The Application Notebook-06-01-2008

June 01, 2008

Short analyses time and high resolution are in great demand from R&D and QC departments within the pharmaceutical industry. Sub-two micron ODS reversed phase columns have recently been introduced to meet these requirements, but these columns require an ultra-high pressure HPLC system to achieve optimum performance. TSK-GEL ODS-140HTP, 2.3mm columns from Tosoh Bioscience have been developed to offer a combination of short analyses time and high resolution separations that can be run at modest pressures, making these columns compatible with conventional HPLC instrumentation. The polylayer bonding chemistry of these columns results in highly efficient and physically stable columns when operated at high linear velocities. In addition, TSK-GEL ODS-140HTP, 2.3mm columns can be efficiently operated at pressures not exceeding 9000psi in UPLC® and other ultra-high pressure HPLC systems, as well as in traditional HPLC systems.

June 01, 2008

Pharmaceutical actives and impurities often contain sulfur in the form of a sulfone or sulfoxide group. Both groups have dipole moments, adding a hydrophilic character to compounds containing these functional groups. The analysis of hydrophilic compounds on traditional alkyl columns (e.g. C18) can be problematic, since alkyl columns depend on hydrophobic interactions for retention. Since the sulfone and sulfoxide groups contain π-π bonds, the biphenyl column's ability to undergo π-π interactions makes it an excellent choice when increased retention or selectivity of compounds containing these groups is desired. To demonstrate the selectivity of the biphenyl phase towards aromatic compounds containing sulfur groups, a set of target compounds was selected and analyzed on C18, phenyl, and biphenyl columns.

June 01, 2008

Synthetic fused silica capillary tubing has proven to be a vital component in many separation science techniques. Recently, it has expanded to serve an array of scientific markets. In most cases, an understanding of the internal surface chemistry is of fundamental importance.

June 01, 2008

Mercury pollution mainly originates from industrial activities such as chlorine production, garbage incineration and above all coal-fueled power generation. The US Environmental Protection Agency (US EPA) considers mercury as highly toxic with a pronounced accumulative and persistent character.

June 01, 2008

MEPS uses a barrel insert and needle (BIN) device to reduce Solid-Phase Extraction (SPE) to a micro-scale suitable for small volume samples and for the online adaptation of conventional SPE techniques. Because the SPE cartridge (BIN) is incorporated into the needle assembly of a gas-tight syringe, MEPS is also a simple field-portable SPE device that may be operated manually without need for sampling pumps or, alternatively, may be incorporated into robotic samplers. MEPS devices are of glass and stainless steel construction allowing them to be fully immersed for sampling at depth or, alternatively, used at needle depth to avoid perturbing the stream from which the sample was drawn. An extension pole allowed MEPS to be used to sample back along pipes or down inspection vents. When sampling from drainage pits and open sumps, there was minimal requirement to remove grates to gain access. An extension pole also allowed sampling from outflows that were offensive and could be readily adapted for safe sampling of..

June 01, 2008

his application note describes the extraction of a variety of basic drugs from plasma using EVOLUTE CX mixed-mode resin-based SPE. The analyte suite includes basic drugs with wide ranging pKa and logP values.

June 01, 2008

There are many misconceptions about what it means to perform fast gas chromatography (GC) and what the term fast GC implies. Fast GC is often associated with the use of hydrogen as a carrier gas and, although this is certainly a good approach, it is not always necessary to shorten the analysis time. A second misconception is that changing column dimension results in time-consuming method development. Using high-efficiency GC columns can greatly reduce the analysis time and when coupled with the method translation software, the time spent on method development can be greatly minimized.

June 01, 2008

Viscotek has been a strong advocate of good chromatography as a prerequisite for GPC (Gel Permeation Chromatography) data accuracy. Our recent work in application development has been driven by extremely difficult samples from industrial, biopharmaceutical, and academic sources. These samples present challenges ranging from sample solubility, column adsorption as well as detection issues. This report will attempt to highlight a new approach that could be very helpful in certain advanced GPC applications.

June 01, 2008

EPA Method 525.2 describes the procedure to determine low ppb levels of semi-volatile organic material in drinking water using solid phase extraction (SPE) or liquid-solid extraction (LSE) techniques. The City of Fort Worth, Water Department implemented an automated SPE process for the analysis of semi-volatiles by EPA Method 525.2, using the Atlantic "Certified for Automation" SPE Disk for EPA Method 525.2. Ethyl acetate, methanol, and water were used to condition the Atlantic disk prior to the extraction step. The extraction solvents used were a 1:1 mixture of methylene chloride and ethyle acetate. Extracts were then analyzed by GC–MS using a splitless injection technique.

