Liquid chromatography - mass spectrometry

Coupling of liquid chromatography to mass spectrometry has not only led to a wide variety of interfaces, it also fostered the development of new ionization methods (Chap. 12) [8,9,12-15,84]. ESI, APCI, or APPI are suitable for LC-MS, then-selection depending on sample properties like molecular mass and polarity (Chap. 12.7). 

Most frequently, high-pressure liquid chromatography (HPLC) is used in LC-MS work, while more recently ultrahigh-pressure liquid chromatography (UHPLC or ultra performance liquid chromatography, UPLC, as it is termed by one manufacturer) has come into use. For very low saiiple amounts, nanoLC, i.e., capillary LC, can directly be interfaced to nanoESI. 

In the analytical practice, LC-MS plays a tremendous role that could be testified here by a large number of applications [9-11,50,85]. However, other than proper adjustment of the interface to the liquid flow from the chromatograph the operation of LC-MS does not require dedicated techniques on the MS side. All scanning and ion monitoring techniques discussed so far in this chapter can equal- 

Example I Impurities in pharmaceutical preparations. UV photodiode array (PDA) and ESI-TOF detection can be combined if the LC effluent is split or the PDA precedes the ESI interface. The detection methods complement each other in that their different sensitivities towards components of a mixture prevent substances from being overlooked. RICs help to differentiate a targeted compound - an unknown impurity in this case - from others and to identify eventually present isomers. Finally, accurate mass measurement helps in the identification of the unknown, the [M+H] ion of which was assigned as [C2oH2iNio] The mass error in the order of 5 ppm was considered perfect for the oaTOF instruments of that time (Fig. 14.13) [27]. 

In addition, co-eluting matrix compounds are often present in a sample from biological material. This will most likely suppress the total ion current formed in the ionization chamber, thus lowering the sensitivity of the method of analysis (Buhrman, Price and Rudewicz, 1996 Chan, 1996 Knebel, Sharp and Madigan, 1995). By using chromatography, ion suppression is reduced. Furthermore, unexpected compounds, metabolites and/or artefacts may be easier to identify with HPLC-MS. With flow or loop injection, these compounds may be overlooked with loss of (important) data as a result. 

Many mass spectrometers today can work in Windows or Macintosh environments. The development of computer hardware and software over the years has made data handling and processing fast and easy. It is now common practice to produce high-quality copies of spectra collected during an analysis run to be copied and pasted into a document while still performing the analysis. This was an impossible task less than ten years ago when one had to wait for the end of analysis before processing any data obtained from that analysis. 

Hydrogenated samples of MGDG and DGDG were analysed by Kim, Yergey and Salem (1987). The classes were separated within 3 min, and 

Odham et al. (1988) compared plasmaspray with thermospray by using ammonium acetate as a buffer, where PSP required no addition of buffer prior to ionization. The total ion current obtained in PSP was significantly higher than previously reported for TS (filament-on). No or very few molecular ions were observed as a result of the harder ionization in PSP compared with TS. Diacylglycerol- and monoacylglycerol-derived fragments of phospholipids were obtained. Cation-exchange HPLC separated PI and PE in a phospholipid extract from bacterial cells Pseudomonas fluorescens). Furthermore, PC, PE, PA, PS, cardiolipin (CL) and polyphosphoinositides (PIP and PIP-2) were also studied. 

Trinitrobenzene derivatives of PE and PS from human red blood cells and rat brain were analysed by Hullin, Kim and Salem (1989). For quantification purposes, reversed-phase HPLC with UV detection was used. HPLC-TS (filament-on) was used for confirmation of molecular species separation. 

LC-MS is a powerful technique for analysis of complex mixtures, where additional analytical information is required to positively confirm the identity of the separated compounds or few separations are obtained. Among the interfacing systems used to couple LC with MS, the newly developed electrospray/ionspray (ES/ISP) mass spectrometric liquid interface allows undoubted advantages in terms of sensitivity and capability to analyze large, thermally labile, and highly polar compounds in addition, tandem MS techniques are useful for structural elucidation studies. 

There are different approaches to the hyphenation of HPLC with MS and some commercial interfaces are available. Atmospheric pressure ionization (API), a soft and highly efficient method suitable for analysis of polar, ionic, high-molecular-mass, and thermally labile compounds amenable to LC has greatly increased the popularity of LC-MS. API-based interfacing systems are ES and ISP, which are liquid-based interfaces. 

