Mass Spectroscopy Articles

The effect of dispersing a solvent into charged droplets, when applying a high electric potential to an effluent capillary, was first described by Zeleny in 1917 (1). It was investigated in more detail in 1955 by Drozin (2) and used in pioneering work for an electrospray interface by Dole et al (3,4). This was continued in 1984 by Yamashita and Fenn (5,6), which finally resulted in a description of an LC–MS interface in 1985 (7). Today electrospray ionization (ESI) is the most widely used ionization technique in LC–MS. This is especially true in protein mass spectrometry, where series of multiply charged ions are observed and used to determine the molecular mass (8,9). Furthermore, after tryptic digestion, the resulting peptides can be measured with LC–ESI-MS, to obtain amino acid sequence information after a second MS/MS-step (10–14).

For decades, selenium has been considered an essential element to human nutrition. It plays an important role in many biological processes, generally exerting its biological affect through selenoproteins. Selenium is incorporated specifically into proteins by a UGA codon that encodes for selenocysteine. There are over 30 known selenoproteins in mammalian systems, including a number of glutathionine peroxidases (GPx), iodothyronine 50-deiodinases (IDI), sperm capsule selenoprotein, and thioredoxin reductase (1, 2). Selenium also is required for the production of triiodothyronine, a thyroid hormone necessary for healthy brain and bone development, normal growth, and thermoregulation (3). Proper immune function also is selenium-dependent, which recently has prompted investigations into its ability to inhibit HIV and AIDS progression (4).

In the pharmaceutical industry, liquid chromatography–tandem mass spectrometry (LC–MS-MS) is already an established method for quality control and quantification of drugs in different matrices. Additionally, this hyphenated analytical tool is becoming more and more important in clinical chemical analysis (that is, in therapeutic drug monitoring).

LC–MS-MS is a very powerful analytical technique because it combines the separation power of LC with the sensitivity and selectivity of MS. However, LC–MS-MS possesses two major drawbacks when analyzing drugs in biological fluids. The first, associated with the mass spectrometer, is that the electrospray source is very susceptible to matrix-related ion-suppression effects. The second drawback concerns the LC. Because of irreversible adsorption and precipitation effects caused by high molecular weight sample components (for example, proteins) complex biofluids cannot be injected and analyzed directly.

Fusarium fungi are capable of producing, to a variable degree, two or more toxins. The major Fusarium mycotoxins are fumonisins, A- and B-trichothecenes, and zearalenone (ZON) (1). Trichothecenes are responsible for a wide range of toxicity in animals, including feed refusal, weight loss and vomiting. In particular deoxynivalenol (DON) can inhibit protein biosynthesis and has been reported as an immunosuppressant (2). To reduce the levels of biogenic toxins, European authorities are currently discussing further regulations on mycotoxins. Within the European Union (EU), harmonized legislation is setting maximum limits for aflatoxins and ochratoxin A in cereals and cereal products. Limits for Fusarium toxins (DON, ZEA, HT2, and T2) are currently being drafted in EU member states; for example, maximum limits for Fusarium toxins (DON 100–500 μg/kg, ZON 20–50 μg/kg) were established in February 2004 (3).

Liquid chromatography-mass spectrometry (LC–MS) has been an important analytical tool in support of drug discovery and drug development for some time. LC–MS provides a combination of detection selectivity and sensitivity, speed of analysis, and robust performance that is well suited to the rapid structural characterization of candidate therapeutic substances and high-throughput screening assessment of pharmacological activity (1-3). More recently, LC–MS has been used for patient monitoring in clinical drug trials (4-6) and for the detection of drugs of abuse (7). As knowledge has grown about the relationship between genetic and environmental factors and individual responses to drugs, clinicians have sought new ways to monitor drug responses accurately to assess and tailor drug therapies more effectively. With its advantageous performance characteristics, LC–MS has proven to be a useful tool for collecting data characterizing these relationships.

GUIDE TO LC–MS.
Summary Mass spectrometry provides a faster of fractions. The procedure described involves initial protein purification on ÄKTA™ purifier 10, automated and rapid desalting of fractions on Ettan microLC, and MS analysis by electrospray ionization time-of-flight mass spectrometry (ESI-ToF MS) using Ettan ESI-ToF. Ettan LC–MS provides high-quality mass spectra of a wide range of high molecular weight proteins including bovine serum albumin, cytochrome C, transferrin, ovalbumin, myoglobin, a monoclonal antibody and two -lactoglobulin variants.
 

Part II of this four-part series on mass calibration looks at the chemical compounds used as mass calibrants in mass spectrometry.

Corrections belong up front. Part I of this column series included a description of mass scales and units, exhorting readers and users to pay attention to basics. An essential part of basics is good proofreading, a skill that seems to have deserted this author in preparing Part I. Table II in Part I (Spectroscopy 19[11], 32–34 [2004]) lists the exact mass for 13C incorrectly as 13.993355. I considered assigning the blame for such an egregious error to electronic voting machines (it was, after all, an election year, and an election month), or fat fingers on the keyboard, but instead I admit to sloppiness and apologize for the error. The correct value for the exact mass of 13C is 13.003355.My thanks to the readers who notified me of the error and demanded a recount.

Mass spectrometry systems have specific vacuum requirements. New developments in oil-free, or dry, primary vacuum pumps have been introduced recently and are discussed in this article with respect to capacity, throughput, and specific pumping requirements for process gases.

 There are many drivers for vacuum configuration in mass spectrometry (MS) and other scientific instrumentation applications. These include: vacuum performance of the primary pump itself (speed, compression, power, and so forth); environmental impact, power, construction, service interval, and the requirements (if any) for oil; regulatory compliance; cleanliness of the vacuum produced; and compatibility with process and target gases–vapors. MS systems have very specific vacuum (physics and engineering) requirements. The systems primarily considered here are liquid chromatography–MS (LC–MS), gas chromatography– MS (GC–MS) and inductively coupled plasma MS (ICP-MS).

This article describes the development of a simple enzymatic reactor and its interfacing with a mass spectrometer for high-throughput mass mapping of peptides. This procedure combines preconcentration and enzymatic digestion in the nano-electrospray emitter.

The recently decoded human genome is believed to be a massive source of information that will lead to improved diagnostics of diseases, earlier detection of genetic predispositions to diseases, gene therapy, rational drug design and pharmacogenomic “custom drugs”. The upcoming “post-genomic” era will then target the gene expression network and the changes induced by effects such as disease, environment or drug treatment. In other words, knowledge of the exact composition of proteins within a living body and the changes in composition that reflect both healthy and diseased states will be used to determine the pharmacological action of potential drugs.

In the last five years, the acceptance and implementation of packed-column supercritical fluid chromatography–mass spectrometry (pSFC–MS) to drug discovery applications has gained momentum. This article describes the pros and cons of pSFC–MS and attempts to demonstrate its broad applicability to such fields as high-throughput analysis, purity assessment, structure characterization and purification. Finally, an outlook for the future of this technique is presented.