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Techniques for the characterization of two boronic acid peptides as their cyclic boronate ester derivatives by positive-ion ammonia chemical ionization (CI) and positive-ion liquid secondary ionization (LSIMS) are described and results presented. These techniques avoid the complications introduced by the thermally induced processes that boronic acids may undergo when the mass spectrometric characterizations of free boronic acids are attempted. Derivatizations for CI analysis were accomplished via a simple benchtop method using several polyfunctional nucleophilic derivatizing agents (ethylene glycol, glycerol, et al.), while derivatizations for LSIMS analysis were accomplished via both benchtop and previously established in situ methods using the same derivatizing agents. Certain previously held misconceptions about the LSIMS mass spectrometry of boronic acids are examined.

Mass spectrometric data can be obtained for compounds in bead-bound combinatorial mixtures by several techniques. However, little specific information is available regarding (1) how well these data differentiate between candidate structures in large combinatorial pools, (2) what precision of data is required to achieve adequate specificity in these analyses, and (3) what are the best strategies for applying these data. In this work, computer modeling is used to address these questions. Strategies employing multiple filters (i.e., those that differentiate possible structures using more that one measured mass spectral parameter) are found to provide better specificity and to be more robust (that is, the specificity is less dependent on the precision of the data) than discrete filters. With moderate precision data (e.g., 50 ppm mass precision, 10% isotope ratio precision), multiple filter strategies are found to give unequivocal results for [approximately]80% of the populations of combinatorial mixtures with most of the remaining degeneracy at the 2-fold level. A simple protocol for the application of multiple filter methods is presented.

The combination of orthogonal TOF/ESI MS exact mass measurement and on-line chromatography represents a powerful analytical tool for identifying unknown components in complex mixtures and is being widely utilized. The precision of these mass data is often incorrectly estimated as the precision or mean deviation obtained for reference standards under standard conditions. But, the precision of a mass measurement is dependent on the number of ions sampled in the measurement and, thus, is likely to be different for every measurement. A simple procedure for correctly estimating the precision of a specific mass measurement is presented, the limits of the procedure are investigated, and the utility and validity of the procedure are demonstrated.

Combinatorial chemistry has opened a new realm of chemical entities in the search for novel therapeutics. Combinatorial chemistry is currently adding hundreds of thousands of compounds to similar numbers available from years of synthesis by medicinal chemistry. It is not unreasonable to expect that over the next several years, nearly a million compounds will be available for screening against each therapeutic target. The number of potential targets will also be increasing with the advances in genomics. With the increasing number of compounds to be screened against an increasing number of targets, it is becoming increasingly difficult and costly to obtain the required amounts of key biological material needed to screen these compounds. One obvious solution is to miniaturize the assays so that the biological reagent supply doesn't need to increase. To this end, we have developed an ultra-high throughput screening system comprised of a new plate design (9600-well), detection system, and liquid handling system. This new format is capable of performing assays in as little as 0.2 Al. The results obtained from this system compare favorably to those obtained in the standard 96-well format. [ABSTRACT FROM PUBLISHER]

A systematic procedure for establishing elemental composition from a concerted high-resolution precise mass analysis of the monoisotopic molecular ion and a low-resolution moment analysis of its isotope pattern is described. The combined method is more selective than a precise mass analysis alone. Factors affecting the enhancement in selectivity and some instrumental factors are discussed. [ABSTRACT FROM AUTHOR]

The basic principles of an approach to mass spectral elemental analysis based on the moments of the low-resolution isotopic pattern are presented. The moment analyses are faster and more general than conventional methods of isotopic pattern analysis. The procedure is applied to the analysis of compounds which exhibit 'pure' molecular ions and the analysis of compounds which produce significant fragment ions at masses overlapping the molecular ion. It appears as though reasonable selectivity can be achieved for low relative molecular mass compounds (relative molecular mass < 400 a) using standard instrumentation and data acquisition softy are. Preliminary studies of instrumental factors affecting the measurement of moments are discussed. [ABSTRACT FROM AUTHOR]

The application of the fundamental equation for isotopic dilution mass spectrometry to real samples often requires using approximations which can lead to significant errors. A new approach relating the composition of isotopic mixtures to the average mass of the mass spectral pattern is presented. The fundamental equations are completely general; they should be applicable to a wide range of problems and may provide improved accuracy and precision in some cases. [ABSTRACT FROM AUTHOR]

One of the remaining challenges in providing effective preparative LC-MS purification is balancing throughput and compound purity. We describe here an approach to optimizing preparative LC-MS methods that provides significantly better chromatographic resolution and, hence, better compound purity than generic preparative LC methods consuming the same amount of time. This approach is easier to implement, is more rugged, and permits significantly greater flexibility than previously reported approaches. The instrument configurations and protocols presented here are specifically tailored for open access support, but the basic approach is equally suitable and effective in high-throughput situations.

Preparative liquid chromatography-mass spectrometry (LC-MS) is widely used in parallel synthesis schemes to expedite purification. Recently, an alternative sample loading scheme, at column dilution, has been shown to dramatically increase the mass loading capacity of LC-MS purification methods. The prototype system utilized separate sample loading and binary gradient pumps. We report here a configuration for effecting at-column dilution using only the two pumps that provide the binary gradient flow. The advantages of a two-pump configuration are reduced cost, reduced space requirements, simplified control, and reduced service and maintenance issues. The two-pump at-column-dilution configuration is demonstrated for large- and small-scale LC-MS purifications. Purification on scales appropriate for high-throughput parallel synthesis can be achieved with small-scale chromatography using at-column dilution; purification of 20 mg of material is demonstrated using a 4.6 mm x 150 mm column and a flow rate of 3 mL/m. Reducing the scale of chromatography required for LC-MS purification has significant benefits, including the following. It requires less expensive columns, consumes less solvent, generates smaller-volume fractions (shorter dry-down time and the ability to collect into small-volume collector formats, such as 96-well plates), and has the potential for faster separations.