1978: 1 st preparative chromatography system 2008: automated chromatography system 2014: automated high performance chromatography system 2016: automated high performance chromatography systems with integrated ELSD acquired
BUCHI Chromatography System Portfolio Flash and Flash/Prep Systems: Reveleris PREP (120bar) Reveleris X2 and X2-UV (14bar) Sepacore X10/ X50 (10bar/ 50bar) Sepacore Easy Synthesis and Extract (10bar/ 50bar) Reveleris PREP Reveleris X2 Reveleris X2-UV Sepacore X10 Sepacore X50 Sepacore Easy Synthesis Sepacore Easy Extract
Reveleris X2 and PREP Overview Portfolio Price UV-Vis Upgrade ELSD Upgrade UV-Vis Upgrade Reveleris Prep Reveleris X2 UV-Vis Upgrade Navigator Upgrade Reveleris X2 (upgradeable) Reveleris X2-UV UV-Vis Upgrade Functionality
Reveleris X2 and PREP Overview Specifications
Reveleris X2 and PREP Overview Specifications Reveleris X2/ X2-UV/ X2 upgradeable Reveleris PREP Pressure 200psi (= 14bar) Up to 1700psi (=120bar) Flow Detectors Columns Solvent delivery Sample injection 1-200mL/min (Accuracy +/- 3%, precision +/- 0.5% for 4-200 ml/min) UV (X2-UV) + ELSD option (X2 upgradeable) + ELSD (X2) Combination of 3 UV and 1 ELSD signal Flash up to 330g; Flash up to 1.5kg (off unit) 4 independent channels; usage of 2 during a run Liquid via a syringe; solid via a solid loader 1-200mL/min (Accuracy +/- 3%, precision +/- 0.5% for 4-200 ml/min) UV+ELSD Combination of 3 UV and 1 ELSD signal Flash up to 330g; Flash up to 1.5kg (off unit); Preparative LC columns 10mm to 75 mm id > =5µ media 4 independent channels; usage of 2 during a run Liquid via a loop + electrical valve; solid via a solid loader
Reveleris X2 and PREP Advanced Software with an Intuitive Interface
Easy to Use Software Viewing Easy view 12 touchscreen Run Setup Fast method set up Auto cartridge recognition populates Adjustable time, flows and pressures Change parameters on the fly Select detection monitoring and threshold settings Simple single screen operation accelerates system set-up Control Upfront and intuitive action buttons
Reveleris X2 and PREP Combination of UV and ELS Detection
Limitations of traditional UV only Chromatography Chromatogram does not reflect sample composition due to differing adsorption coefficients Non-chromophoric molecules are not detected - can t use for nonchromophoric targets and impurities Choice of solvents can be limited by UV interference (e.g. acetone) Purification is not efficient due to additional work required
How does ELSD work? Step 1: Nebulization Column effluent mixes with gas flow to form a dispersion of droplets Step 2: Mobile Phase Evaporation Mobile phase evaporates from around sample particle leaving dried particles in solvent vapor Step 3: Detection Sample particles pass through laser light in a flow cell. The particles scatter light that is collected by a photodiode ELSD detects any compound less volatile than mobile phase
Benefits of ELSD versus UV UV detector limitations: Only detects compounds that absorb UV or visible radiation Detection based on extinction coefficient Baseline drift with gradients at low wavelength Solvent front interference at low wavelength Solvent limited by UV cut-off ELSD benefits: Detects any compound less volatile than the mobile phase Near equivalent response Little or no baseline drift during gradients No response from solvents Compatible with all volatile mobile phases
ELSD Quantification UV signal is variable and response dependent on extinction coefficients UV Peak 3 Peak 1 Peak 2 Weight Percent Material Actual UV Peak 1 42% 5% Peak 2 40% 5% Peak 3 18% 90% ELSD detection provides a more accurate estimate of relative weight percent and purity than UV-only detection
ELSD Quantification ELSD signal strength directly proportional to compound mass Peak 1 ELSD Peak 2 Peak 3 Weight Percent Material Actual ELSD Peak 1 42% 40% Peak 2 40% 40% Peak 3 18% 20% ELSD detection provides a more accurate estimate of relative weight percent and purity than UV-only detection
ELSD Allows Detection of all the Components in a Sample Nonchromophores NOT VISIBLE UV Nonionized compounds NOT VISIBLE MS Detects all non volatiles ELSD Detecting all components eliminates guesswork and wasted time!
