SHIMADZU Solutions for Science Since 1875 Since 1875 HPLC Method Development Doug Catron Field Sales...
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![Page 1: SHIMADZU Solutions for Science Since 1875 Since 1875 HPLC Method Development Doug Catron Field Sales Engineer Shimadzu Scientific Instruments, Inc. .](https://reader035.fdocuments.us/reader035/viewer/2022062407/56649cca5503460f9499313c/html5/thumbnails/1.jpg)
SHIMADZUSHIMADZUSolutions for ScienceSolutions for Science Since 1875Since 1875
HPLC Method DevelopmentHPLC Method Development
Doug CatronDoug Catron
Field Sales EngineerField Sales Engineer
Shimadzu Scientific Instruments, Inc.Shimadzu Scientific Instruments, Inc.www.ssi.shimadzu.com
www.ssi.shimadzu.com/LCVAwww.ssi.shimadzu.com/LCVA
![Page 2: SHIMADZU Solutions for Science Since 1875 Since 1875 HPLC Method Development Doug Catron Field Sales Engineer Shimadzu Scientific Instruments, Inc. .](https://reader035.fdocuments.us/reader035/viewer/2022062407/56649cca5503460f9499313c/html5/thumbnails/2.jpg)
SHIMADZUSHIMADZUSolutions for ScienceSolutions for Science Since 1875Since 1875
Types Of MethodsTypes Of Methods
• PreparativePreparative• QuantitativeQuantitative• QualitativeQualitative
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SHIMADZUSHIMADZUSolutions for ScienceSolutions for Science Since 1875Since 1875
PreparativePreparative
• Sample PreparationSample Preparation• CapacityCapacity
• Big ColumnsBig Columns• High Flow RatesHigh Flow Rates
• Large Injection VolumesLarge Injection Volumes• Blinding The DetectorBlinding The Detector
![Page 4: SHIMADZU Solutions for Science Since 1875 Since 1875 HPLC Method Development Doug Catron Field Sales Engineer Shimadzu Scientific Instruments, Inc. .](https://reader035.fdocuments.us/reader035/viewer/2022062407/56649cca5503460f9499313c/html5/thumbnails/4.jpg)
SHIMADZUSHIMADZUSolutions for ScienceSolutions for Science Since 1875Since 1875
PreparativePreparative
Heart CuttingHeart Cutting
![Page 5: SHIMADZU Solutions for Science Since 1875 Since 1875 HPLC Method Development Doug Catron Field Sales Engineer Shimadzu Scientific Instruments, Inc. .](https://reader035.fdocuments.us/reader035/viewer/2022062407/56649cca5503460f9499313c/html5/thumbnails/5.jpg)
SHIMADZUSHIMADZUSolutions for ScienceSolutions for Science Since 1875Since 1875
QuantitativeQuantitative
• Sample PreparationSample Preparation• SeparationSeparation• DetectionDetection• CalibrationCalibration• QCQC
![Page 6: SHIMADZU Solutions for Science Since 1875 Since 1875 HPLC Method Development Doug Catron Field Sales Engineer Shimadzu Scientific Instruments, Inc. .](https://reader035.fdocuments.us/reader035/viewer/2022062407/56649cca5503460f9499313c/html5/thumbnails/6.jpg)
SHIMADZUSHIMADZUSolutions for ScienceSolutions for Science Since 1875Since 1875
QualitativeQualitative
• Sample PreparationSample Preparation• SeparationSeparation• DetectionDetection• Data ComparisonData Comparison
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SHIMADZUSHIMADZUSolutions for ScienceSolutions for Science Since 1875Since 1875
Sample PreparationSample Preparation
• SamplingSampling• HomogenationHomogenation• Sub samplingSub sampling• DissolutionDissolution• Clean UpClean Up• Chill OutChill Out
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SHIMADZUSHIMADZUSolutions for ScienceSolutions for Science Since 1875Since 1875
Clean UpClean Up
• Phase SeparationPhase Separation• DigitizationDigitization• Solid Phase ExtractionSolid Phase Extraction• Solid Phase Micro extractionSolid Phase Micro extraction• CentrifugeCentrifuge• Filtering!!!Filtering!!!
Don’t Overdo Don’t Overdo It!It!