June 01, 2008

In pharmaceutical development, it is important to analyze small molecules or their metabolites in biological fluids. For this purpose, the analytical methods such as sample pretreatment, 2D-LC and LC–MS have been developed. However there are still problems of resolution and protein adsorption. As a result, satisfying analytical results have not always been achieved.

June 01, 2008

An effective UHPLC-MS method for high throughput separation, identification and quantitation of pseudoephedrine was developed on a Hypersil GOLD™ PFP 1.9 µm, 2.1 x 100 mm column.

June 01, 2008

With the forthcoming USP monograph , many laboratories are in the process of reexamining their high performance liquid chromatography (HPLC) instrumentation qualification practices. This article demystifies the qualification procedures and proposes a well designed, easy and simple set of experiments upon which to establish internal standard operating procedures (SOPs) for the complete qualification of HPLC instruments. A key concept is the development of a consistent test system, comprised of premade test solutions, a prequalified HPLC column, standardized protocols, and validated software that can be prepared in-house or purchased commercially as a kit. This system can be applied to any HPLC system worldwide, to produce comparable test results under uniform conditions. The test system is designed to be rapid, with a comprehensive performance qualification being completed in about 2 h for isocratic, and 3 h for quaternary gradient systems.

June 01, 2008

Two of the most commonly occurring unpleasant odor-causing compounds in drinking water are geosmin and 2-methylisoborneol (MIB) (Figure 1). Geosmin is produced primarily by blue-green algae (cyanobacteria) and actinomycete bacteria, and MIB is produced by certain species of cyanobacteria, primarily Oscillatoria. Many environmental laboratories are required to detect these compounds as low as 1–3 ppt concentration and measure quantitatively at 5 ppt.

June 01, 2008

Dionex has developed a new standard for flow-through solvent extraction which allows accelerated solvent extraction (ASE®) of matrices that have undergone acid or alkaline pretreatment or digestion. The new ASE 150 and ASE 350 systems use extraction cells and post-cell solvent pathways constructed of Dionium™ material. This pH-hardened substance resists corrosion under acidic or alkaline conditions used in standard pretreatments, widening the scope of ASE applications and significantly expanding its capabilities.

June 01, 2008

Several common birth control formulations contain both drospirenone and ethinyl estradiol. A highly selective and sensitive analytical method for the analysis of drospirenone in human plasma has been developed for use in bioequivalence studies. The solid-phase extraction (SPE) and UPLC®–MS–MS methodologies are described as well as performance against validation parameters.

June 01, 2008

The use of mobile phase pH to control analyte ionization states (pH-LCâ„¢) in reversed phase HPLC separations is a highly effective way to change selectivity. The ionized species of an analyte is shown to have higher polarity (less hydrophobicity) than the neutral species, which results in a loss of expected retention for that analyte. This can be attributed to less interaction with the hydrophobic stationary phase and greater affinity with the aqueous portion of the mobile phase. Ionized species also participate in ionic interactions with exposed and activated silanols, which impact peak shape and reproducibility.

June 01, 2008

The purge-and-trap (P&T) technique for analysis of volatile organic compounds (VOCs) was pioneered in the 1970s at the United States Environmental Protection Agency (USEPA) research laboratory in Cincinnati. Many of the operational parameters developed during this time period are still included in USEPA methods. While these parameters still produce good analytical results, they do not take advantage of advances in instrumentation that enable analysis of emerging contaminants such as fuel oxygenates, and increased sample throughput.

June 01, 2008

H-SRM provides excellent selectivity for accurate identification and quantification of pesticides in matrix, demonstrating high productivity for effective control at international maximum residue levels (MRLs).

June 01, 2008

Bradykinin, a 9 amino acid peptide, is a physiologically and pharmacologically active peptide of the kinin group of proteins, which is used in the development of antagonists and therapies for hereditary angioedema. In this application, 50 mg of crude bradykinin, synthesised on a StratoSpheresâ„¢ PL-Rink resin, is purified using an HPLC method that can be scaled from the laboratory through to full production.

June 01, 2008

The United States Geological Survey (USGS) has found pharmaceuticals and personal-care products (PPCPs) containing known or suspected endocrine-disruptors in U.S. rivers. As such, it is important to use adequate techniques to help identify these compounds and possible metabolites.

June 01, 2008

With recent advances in technology and the use of ultra high pressure chromatography systems becoming more commonplace, it is of use to compare reproducibility of retention time and peak area of UHPLC vs HPLC.