Active Splitting of the Sample Flow ELSD is a disruptive detection method (sample loss) An active splitter allows to minimized sample loss Flow from the outlet of the column Flow from the make-up pump Flow from the outlet of the column Flow from the make-up pump Mass to be transferred Mass transferred Rapid switching valve Switch Flow to the UV detector and fraction collector Flow to the ELSD Reveleris sample loss: 30µl/min Flow to the UV detector and fraction collector Flow to the ELSD
Advantages of the Reveleris X2/ Prep integrated ELSD Sample loss only at 30µl/min No setting adjustment (temperature) needed The dry air supply (accessory) provides compressed and moisture and oil free air Automatic switch ON/OFF of the air supply
What is RevealX Detection Technology? Patented Detection Technology Intelligently combines multiple UV and ELSD signals Triggers fractions based on the combined signal Automatically accounts for signal delay between UV and ELSD Does not require detector optimization for each run Have confidence that you are detecting everything from your sample and fractions are being triggered correctly
Detect Hidden Unknowns with ELSD and RevealX Detection Technology UV-inactive impurities not detected or resolved Triggers collection of 1 impure fraction UV
Detect Hidden Unknowns with ELSD and RevealX Detection Technology Significant quantities of nonchromophoric impurities present. Triggers collection of 2 pure fractions UV ELSD Detecting more components from a crude mixture not only reveals more about the sample, but frees time for more value-added work!
Reveleris X2 and PREP Reveleris Navigator Software: From TLC to Flash Method
Reveleris Navigator Technology Improves target purities and recovery, increasing your productivity Simplifies method optimization by interpreting TLC or HPLC scouting runs Reduces solvent waste costs Patented algorithm allows a choice to optimize for the fasted run, or the highest recovery Warns if the compound is difficult to separate with selected solvents, before you begin the run is an optional upgrade can be purchased with your system, or added to your system later
Reveleris Navigator Technology Flash mode method development using the Reveleris Navigator software feature optimizes the purification separation and gives confidence of first-time success. Software Speed Method Reduce concerns when scaling up closely eluting components on flash
Reveleris Navigator Technology Flash mode method development using the Reveleris Navigator software feature optimizes the purification separation and gives confidence of first-time success. Software Purity Method Reduce concerns when scaling up closely eluting components on flash
Reveleris Navigator Technology Reveleris Navigator Purity Method Reveleris Speed Method Traditional Method Method Development Time (min) 52 52 78 Solvent Used (ml) 50 50 75 Resolution 2.9 2.04 1.50 Method Performance Time 8.1 4.5 19.2 Solvent Used (ml) 292 162 691 Total Time (min) 60.1 56.5 97.2 Total Solvent Used (ml) 342 212 767 Reveleris Navigator TM Purity Method: Improved resolution 2x from 1.5 to 3 Reveleris Navigator TM Speed Method: Reduce run time from 19 to 4.5 minutes and reduce solvent use from 766 to 212 ml
Reveleris PREP Flash and prep HPLC in a Single Compact System
Reveleris PREP Benefits Time and Cost Savings Can be used in flash and preparative modes separately or sequentially, saving time by performing preparative LC on the same system Dual-function capability in a single system saves space in the laboratory, and helps to reduce the total capital cost of flash and preparative LC purification by 40%-50% Control Allows the user to seamlessly switch between flash and preparative modes with a single click The ability to perform flash and preparative LC on a single system, allows the user to maintain custody of the sample Efficiency Perform a simple flash separation to clean up samples before injecting onto a preparative LC column, preventing degradation of expensive preparative LC columns and prolonging their lifetimes
