Liquid Chromatography (LC/HPLC)

In any field there are often "misconceptions" or "myths" that are perpetuated and passed on to the next generation. These myths are often driven by a lack of understanding by practitioners of the real issues. In this instalment of "Column Watch", the 10 most popular myths around high performance liquid chromatography (HPLC) column technology will be demystified by discussing the issues at hand. Among some of the popular myths that will be dispelled are that "All C18 (L1) columns are the same", "You can't reverse an HPLC column", "High temperature always leads to better separations" and "The higher the carbon load the better the reversed-phase column".

This article describes and compares a number of approaches to increase the speed of liquid chromatographic separations. On a standard LC column, a gain of a factor two in the run time (from 10 to 5 minutes) was achieved by increasing the flow-rate two-fold. On a monolithic column, a column operated at high temperature (120 ?C) and a short column, flow-rates could be increased typically five-fold, resulting in run times in the order of 2 minutes. This was accompanied by a sometimes considerable loss in separation efficiency. A combination of a very short run time and unaffected separation efficiency was realized on a UPLC system, designed for use at higher pressure. By working at approximately 800 bar, the analytes could be well separated within 30 seconds.

This month's instalment of "LC Troubleshooting" presents two examples of sample degradation inside the liquid chromatography (LC) column. Depending upon the type of samples you analyse, sample degradation might or might not be a problem that you encounter regularly. However, most of us run a sufficiently wide variety of sample types over our careers that we will probably run into some samples that do not behave as expected.

Emerging as a complementary analytical tool to LC-MS, LC-ICP-MS brings added value to the determination of inorganic element containing molecules of biological importance.

Chromatographers worldwide suffer from the same problems.

The environmental market peaked about fifteen years ago when the United States government passed a large body of new environmental laws and strengthened existing ones, dramatically increasing the number of mandated tests. As a result, the analytical instruments market received a boost in new instrument sales. While growth from the environmental market has slowed considerably since, the total market is still quite significant.

The analysis of peptides and additives using a new HPLC innovative column technology, Pathfinder, was shown. For the analysis of flavours in lemon juice GC–MS with a new MS library, FFNSC, with linear retention indices was used.

Soft drink formulas often include preservatives, artificial sweeteners, flavours or caffeine in their list of ingredients. Using the Acclaim OA column, as many as eight common additives may be determined in a single run. Many of these additives are hydrophilic organic acids for which this column was designed. Notably, benzoate and sorbate, which do not resolve on C18 columns at low pH, are fully separated.

Column lifetime is a more and more important issue when developing an analytical method for HPLC. Besides sample treatment, column cleaning and storage, operational parameters of the analytical method will have an influence on column lifetime. This question may not always be addressed early enough in the methods development process.

The authors describe a study that was conducted to evaluate the correlation between the slope of the response line and the response factor of an ELSD that are ordinarily regarded as independent numerical coefficients.

Sometimes it is easy to ignore the fittings and tubing that are used to connect various parts of the liquid chromatography (LC) system. After all, it's the pump, injector, column and detector that do all the work - right? Well, yes and no. It is possible to compromise an otherwise excellent separation by the improper use of fittings, but with reasonable care, you should not have problems with most applications. This month's instalment of "LC Troubleshooting" takes a look at the important components that are used to connect various parts of the LC system and how to use them wisely.

In a good laboratory practice (GLP) environment, data-handling software cannot be used until it is validated. This even applies to the most simple program that performs calculations or stores data. A detailed documentation of the set-up and the performance of the software - called software validation - is required. The development and validation requirements are described in this article and illustrated with a software for robustness testing (SRT), which guides the user step-by-step through the experimental set-up and interpretation of robustness tests. This software was developed in an Excel (Windows XP) environment and is used as part of method validation in laboratories that require compliance with GLP and 21 CFR Part 11. The software was subjected to software validation regulations and is compliant with electronic records and signature rules (21 CFR Part 11) as it creates, delivers and stores electronic data. The validation tests are based on the computerized system validation (CSV) -..

This article assesses the advantages of including certified reference materials in collaborative method validation studies in food analysis. A recently conducted collaborative trial on the determination of acrylamide in bakery and potato products is described to illustrate this.

Since the first Pittsburgh Conference on Analytical Chemistry and Applied Spectroscopy was held in 1950, Pittcon, as it is called today, has become a key meeting place where renowned scientists present their pioneering research and developers showcase the latest instruments that make many of those research scientific breakthroughs possible. This two-part article series will present a historical perspective on just a few of the significant scientific advancements that were discussed and demonstrated at Pittcon over the years. Part I will focus on new instruments and techniques that were presented at Pittcon from 1950 to 1980, and Part II will discuss a few of the many notable scientists and their discoveries that have been enabled by advances in mass spectrometry, capillary electrophoresis, and other instrumental techniques since 1980.

Serum protein profiling using mass spectrometry (MS) is one of the most promising approaches for biomarker identification.

What do you do when the sample changes inside the column?

Measurement of chiral purity is a necessary means of quality control for drug substances that exhibit chiral centers. This article describes a simple and practical approach to setting up system suitability and validation for chiral purity assays.

In any field there are often "misconceptions" or "myths" that are perpetuated and passed on to the next generation.

The laboratory life science and analytical instrument industry is comprised of a large, diverse group of organizations that manufacture and distribute their products worldwide. However, overall industry sales are concentrated among a small set of companies. Approximately 950 firms are included in the industry, but only about 100 companies have sales above $50 million.

Software traditionally used to control and process data in analytical instruments is now being expanded to include functions such as automated system monitoring with maintenance feedback alerts and interactive diagnostic routines that enable users to quickly track and resolve the source of many system problems. As a back up to these functions, a second layer of expert support is being made available by some instrument vendors to address more complex servicing issues. For larger or more elaborate instrument installations, monitoring and diagnostic software functions can be integrated with remote servicing expertise in a networked configuration to provide continuous proactive instrument maintenance. The benefits of these innovations include lower and more predictable operating costs, reduced system downtime, and more efficient utilization of the instrument user's time.