HPLC Application Collection


Our chromatography experts have compiled an extensive collection of analytical HPLC and UHPLC application notes and methods. Whether you are developing new methods or optimizing existing processes, you can take advantage of this wealth of information.

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For an overview of these notes and methods, see below:
  • Chinese Pharmacopoeia 2015 Monograph Methods

    Solutions for regulated Pharmaceutical Instrumental Analysis (2016-2)

    Merck:/Freestyle/LE-Lab-Essentials/Chromatography/LE-Chinese Pharmacopoeia MM-06082016.jpg

    This application compilation focuses on the new 2015 Chinese pharmacopeia (ChP 2015) edition that officially replaced the previous ChP 2010 on December 1, 2015. 

    In all examples we have used modern particle packed Purospher® STAR (RP-18 endcapped and Phenyl) HPLC columns and monolithic Chromolith® HighResolution RP-18 endcapped columns, Lichrosolv® high purity solvents (isocratic grade) and EMSURE® high purity reagents.
    The monograph methods have been selected from both chemical medicines as well as Traditional Chinese Medicines (TCM’s). You will learn about Chromatographic aspects of testing, implementation, and validation of monograph methods from the new ChP 2015.

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  • How to use Monolithic Columns in Regulated Pharmaceutical Analysis

    Scaling of Monograph Methods (2015-4)

    How to use Monolithic Columns in Regulated Pharmaceutical Analysis

    This compilation focus on United States pharmacopeia (USP) monograph methods, and how method transfer of isocratic HPLC methods is carried out from particulate to monolithic columns following the updated USP general chapter 621.

    In all examples we have used modern particle packed HPLC columns and monolithic Chromolith® HighResolution RP-18 endcapped columns, Lichrosolv® high purity solvents (isocratic grade) and EMSURE® high purity reagents.

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  • Complete Solutions for Dissolution Testing

    Monograph Methods for Tablets and Capsules (2015-2)

    In this compilation you will find complete solutions for dissolution testing of a few selected formulated small molecule drugs. The highlighted methods follow the current USP37-NF32 monographs, and are all compliant with USP38-NF33 and the given system suitability requirements.

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  • Pharmacopoeia Monograph Methods

    HPLC and UHPLC Columns for Regulated Drug Analysis

    This application compilation introduces the new guidelines in general chapter 621 of USP and details how this affects scaling of methods as well as what considerations must be taken. Both USP and EP methods are included in this compilation both under HPLC and UHPLC conditions. In addition, there is new information about how to perform validation of a monograph method to meet regulatory demands.

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  • U.S. Pharmacopeia Methods

    Regulated Methods for HPLC with Purospher® STAR Columns

    The process of validating a new analytical procedure for compendial usage is addressed in United States Pharmacopeia (USP) general Chapter 1225 "Validation of Compendial Procedures". However, even with a fully validated procedure, scientists may not have assurance that the procedure is suitable for use with a specific ingredient or product in a specific laboratory with specific personnel, equipment, consumables and reagents. USP, therefore, developed chapter 1226 in response to the industry's request to provide instructions for verifying compendial procedures in specific situations.

    In this compilation, we have addressed USP's proposed new general chapter 1226 "Verification of Compendial Procedures", intended to fill the gap in the proper usage of compendial procedures by outlining a process for verifying their suitability.

    The role of HPLC columns is of immense importance to meet system suitability test (SST) criteria in compendial methods. This is illustrated in this compilation by analysing important drugs on Purospher® STAR as per pending USP monographs.

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  • Small Molecule Bioanalysis by LC-MS and LC-MS/MS

    Analysis of Endogenous and Exogenous Compounds in Biological Samples

    Food safety and food quality control chromatography application compilations using monolith HPLC Columns

    Many parties require methods for accurate quantification of drugs and endogenous substances in biological samples to provide qualitative and quantitative measures of the active drug and/or its metabolite(s) for the purpose of pharmacokinetics, toxic kinetics, bioequivalence and exposure–response (pharmacokinetics/pharmacodynamic studies).

    On the following pages, we have included a number of applications with relevance to clinical bioanalysis, focusing on a number of molecules either being endogenous (naturally occurring) or molecules being administered to humans (exogenous).