Easy Switch Between Flash and Prep HPLC Mode One Click to Switch Modes Prep Mode Screen Flash Mode Screen
Inverted Cartridge Flow Direction in Reveleris PREP and X2 Uniform flow of trapped air Better separation of unpolar compounds when working woth C18 columns Flow
BUCHI Chromatography Column Portfolio Flash Cartridges Reveleris (RFID) GraceResolv (no RFID) Prep HPLC Columns
Reveleris and GraceResolv Cartridges The main difference between Reveleris and GraceRresolv Cartridges is the RFID tag The RFID tag allows to follow the history of the column and automatically sets the optimal flow rate and max pressure limit Only availabel with Reveleris Cartridge
Reveleris Cartridges 40µm High capacity cartridges improve results & reduce costs Better peak shapes and resolution from robust & efficient packing Collect higher purity fractions and avoid collecting non-productive fractions due to better resolving cartridges Superior resolution - increased yields and higher recovery Load more in one injection on equivalent sized cartridge Cut costs by optimizing the cartridge size while increasing yield and loading Irregular particles with 60Å pore size
Reveleris Cartridges 40µm Chemistry Silica C18 Amino Diol Ideal For Conventional applications, polar compounds Non-polar compounds Better peak shapes with polar and basic compounds, ideal for carbohydrates Alternate selectivity for a variety of polarity types, ideal for lipids Stationary Phase Silica Cartridge Size 4g, 12g, 24g, 40g, 80g, 120g, 220g, 330g C18 4g, 12g, 24g, 40g, 80g, 120g, 220g, 330g Amino/ Diol 4g, 12g, 40g,
Reveleris Cartridges 40µm Grace Flash Cartridge Competitor Flash Cartridge Cartridge: 40g Silica Cartridge Sample: Peak 1: Toluene Peak 2: Dimethyl Phthalate Mobile Phase: Hexane/Ethyl Acetate (70/30) Flow Rate: 40mL/min Detector: UV at 254nm Peaks are more fully resolved at identical sample load on a higher capacity silica.
Reveleris Cartridges 40µm Reveleris Flash Cartridge Other Flash Cartridge 3 impurity peaks 7 fractions 0 impurity peaks 19 fractions Efficient packaging processes Better peak shapes and resolution Ability to withstand higher pressures and flow rates Smaller fraction volumes Less fractions Fewer, higher purity fractions result from better resolving cartridges
Reveleris Cartridges 40µm Reveleris Cartridge Competitor A Cartridge Competitor B Cartridge Fraction Purity Analysis by TLC Fraction Purity Analysis by TLC Fraction Purity Analysis by TLC Yield: 76% Yield: 23% Yield: 26% Improved resolution yields 3 times higher recovery even in cases of poor peak shapes
Reveleris Cartridges 40µm Reveleris Flash Cartridge 9.12g Other Flash Cartridge 3.84g Grace Reveleris 9.12g Cartridge Competitor 3.84g Cartridge Load more in one injection on equivalent sized cartridge In scale-up, process more grams with less injections Reduce consumable, solvent and waste costs Depending on application and purification requirements, studies have indicated between a 30-65% savings
Reveleris Cartridges HP 20µm Covers all advantages of the standard Reveleris Cartridges High resolution 20µm silica media with low fines Ideal for complex samples Irregular particles with 60Å pore size Stationary Phase Silica Cartridge Size 4g, 12g, 24g, 40g, 80g, 120g, 220g
Reveleris Cartridges HP 20µm Reveleris 40g Cartridge Neem Oil, 500mg on a standard Reveleris flash cartridge Reveleris HP 40g Cartridge Neem Oil, 500mg on a Reveleris HP flash cartridge Reveleris HP flash cartridges offer superior performance with greater efficiency and resolution