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SHIMADZUSHIMADZUSolutions for ScienceSolutions for Science Since 1875Since 1875
Clean UpClean Up
![Page 10: SHIMADZU Solutions for Science Since 1875 Since 1875 HPLC Method Development Doug Catron Field Sales Engineer Shimadzu Scientific Instruments, Inc. .](https://reader035.fdocuments.us/reader035/viewer/2022062407/56649cca5503460f9499313c/html5/thumbnails/10.jpg)
SHIMADZUSHIMADZUSolutions for ScienceSolutions for Science Since 1875Since 1875
Now Lets Get To BusinessNow Lets Get To Business
Don’t Reinvent The WheelDon’t Reinvent The Wheel
•Standard MethodStandard Method
•Column ManufacturersColumn Manufacturers
Check Your SourcesCheck Your Sources
Standards vs. Real Standards vs. Real SamplesSamples
•MatrixMatrix
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SHIMADZUSHIMADZUSolutions for ScienceSolutions for Science Since 1875Since 1875
Tools Of The TradeTools Of The Trade
AffinityAffinity
•Reverse PhaseReverse Phase
•Normal PhaseNormal Phase
•Strong SolventStrong Solvent
Gel PermeationGel Permeation
Ion Pairing/Ion ExchangeIon Pairing/Ion Exchange
Chiral SeparationsChiral Separations
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SHIMADZUSHIMADZUSolutions for ScienceSolutions for Science Since 1875Since 1875
Move A Mountain One Peak At A Move A Mountain One Peak At A TimeTime
Lower RT = Stronger Lower RT = Stronger SolventSolvent
Higher RT = Weaker Higher RT = Weaker SolventSolvent
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SHIMADZUSHIMADZUSolutions for ScienceSolutions for Science Since 1875Since 1875
Move A Mountain One Peak At A Move A Mountain One Peak At A TimeTime
Do Both With A GradientDo Both With A Gradient
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SHIMADZUSHIMADZUSolutions for ScienceSolutions for Science Since 1875Since 1875
Move A Mountain One Peak At A Move A Mountain One Peak At A TimeTime
Leave Room For Uninvited Leave Room For Uninvited GuestsGuests
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SHIMADZUSHIMADZUSolutions for ScienceSolutions for Science Since 1875Since 1875
When The Mountain Won’t MoveWhen The Mountain Won’t Move
A Stationary Phase For Any A Stationary Phase For Any OccasionOccasion
•C18C18
•C8C8
•C4C4
•PhenylPhenyl
•Get NormalGet Normal
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SHIMADZUSHIMADZUSolutions for ScienceSolutions for Science Since 1875Since 1875
OptimizeOptimize
How Good Is Good EnoughHow Good Is Good Enough
•Short Is GoodShort Is Good
•ResolutionResolution
Leave A Little Wiggle RoomLeave A Little Wiggle Room
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SHIMADZUSHIMADZUSolutions for ScienceSolutions for Science Since 1875Since 1875
The Chromatographer’s TriangleThe Chromatographer’s Triangle
SpeedSpeed
ResolutionResolution CapacityCapacity
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SHIMADZUSHIMADZUSolutions for ScienceSolutions for Science Since 1875Since 1875
Do My Peaks Look Fat?Do My Peaks Look Fat?
Free Mean PathsFree Mean Paths
Laminar FlowLaminar Flow
VoidsVoids
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SHIMADZUSHIMADZUSolutions for ScienceSolutions for Science Since 1875Since 1875
Hot PeaksHot Peaks
1ºC ~ 1% RT1ºC ~ 1% RT
Different For Every CompoundDifferent For Every Compound
Room Temp May Not Be The BestRoom Temp May Not Be The Best
Trial and ErrorTrial and Error
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SHIMADZUSHIMADZUSolutions for ScienceSolutions for Science Since 1875Since 1875
Straighten The PeaksStraighten The Peaks
Competing ChromatographyCompeting Chromatography•Size ExclusionSize Exclusion•ModifiersModifiers
OverloadingOverloading
Resonance StructuresResonance Structures
Methanol vs. AcetonitrileMethanol vs. Acetonitrile
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SHIMADZUSHIMADZUSolutions for ScienceSolutions for Science Since 1875Since 1875
DetectionDetection
SpecificitySpecificity
SensitivitySensitivity
Post And Pre Column DerivativesPost And Pre Column Derivatives
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SHIMADZUSHIMADZUSolutions for ScienceSolutions for Science Since 1875Since 1875
DetectionDetection
UV/VisUV/Vis•254nm254nm
RIDRID
FluorescenceFluorescence
ConductivityConductivity
ELSDELSD
MSMS
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SHIMADZUSHIMADZUSolutions for ScienceSolutions for Science Since 1875Since 1875
CalibrationCalibration
How Many PointsHow Many Points
Pick An EquationPick An Equation
SpacingSpacing
UncertaintyUncertainty
How Long Is The Curve ValidHow Long Is The Curve Valid
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SHIMADZUSHIMADZUSolutions for ScienceSolutions for Science Since 1875Since 1875
Set’em UpSet’em Up
How Much SampleHow Much Sample•100 mg100 mg
Range Range •10 – 100 ppm10 – 100 ppm
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SHIMADZUSHIMADZUSolutions for ScienceSolutions for Science Since 1875Since 1875
ValidationValidation
Sample / Standard StabilitySample / Standard Stability
AccuracyAccuracy
PrecisionPrecision
LODLOD
LOQLOQ
ToleranceTolerance•ColumnColumn•MatrixMatrix
High ConcentrationHigh Concentration
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SHIMADZUSHIMADZUSolutions for ScienceSolutions for Science Since 1875Since 1875
Doing It BetterDoing It Better
Smaller ColumnsSmaller Columns
Smaller ParticlesSmaller Particles•A=A=ππr²r²
Capillary HPLCCapillary HPLC
UPLCUPLC
This product was funded by a grant awarded under the President’s High Growth Job Training Initiative as implemented by the U.S. Department of Labor’s Employment & Training Administration. The information contained in this product was created by a grantee organization and does not necessarily reflect the official position of the U.S. Department of Labor. All references to non-governmental companies or organizations, their services, products, or resources are offered for informational purposes and should not be construed as an endorsement by the Department of Labor. This product is copyrighted by the institution that created it and is intended for individual organizational, non-commercial use only.