HPLC-HRMS: The New Paradigm for New Drug Discovery Assays
Walter Korfmacher Sr. Director, Bioanalytical, Medpace Bioanalytical Laboratories
A One-Step Procedure for Simultaneous Protein Precipitation and Phospholipid Removal Prior to LC-MS Analysis
Xiaoning Lu Supelco Division, Sigma-Aldrich
Antibodies, Proteins and Peptides - Approach to Make Complex Separations Simpler With More Resolution and Speed
Tim Rice Agilent Technologies
Lipids, Proteomics and Non-chromophoric Compounds Using HPLC-ELSD
Tim Reyburn Ticoscen, Inc.
Bioanalytical Method Development Strategy for Therapeutic Peptides
Mary Lame Waters Corporation
A Case Study on What Information HPSEC / MALS / RI Analysis Tells Us
Jian He Merck & Co, Inc.
The Bioanalysis of Large Molecules: An Overview
Shannon Chilewski Bristol-Myers Squibb Co.
Abstracts
HPLC-HRMS: The New Paradigm for New Drug Discovery Assays
Walter Korfmacher Sr. Director, Bioanalytical, Medpace Bioanalytical Laboratories
There is a continuing demand for higher throughput assays for compounds in the drug discovery stage. Specifically, there is a need for techniques that can be used to increase sample throughput in the analysis of drug discovery samples obtained from various absorption, distribution, metabolism, excretion (ADME) and pharmacokinetic (PK) studies. Currently, most of these assays rely on LC-MS/MS for the quantitative analysis step. In the future, most of these assays will likely be performed using high resolution mass spectrometry (HRMS). The focus of the presentation will be to provide a view of how this paradigm shift will occur.
A One-Step Procedure for Simultaneous Protein Precipitation and Phospholipid Removal Prior to LC-MS Analysis
Xiaoning Lu Supelco Division, Sigma-Aldrich
Endogenous proteins and phospholipids are two of the major causes of matrix effects in LC-MS/MS analyses. The concentrations of these proteins and phospholipids are often high and variable. Liquid-liquid extraction and solid phase extraction are two commonly used procedures for cleanup of such proteins and phospholipids during the preparation of biological samples. Both of the procedures, however, involve multiple steps and tend to be time consuming and labor-intensive. While the use of protein precipitation affords a faster method for sample preparation, this technique is not effective in removing phospholipids. The presence of phospholipids in the sample extract can significantly impact the quality of the data derived from using this technique.
We have developed a one-step procedure for simultaneous protein precipitation and phospholipid removal from biological matrices. The procedure involves the use of zirconia-modified silica sorbents to selectively remove phospholipids from biological samples. And the proteins are precipitated with organic solvents. Protein precipitation and phospholipid removal are realized in a single step. Our results indicate that phospholipids in various biological fluids such as plasma and serum were effectively eliminated and ion suppression of the MS signals was greatly reduced. We will demonstrate with several application examples that our procedure leads to higher sensitivity and improved data quality.
Antibodies, Proteins and Peptides - Approach to Make Complex Separations Simpler With More Resolution and Speed
Tim Rice Agilent Technologies
A quick look at the most recent new drug filing reveals how much interest there in biologically derived molecules as therapeutic agents. The majority of the filings are for New Biological Entities rather than New Chemical Entities with the largest NBE group being Antibodies. As we move in this direction regulatory agencies are asking for more and more data about the make up of these molecules, degradation products and improperly made versions of the molecule. The need for this information about these complex systems makes efficient separations very important to speeding up development as well as monitoring production of new NBEs.
This presentation will look at new separation products designed to improve resolution and reproducibility in SEC, IEX, and RP separations. We will explore:
Improved resolution in SEC separations by utilizing 3u particles and simpler mobile phases - better aggregate vs monomer resolution
Improved resolution for weak and strong IEX using 5, 3.5, and sub-two micron particles. High efficiency is finally here!
Improved resolution and drastically reduced assay time for whole protein and peptide map separations
The presentation will review the mechanism of these separation techniques and show how to optimize separation conditions for the best resolution in the least amount of time with HPLC, UHPLC and LC/MS instrumentation. There might be a few surprises!
Lipids, Proteomics and Non-chromophoric Compounds Using HPLC-ELSD
Tim Reyburn Ticoscen, Inc.
Bioanalytical Method Development Strategy for Therapeutic Peptides
Mary Lame Waters Corporation
A Case Study on What Information HPSEC / MALS / RI Analysis Tells Us
Jian He Merck & Co, Inc.
The Bioanalysis of Large Molecules: An Overview
Shannon Chilewski Bristol-Myers Squibb Co.