Reveleris C18-WP Flash Cartridges 20µm High resolution 20µm silica media with low fines Ideal for compounds such as peptides, proteins, lipids, biopharmaceuticals and carbohydrate based molecules Suitable for molecular weights up to 70,000 daltons Irregular particles with 150Å pore size Stationary Phase Silica Cartridge Size 4g, 12g, 40g
Reveleris C18-WP Flash Cartridges 20µm Comparison of peptide retention and peak shapes on Reveleris C18 and Reveleris C18-WP flash cartridges. Chromatograms of small, medium and large sized peptide/protein are shown. Z-Gly-Phe (MW 357) Lysozyme (MW 14700) Bovine Serum Albumin (MW 66000) Reveleris C18 cartridge Reveleris C18 cartridge Reveleris C18 cartridge Reveleris C18-WP cartridge Reveleris C18-WP cartridge Reveleris C18-WP cartridge Reveleris C18-WP cartridges can be used to 70,000 Daltons
Reveleris C18-WP Flash Cartridges 20µm Mixture of Small Peptides (Ampicillin + Piperacillin) Mixture of Small Molecules (Eugenol + Citral) Reveleris C18 cartridge, 40µm, 60Å Reveleris C18 cartridge, 40µm, 60Å Reveleris C18-WP, cartridge 20µm, Wide Pore Reveleris C18-WP, cartridge 20µm, Wide Pore Baseline separation is achieved for both large and small molecules and their various impurities. with the Reveleris C18-WP 20µm flash cartridge
GraceResolv Cartridges Same silica material as Reveleris Cartridges Low budget consumable: no RFID Irregular particles with 60Å pore size Two hardware options for outlet: Luer slip or Luer lock (with Luer slip adapter) Stationary Phase Silica (Luer slip outlet) Silica (Luer lock outlet with Luer slip adapter) Cartridge Size 4g, 12g, 24g, 40g, 80g, 120g, 220g, 330g, 750g, 1500g 4g, 12g, 24g, 40g, 80g, 120g, 220g, 330g
BUCHI Prep HPLC Column Portfolio High resolution achieved using spherical and small particle size silica Constant quality thanks to the ultra-pure spherical silica Purification capacity of up to 1 g sample on columns with an ID up to 50 mm Stationary Phase Prep HPLC Column Size Silica 10µm, 60Å Silica 15µm, 60Å C18 10µm, 100Å C18 15µm, 100Å 10, 20, 30, 50mm ID 150, 250mm lengths 10, 20, 30, 50mm ID 150, 250mm lengths 10, 20, 30, 50mm ID 150, 250mm lengths 10, 20, 30, 50mm ID 150, 250mm lengths
Sample Loading Techniques Loading Technique Liquid loading Solid loading Option 1 Prep injector Sample injected via the loop Dry loading column replacing the loop Option 2 Flash injector Sample loaded directly on top of the purification column Dry loading column connected on top of the purification column
Reveleris Solid Loader Available sizes empty tubes Available sizes silica filled tubes 3g, 15g, 40g 3g, 15g, 40g
Reveleris Empty Screw Cap Solid Loaders Available sizes 12g, 20g, 40g
Why Buy a Reveleris Purification System? BUCHI is a leading innovator in flash chromatography technology Buchi is the only flash instrument manufacturer with its own ELSD technology, specifically optimized for use in Reveleris purification systems Buchi has many years experience with patented ELSD technology Buchi is the only supplier of a higher pressure capable flash instrument with a fully integrated and self optimizing ELSD Buchi s dual detection technology offers superior performance over the competition Buchi has significant IP in flash technology Buchi has expertise in packing flash cartridges and Prep columns Buchi has global reach and support available to customers
BÜCHI Labortechnik AG Sepacore instrumentation
Sepacore Flash Systems X10 / X50 Binary gradient up to 250mL/min, 10 bar (X10) or 50 bar (X50) Injection valve for liquid sample injection (0 20 ml) Fraction collection, 4 racks capacity UV-Vis detection Up to 8 signal inputs SepacoreControl software Integrated touchscreen or separate computer control
Questions?