June 01, 2008

Azo dyes are used widely in the manufacture of various consumer goods such as leather, textiles, plastics, paper, hair care products, and cosmetics. On September 11, 2003, the European Union enacted European Parliament Directive 2002/61/EC, prohibiting the manufacture and sale of consumer goods containing specified azo dyes (1). The azo dyes of concern are those that can be reduced to aromatic amines. There are 22 aromatic amines classified as carcinogenic or potentially carcinogenic to humans.

June 01, 2008

Size exclusion chromatography is a powerful tool for the characterization of molecules differing in size and molar mass. It is widely used and well known for the determination of molar mass distributions and molar mass averages. For membranes GPC-SEC is a useful characterization method as it can measure the membrane characteristic pore size distribution, average pore size and molecular cut-off significantly faster than other methods.

June 01, 2008

Aminoglycoside antibiotics are among those commonly used in animal feeds to manage intestinal microorganisms. The beneficial effects include improved growth and generally healthier animal populations. Use of antibiotics creates a demand for analytical procedures to verify concentrations in pre-mixes and feeds and in some instances for residue analysis in animal products.

June 01, 2008

Mycotoxins, toxic secondary metabolites of several fungal species, represent food safety issues of high concern. Deoxynivalenol, the most abundant trichothecene mycotoxin, can be found worldwide as a contaminant of wheat, barley, maize and other cereals (1,2). The transmission of deoxynivalenol from barley into beer has been reported in several studies (3,4). Therefore, its levels should be controlled.

June 01, 2008

A new phenylhexyl bonded phase provides an alternative selectivity for optimizing HPLC method. This also applies when different types of phenyl bonded phases are compared.

June 01, 2008

Supercritical fluid chromatography (SFC) is a normal phase separation technique. In SFC, supercritical CO2 in combination with one or more polar organic solvents, most commonly alcohols, is used as the mobile phase. Owing to the lack of intermolecular interactions, supercritical fluid typically possesses lower viscosity and higher diffusivity than those solvents used in traditional high performance liquid chromatography (HPLC). This allows for higher flow rates, faster analyses, and the use of longer columns for higher chromatographic efficiencies. Initially deemed a niche chromatographic technique for chiral separation, the horizon of SFC applications has rapidly expanded to include achiral analyses of natural products, biodiesel, oligomer, pesticides/herbicides, and peptides. This is due, in part, to the improvements in detection choices and performances for SFC. Evaporative light scattering detectors (ELSDs) coupled with SFC have found wide use in many pharmaceutical and chemical laboratories (1).

June 01, 2008

This application note describes a fast and sensitive LC-MS method using a Hypersil GOLDâ„¢ column on a Thermo Scientific LC-MS system for the quantitative analysis of two widespread PFCs, perfluorooctanoic acid (PFOA) and perfluorooctansulfonate (PFOS).

June 01, 2008

Supercritical Fluid Chromatography (SFC) is quickly becoming a popular choice by chromatographers for analytical and preparative separations.

June 01, 2008

In the last few decades, the novel functions of polysaccharides have provided a major impetus for increasing scientific attention. Among the most promising aspects are their immunomodulatory and antitumor effects, thickening agents and stabilizer effects.

June 01, 2008

Glucosamine (GlcN) is a major structural component in the biosynthesis of glycosaminoglycans, compounds involved in normal joint function. Use of GlcN as a dietary supplement in the management of osteoarthritis has attracted considerable attention; it is one of the five nonvitamin, nonmineral supplements most frequently used by US adults (1). The US FDA currently regulates dietary supplements to ensure that they are produced under cGMP (2).

June 01, 2008

Urea is commonly used in protein purification, including large-scale purification of recombinant proteins for commercial purposes, and in recombinant protein manufacturing to denature and solubilize proteins (1). In aqueous solutions, urea degrades to cyanate and ammonium, with the maximum degradation rate occurring at neutral pHs commonly used in biological buffers (2). Cyanate is problematic in urea solutions because it carbamylates proteins, which causes unwanted modifications that can alter the protein's stability, function, and efficiency. Therefore, an accurate, sensitive method for determining cyanate in urea-containing buffers is required.

June 01, 2008

There has been an increasing interest in the presence and availability of compounds in plant materials that may possess bioactive properties, in particular, antioxidant activity. Some of these compounds have been attributed to possess anticancer, antiaging, and antimutagenic properties as well as other health benefits (1). The types of plants that have been investigated cover a vast range from common foodstuffs to regional or exotic materials. Plant parts under study have included portions that are traditionally known to be edible, as well as sections that are considered "waste" or used for animal forage. Because most screening techniques involve lengthy separations, high throughput HPLC methods are desirable.

