High performance liquid chromatography software

Fig. 6.1.4 Components of the high-performance liquid chromatography (HPLC) system for oxidized pterins with column switching. The autosampler starts the chromatography software, which controls valves and pumps. Pump 3 is used only for rinsing the precolumn. Details of the three valves and their different positions are explained in Fig. 6.1.5...

The methods for each study are divided into the initial protein separation step, a second separation step if applicable, the type of mass analysis, and the software used for peptide identification. ID = one dimensional polyacrylamide gel electrophoresis, 2D = two dimensional polyacrylamide gel electrophoresis, MS = mass spectrometry (peptide mass fingerprinting), MS/MS = tandem mass spectrometry, MALDI-TOF = matrix assisted laser desorption/ionization-time of flight, MS FIT = software from Protein Prospector, http //prospector.ucsf edu/, ESI = electrospray ionization, Q-TOF = quadrupole-time of flight, PPSS2 =Protana s Proteomic Software Suite (ProtanaEngineering, Odense, Denmark), Mascot = Matiix Science, http //www.matrixscience.com/, TOF-TOF = MALDI plus TOF tandem mass spectrometry, RP-HPLC = reverse phase high performance liquid chromatography, Q-IT = quadrupole ion trap, LIT = linear ion trap. Bioworks = Thermo Electron Corporation. 

Current proteomic methods of protein separation by 2D gel and/or high-performance liquid chromatography (HPLC), as well as the identification of proteins by mass spectrometry (MS) in conjunction with protein databanks and several software programs are satisfactory. These methods must be improved in the future to be more efficient. Some improvements that are expected in the near future must include the following strategies as discussed by others as well ... 

Mixtures can be identified with the help of computer software that subtracts the spectra of pure compounds from that of the sample. For complex mixtures, fractionation may be needed as part of the analysis. Commercial instruments are available that combine ftir, as a detector, with a separation technique such as gas chromatography (gc), high performance liquid chromatography (hplc), or supercritical fluid chromatography (96,97). Instruments such as gc/ftir are often termed hyphenated instruments (98). Pyrolyzer (99) and thermogravimetric analysis (tga) instrumentation can also be combined with ftir for monitoring pyrolysis and oxidation processes (100) (see Analytical methods, hyphenated instruments). 

Many components of SEC systems are well known and ubiquitous in an analytical lab. Size exclusion chromatography systems require at least an isoaatic pump, a high-pressure injection systan (manual or automated), one or more SEC colunm(s), one or more detector(s), and data evaluation units with software to acquire, calibrate, and analyze data. Other components, such as degassers or column compartments are often optional and depend on the particular application and on the lab environment and conditions. Although SEC and high-performance liquid chromatography (HPLC) seem to be very similar in instrumentation requirements, the theoretical background and separation mechanism of both techniques are entirely different. The major differences in the practice of SEC are outlined in Table 9.2. 

SEC was performed according to the method of Chin et al. (1994). A Shodex KW802.5 SEC column (Waters Corp., Milford, MA., USA) was used and a Waters liquid chromatography system consisting of the following components was used for the analysis Waters 501 high pressure pump. Waters 717 autosampler, InterAction column temperature control oven, Waters 484 UV/VIS detector, and Waters Millenium 2.0 computer software package. 

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