Purification mass-directed preparative

The technique of preparative LC/MS, introduced in the late 1990s was the first technique to greatly simplify the purification process. For the first time, preparative LC/MS (PrepLCMS) methods allowed the concept of one compound/one fraction to be realized [51-55]. In the Prep LC-MS mode, the mass 

Debate exists as to whether UV-based or mass-based fraction collection is the more appropriate tool for purifying compound libraries. The choice of 

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To deal with this problem we set about coupling an autoprep system to a mass spectrometer [14]. The addition of a mass spectrometer enabled the system to become far more specific and by inputting the molecular weight, the system would collect only the compound(s) of interest. In the majority of cases, only one desired component is required to be collected so this means that one sample will prcxluce only one purified fraction, two samples will produce only two and similarly, a plate of 80 samples will produce only 80 purified fractions. A mass-directed preparative HPLC (MS-prep) instrument would therefore eliminate the re-analysis and recombination steps from the purification process. 

Figure 14 High-throughput mass-directed auto purification system schematic. System as shown is configured for preparative two-column regeneration and an analytical column for method development or to reanalyze fractions.

Zhang, X. et al., Development of a mass-directed preparative supercritical fluid chromatography purification system, J. Comb. Chem., 8(5), 705, 2006. 

Since application of fast, generic LC methods with mass spectrometry (LC/MS) is emerging as the technique of choice for assessing the progress and final quality of large combinatorial arrays in drug discovery, it will be discussed in some detail, along with other detection techniques. Mass-directed purification and characterization on the preparative scale will also be addressed. 

Xu R, Cai Z, Fogelman K, Wikfors R, Worle V, Stublen N, Kassel DB. Mass-directed purification of compound libraries by automated semi-preparative SFC/MS. Oral presentation and poster. American Society for Mass Spectrometry. Orlando, FL, 2002. 

Wang and co-workers have reported that a preparative SFC system can be interfaced with a single quadrupole mass spectrometer for mass-directed fraction collection. Samples with no chromophore (Ginsenoside Rb, Ginsenoside Rc, and Ginsenoside Re) were isolated near homogeneity. A more sophisticated preparative SFC system was patented by Maiefski et al. There are four parallel ehannels in this system, and there is a UV detector for each channel. Since the eluent can be also splitted into a mass spectrometer, this system is capable of both UV and MS directed purification. 

The inherent lower viscosity of the SFC mobile phase relative to HPLC yields significant advantages for chiral separation applications.Indeed, preparative SFC is experiencing an increase in utilization in the pharmaceutical industry for chiral purification for a wide range of scales. Now, mass-directed purification technology has been incorporated with SFC for both chiral and achiral purification applications,and commercial prep-SFC/ MS systems are currently in development. 

The development of many novel techniques that make our existence so comfortable has been intimately associated with the accessibility of suitable analytical methods. Liquid chromatography-mass spectrometry (LC-MS) is a powerful technique used for various applications based upon its very high sensitivity and selectivity. Generally, its applications are oriented toward the detection and identification of chemicals in a complex mixture. Preparative LC-MS systems can be used for fast and mass-directed purification of natural product extracts and new molecular eutities important to food, pharmaceutical, agrochemical, and other industries. 

Purification of the radioactive tracer was modified to include a fractional sublimation before a single extraction—recrystallization cycle to conserve the tracer material. Microgram samples were prepared in melting point capillaries for assay by mass spectroscopic analysis (Table III), made by direct probe injection of the sample into the ion source (18). The probe was heated rapidly to 200°C, and mass spectra were obtained during vaporization of the sample. Tri-, tetra-, and pentachlorodibenzo-p-dioxins vaporized simultaneously with no observed fractionation. 

Specifically for triazines in water, multi-residue methods incorporating SPE and LC/MS/MS will soon be available that are capable of measuring numerous parent compounds and all their relevant degradates (including the hydroxytriazines) in one analysis. Continued increases in liquid chromatography/atmospheric pressure ionization tandem mass spectrometry (LC/API-MS/MS) sensitivity will lead to methods requiring no aqueous sample preparation at all, and portions of water samples will be injected directly into the LC column. The use of SPE and GC or LC coupled with MS and MS/MS systems will also be applied routinely to the analysis of more complex sample matrices such as soil and crop and animal tissues. However, the analyte(s) must first be removed from the sample matrix, and additional research is needed to develop more efficient extraction procedures. Increased selectivity during extraction also simplifies the sample purification requirements prior to injection. Certainly, miniaturization of all aspects of the analysis (sample extraction, purification, and instrumentation) will continue, and some of this may involve SEE, subcritical and microwave extraction, sonication, others or even combinations of these techniques for the initial isolation of the analyte(s) from the bulk of the sample matrix. 

Direct injection API-Electrospray MS is capable of analyzing much larger and less volatile substances than either EI/MS or CI/MS. As a result, this method is often used to provide structural information on peptides, proteins, and polymers derived from both natural and synthetic processes it is also useful in the analysis of many natural compounds including molecules such as saponins and flavonol glycosides, derived from plants. When using direct injection API-electrospray, partial purification and EC preparation are performed elsewhere and a collected fraction is dissolved in an appropriate solvent and injected as a bolus into the mass spectrometer (flow or direct injection or syringe infusion). This has an advantage, as the mass... 

Semiconductors made from elements in Groups 3A(13) and 5A(15) are typically prepared by direct reaction of the elements at high temperature. An engineer treats 32.5 g of molten gallium with 20.4 L of white phosphorus vapor at 515 K and 195 kPa. If purification losses are 7.2% by mass, how many grams of gallium phosphide will be prepared ... 

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