June 01, 2008

The analysis of polar compounds in support of clinical and preclinical pharmacokinetic studies requires an analytical methodology capable of achieving ultra-low detection and quantification limits. The high sensitivity afforded by coupling HPLC with tandem mass spectrometry (MS-MS) has made it the technique of choice in this environment, but it is subject to the following limitations when reversed phase liquid chromatography (RPLC) is used:

June 01, 2008

The Zebron ZB-Bioethanol GC column reduces fuel ethanol analysis time to less than 8 min, while providing complete resolution of methanol and ethanol from all denaturant components. This newly developed GC column is suitable for analysis following ASTM Method D 5501 and Europe prEN 15376.

June 01, 2008

A method for trace odor components, isopropyl-methoxypyrazine (IPMP), isobutyl-methoxypyrazine (IBMP), methylisoborneol (MIB), and geosmin in drinking water involving the use of solid phase microextraction (SPME) and the SLB-5ms capillary column using gas chromatography/mass spectrometry (GC–MS).

June 01, 2008

Amylose is an occasionally-branched biopolymer and, together with amylopectin, the hyper-branched component, a constituent of starch. Determination of branching in amylopectin on the basis of amyloses may be performed with the help of synthetic amyloses. Synthetic amyloses from enzymatic (phosphorolytic) reaction were checked for their linearity.

June 01, 2008

Like any other segment of the world economy, current events and the realities of the world around us go a long way toward shaping the direction of chromatographic research and development, and consequently, the financial trends that have a direct impact on the industry in which we all live and work. In this sense, the LCGC Application Notebook has become something of a market barometer over the years.

June 01, 2008

In March 2007, several North American manufacturers of pet food voluntarily issued nationwide recall notices for some of their products that were reportedly associated with renal failure in pets. The raw material wheat gluten, used to manufacture the pet food, was imported from China and was identified as the source of contamination.

June 01, 2008

The United States Pharmacopeia (USP) has implemented a revised method for the determination of residual solvents, chapter 467; this revision has brought the methodology of USP 467 into close alignment with European Pharmacopeia (EP) method 2.4.24. The USP and EP determination of class 1 and class 2 residual solvents is performed with static headspace (HS) sample introduction and gas chromatography (GC) with flame ionization detection (FID); class 3 has flexibility in the technique, however, it is often included in the HS analysis.

June 01, 2008

Urea and allantoin are added to cosmetic products for skin protection and regeneration, especially for the treatment of dry skin, and analyzed for QC purposes. As polar compounds, they are not ideal for reversed-phase HPLC separations. Neutral hydrophilic compounds like urea and allantoin are best analyzed by hydrophilic interaction chromatography (HILIC). Traditional HILIC columns use silica modified with a hydrophilic group such as diol or cyano. Analytes are adsorbed and subsequently eluted with mobile phases containing high percentages of organic solvent (>75%).

June 01, 2008

Gel permeation chromatography (GPC) has been used as an effective cleanup procedure for removing high molecular weight interfering molecules such as lipids, pigments, proteins, and polymers before GC or HPLC analysis. The GPC cleanup method has been extensively documented (1–3) and is also recommended in US EPA SW-846 Method 3640A. To demonstrate the efficacy of this method to extract polar and nonpolar substances by using the KNAUER Smartline GPC Cleanup Unit 6500, olive oil samples were investigated by spiking these with different types of organic pollutants, including PAHs, phthalates, phenols, and triazine.

June 01, 2008

Use imaging mass spectrometry to determine the biodistribution of erlotinib (Tarceva®) in pancreatic tumors from treated and untreated mice.

June 01, 2008

The QC for the pharmaceutical industry is based on methods developed on a variety of columns. At Orochem, we have attempted to address method development utilizing unique chemistries such as the OROSIL C18 to analyze a wide range of acidic, basic, and neutral compounds. Data is presented on a few representative compounds and unique separation criteria with their associated chromatograms.

June 01, 2008

The continuing and growing trend toward high-speed analysis in all fields of chromatography is also reflected in enantiomer separations. A variety of new 3-μm columns has been designed to meet this need. Applications of CHIRALCEL® OD-3 and CHIRALPAK® AD-3 in some enantiomer separations demonstrate the benefits of transitioning to such media.

June 01, 2008

The natural colors of the food we eat add greatly to the enjoyment of life. Consequently, the art and science of recreating food's natural colors through the use of synthetic colorants is an important success factor in the food industry.