High-throughput ultrafiltration

We have developed a high throughput ultrafiltration affinity screening method coupled to MS (affinity selection/mass spectrometry ASMS), which works with any soluble target and small molecule library (including natural products). ASMS is amenable to parallelization, efficient and robust enough to allow study... 

Zhang, J. Musson, D.G. Investigation of high-throughput ultrafiltration for the determination of an unhound compound in human plasma using liquid chromatography and tandem mass spectrometry with electrospray ionization. J. Chromatogr. B, 2006, 843, 47-56. 

Pulsed ultrafiltration MS (PUF-MS) represents an inline high throughput affinity screening method with a variety of potential uses in the discovery and development of pharmaceuticals [22]. The in-line combination of solution-phase equilibration, ultrafiltration, and electrospray liquid chromatography mass spectrometry (LC-ESI-MS) facilitates the identification of high affinity target-specific... 

Screening for inhibitors of dihydrofolate reductase using pulsed ultrafiltration mass spectrometry. Comb Chem High Throughput Screen 1998, 1, 47-55. 

Plasma protein binding is also an important parameter in the pharmacokinetic field. Frontal analysis combined with capillary zone electrophoresis (CZE-FA) (67-69) is a powerful technique for high-throughput assay, because it is relatively rapid and easy to automate, in comparison with conventional methods such as dialysis, ultrafiltration, and ultracentrifugation. Recently, we introduced the EKC approach with ionic CDs to frontal analysis for anionic drugs that cannot be analyzed by conventional CZE-FA (70). In this approach, ionic CDs work as an EKC pseudostationary not for proteins but for small solutes. 

If automated 96 well plate ultrafiltration system is used, high throughput with sample handling by robotic system. 

The UF can either performed in a single ultrafilter unit or in a 96 well plate ultrafiltration system, latter in a semi-automatic high-throughput determination with sampling handling by a robotic system (Fung et al. 2003 Jordan et al. 2000). 

Plastic microdevices for high-throughput screening with MS detection were also prepared for detection of aflatoxins and barbiturates. These devices incorporated concentration techniques interfaced with electrospray ionization MS (ESI-MS) through capillaries [2], The microfluidic device for aflatoxin detection employed an affinity dialysis technique, in which a poly (vinylidene fluoride) (PVDF) membrane was incorporated in the microchip between two channels. Small molecules were dialyzed from the aflatoxin/antibody complexes, which were then analyzed by MS. A similar device was used for concentrating barbiturate/antibody complexes using an affinity ultrafiltration technique. A barbiturate solution was mixed with antibodies and then flowed into the device, where uncomplexed barbiturates were removed by filtration. The antibody complex was then dissociated and electrokinetically mobilized for MS analysis. In each case, the affinity preconcentration improved the sensitivity by at least one to two orders of magnitude over previously reported detection limits. 

In principle, ultrafiltration can be an easy way to quantify free drug fraction present in plasma, serum, or other biological fluids. The approach is not without pitfalls, however, in that ion suppression, clogged membranes, and poor sensitivity due to extensively bound drugs can derail this type of assay. Still, ultrafiltration has a lot of untapped potential and could become pervasive as membranes are made more robust and adapted to high-throughput formats such as 96-well plates. A related sample preparation technique, microdialysis, is discussed in Chapter 12. 

Nikolic, D. Van Breemen, R.B. Screening for Inhibitors of Dihydrofolate Reductase Using Pulsed Ultrafiltration Mass Spectrometry, Comb. Chem. High Throughput Screening 1,47-55 (1998). 

Nikolic D, Fan PW, Bolton JL, van Breemen RB. Screening for xenobiotic electrophilic metabolites using pulsed ultrafiltration—mass spectrometry. Comb Chem High Throughput Screen 1999 2 165-175. 

Gu,C. Nikolic,D. Lai,J. Xu,X. VanBreemen,R.B. Assays of ligand-human serum albumin binding using pulsed ultrafiltration and liquid chromatography-mass spectrometry. Comb. Chem. High Throughput Screen. 1999,2, 353-359. 

The second solution to increasing the throughput of pulsed ultrafiltration mass spectrometry has been to miniaturize the ultrafiltration chamber volume while maintaining the flow rate and chamber pressure. Because the ultrafiltration membrane cannot withstand high pressure without rupturing, the ultrafiltration process cannot be accelerated simply by increasing the flow rate through the chamber. The approach of Beverly et al. (72) was to fabricate a 35-/u,L ultra-... 

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