Selected Examples of Isoflavone and Lignan Analysis by High-Performance Liquid Chromatography (HPLC) and Liquid Chromatography-Mass Spectrometry (LC-MS) 

Human urine and plasma (premenopausal women) Ingestion of C-labeled isoflavone daidzein and genistein 7-day dietary diaries and 24-h pooled urine and collection of blood sample (10 mL) from (2-72 h) 

Rat plasma daidzein, genistein, glycitein, equol, 4-ethyl phenol, and biochanin 

Jones and co-workers [228] used particle beam liquid chromatography coupled with a MS detector to determine styrene oligomers up to the w-18 oligomer of PS. Bryant and Semiyen [229] analysed cyclic oligomers of polybutylene terephthalate using column chromatography GPC (size exclusion), fast atom bombardment mass spectrometry, and tandem mass spectrometry (MS-MS). They obtained molecular weight distributions. 

Dwyer [96] investigated the combination of SEC and matrix-assisted laser desorption/ ionisation mass spectrometry (MALDI-MS). He showed that there are a number of factors that can influence results. Raghaven and Egwim [230] used LC-MS in degradation studies of polyester films in alkali solution. 

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Liquid chromatography/mass spectrometry (LC/MS) is an analytical technique combining the advantages of an LC instrument with those of a mass spectrometer. 

Brown, M.A., Liquid Chromatography/Mass Spectrometry Applications in Agricultural, Pharmaceutical and Environmental Chemistry, Oxford University Press, Oxford, 1998. 

Niessen, W.M.A. and van der Greef, J, Liquid Chromatography-Mass Spectrometry, Marcel Dekker, New York, 1992. 

Yergey, A.L., Edmonds, C.G., Lewis, I.A.S., and Vestal, M.L., Liquid Chromatography/Mass Spectrometry Techniques and Applications, Plenum Press, New York, 1990. 

During gc/ms or liquid chromatography/mass spectrometry (Ic/ms) acquisitions, it is possible to perform a mixture of the experiments described in Table 2 for different time windows, with the experimental parameters, such as the coUision energy, optimized for each analyte. 

A liquid chromatography-mass spectrometry (LC-MS) method that can quantitatively analyze urinar y normal and modified nucleosides in less than 30 min with a good resolution and sufficient sensitivity has been developed. Nineteen kinds of normal and modified nucleosides were determined in urine samples from 10 healthy persons and 18 breast cancer patients. Compounds were separ ated on a reverse phase Kromasil C18 column (2.1 mm I.D.) by isocratic elution mode using 20 mg/1 ammonium acetate - acetonitrile (97 3 % v/v) at 200 p.l/min. A higher sensitivity was obtained in positive atmospheric pressure chemical ionization mode APCI(-i-). 

Very little in the way of advances has occurred since 1971 in the applications of ultraviolet or infrared spectroscopy to the analysis of fluonnated organic compounds Therefore, only gas-liquid chromatography, liquid chromatography, mass spectrometry, and electron scattering for chemical analysis (ESCA) are discussed The application of nuclear magnetic resonance (NMR) spectroscopy to the analysis of fluonnated organic compounds is the subject of another section of this chapter... 

The ion spray liquid chromatography/mass spectrometry (LC-MS) interface coupled via a postsuppressor split with an ion chromatography (IC) has been used in the analysis of alcohol sulfates. The IC-MS readily produces the molecular weight while the tandem mass spectrometric detection IC-MS-MS provides structural information [305]. 

Hummel D, Loffler D, Fink G, Temes TA (2006) Simultaneous determination of psychoactive drugs and their metabolites in aqueous matrices by liquid chromatography mass spectrometry. Environ Sci Technol 40 7321-7328... 

Farre M, Ferrer I, Ginebreda A, Figueras M, Olivella L, Tirapu L, Vilanova M, Barcelo D (2001) Determination of drugs in surface water and wastewater samples by liquid chromatography-mass spectrometry methods and preliminary results including toxicity studies with Vibrio fischeri. J Chromatogr A 938 187-197... 

Liquid Chromatography -Mass Spectrometry An Introduction. Robert E. Ardrey... 

In this book, I have tried to show the way in which high performance liquid chromatography-mass spectrometry (LC-MS) has developed, somewhat slowly it has to be said, into a powerful hybrid analytical technique. 

The characteristics of an ideal liquid chromatography-mass spectrometry interface have been discussed, with emphasis having been placed upon the major incompatibilities of the two component techniques that need to be overcome to allow the combination to function effectively. 

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