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  • Polar Hydrophilic Compounds in Pharmaceutical Analysis – Part II

    Monograph Modernization Using SeQuant® ZIC®-cHILIC and ZIC®-pHILIC HPLC Methods

    Food safety and food quality control chromatography application compilations using monolith HPLC Columns

    Reversed phase (RP) HPLC has been the backbone of pharmaceutical analysis for more than 25 years. It has been particularly successful for Active Pharmaceutical Ingredients (APIs) since most APIs have traditionally followed the Lipinski rule of five and been relatively hydrophobic molecules. Roughly 10% of all marketed drugs, however, do not follow Lipinski's rule. RP chromatography has been used most often for these APIs. The poor suitability of RP for polar APIs have, however, been acknowledged by the USP, and most of the pharmaceuticals on the 'Monograph Modernization List' are polar molecules.

    Application examples are presented here, with full experimental details, to demonstrate the usefulness of HILIC for pharmaceutical quality control and impurity profiling methods.

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  • Food Safety and Food Quality Control - Dirty Sample Analysis

    Dirty Sample Analysis Even Faster with Monolith Columns Monolith Chromatography Columns – The Ideal Tool for Dirty Sample Analysis

    Merck:/Freestyle/LE-Lab-Essentials/Chromatography/LE-Food safety and food quality control-150x212-02052014.jpg

    With the current focus on food issues, there is an increasing need for new analytical methods able to cope with large numbers of analytes in complex matrices. These new analytical assays must provide sensitivity, robustness and high resolution within an acceptable analysis time. Many modern approaches in HPLC analysis enable the reduction of analysis time without compromising resolution and/or separation efficiency: the use of monolith columns, liquid chromatography at high temperatures, and ultrahigh pressure liquid chromatography (UHPLC) either using small particle packed columns or superficially porous particle columns. A viable approach to meet these demands is to develop the HPLC assays using Chromolith® columns since monolithic columns are more matrix tolerant than particles packed columns.

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  • UHPLC - HIgh-Throughput Separations Applications

    Guidelines for High-Throughput Separations
    Applications for UHPLC with Purospher® STAR and Chromolith® Columns

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    Fast and ultra-fast separations have become particularly important due to the need for high sample throughput and higher productivity in daily lab work. Using UHPLC methods with short columns, narrow inner diameters and small particles sizes, it is possible to speed up analyses up to ten-fold.

    The choice of UHPLC columns depends on the sample being analyzed. To ensure that you have the most appropriate solution for your application, Merck offers two different column materials for UHPLC separations: Relatively clean samples are optimally separated on columns with narrow inner diameters and small particle sizes like Purospher® STAR UHPLC columns. Samples with more complex matrices usually require intensive sample preparation to avoid column contamination or clogging. In such cases, Chromolith® columns are the best choice, since they enable fast separations with minimum sample preparation – saving time and reducing costs.

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  • Food Safety and Food Quality Control - Analysis of Polar Compounds in Complex Samples

    Analysis of Polar Compounds in Complex Samples

    Applications for HPLC and LC-MS with SeQuant® ZIC®-HILIC columns

    Merck:/Freestyle/LE-Lab-Essentials/Chromatography/LE-Analysis of polar compounds cover-250x206-02052014.jpg

    This compilation covers Food Safety and Food Quality Control and focuses on the Analysis of Polar Compounds in Complex Samples (for example, melamine in milk powder formulations). The quality of our daily food is of concern for producers, consumers, and controlling authorities. The globalization process changes the demands and requirements almost constantly. Merck offers analytical solutions and prompt technical support.

    The initial part of this compilation covers trends and news regarding food safety legislation and has been compiled by the Merck Regulatory Department.

    The subsequent sections give an account as to why the analysis of polar compounds is important for food and beverage control. Analytical challenges are highlighted, and an explanation is given about why Hydrophilic Interaction Liquid Chromatography (HILIC) in general, and SeQuant® ZIC®-HILIC columns in particular, is the ideal tool for analysis of polar compounds in food and other complex samples. Several methods are presented, with complete workflows and, as you will see, Merck offers everything but the instrument to successfully implement these methods in your laboratory.

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