Platelets, human chromatography

A. General description Eptifibatide is a cyclic heptapeptide containing six amino acids and one mercaptopropionyl residue. An interchain disulfide bridge is formed between the cysteine amide and the mercaptopropionyl moieties. Eptifibatide binds to the platelet receptor glycoprotein (gp) Ilb/IIIa of human platelets and inhibits platelet aggregation. The eptifibatide peptide is produced by solution-phase peptide synthesis, and is purified by preparative reverse-phase liquid chromatography and lyophifized. 

Immler, D. Gremm, D. Kirsch, D. Spengler, B. Presek, R Meyer, H. E. 1998. Identification of phosphorylated proteins from thrombin-activated human platelets isolated by two-dimensional gel electrophoresis by electrospray ionization-tandem mass spectrometry (ESI-MS/MS) and liquid chromatography-electrospray ionization-mass spectrometry (LC-ESI-MS). Electrophoresis, 19,1015-1023. 

An important (and early) objective of this chapter was to illustrate an approach to evaluation of the lipid classes present in a mammalian cell, such as the human platelet. Thus, armed with the information presented above, it is now feasible to start to explore this problem. As emphasized earlier, TLC can provide significant information on the types (or classes) of lipids present in a lipid extract analytical and/or structural technique to elucidate the chemical nature of the individual groups of compounds. A brief examination of the results that can be obtained in two types of thin-layer chromatography follows. 

Using the total platelet lipid extract as described in the section entitled Isolation of Human Platelets, single-dimensional thin-layer chromatography on a 250- pm-thick silica gel G-coated plate with a solvent system of chloroform-methanol-water (65 35 7, v/v) will yield the pattern shown in Figure 3-2. 

J2. Jiang, H., and Balazy, M., Detection of 3-nitrotyrosine in human platelets exposed to peroxynitrite by a new gas chromatography/mass spectrometry assay. Nitric Oxide 2, 350-359 (1998). 

Human-platelet membranes have been extracted with lithium 3,5-diiodosalicylate, and the soluble glycoproteins separated on O-phosphonocellulose.849 The purified, membrane glycoprotein, molecular weight 100,000, was immunochemically identical to a sample purified from platelet-membrane extract by means of con A-Sepharose affinity chromatography. No further characterization was reported,... 

Borg C, Lim CT, Yeomans DC, Dieter IP, Komiotis D, Anderson EG, LeBreton GC. Purification of rat brain, rabbit aorta, and human platelet thranboxane A / prostaglandin H pecqitors by immunoaffinity chromatography employing anti-peptide and anti-receptor antibodies. J Biol Chem 1994 269 6109-16... 

Purified by affinity chromatography from human platelets. 

Blandini F, Martignoni E, Pacchetti C, Desideri S, Rivellini D, Nappi G. Simultaneous determination of L-dopa and 3-O-methyldopa in human platelets and plasma using high-performance liquid chromatography with electrochemical detection. J Chromatogr B Biomed Sci Appl 1997 700 278-82. 

We have reported that bulbs of Crocus sativus contain a poly-histidine protein factor, which aggregates human platelets in vitro. The molecular weight of this factor as found by gel filtration chromatography on a Sephadex G-75 column and sodium dodecylsulfate (SDS) polyacrylamide slab gel electrophoresis is 42,000Da [79], In addition, another low molecular weight protein factor isolated from the same crude extract was found to inhibit human platelet aggregation in vitro [80]. The effect of Crocus sativus on blood coagulation has been also examined by Nishio et al [81]. 

A glycoprotein (mol. wt, 1.5 X 10 ) has been isolated from whole human platelets by affinity chromatography on immobilized wheat-germ agglutinin. Extraction of the whole platelets with lithium di-iodosalicylate released three major glycoprotein fractions. 

Purification of the peptidase active against insulin B-chain, etc., from rat kidneys Separation of enzyme cofactors Afllnity chromatography of anti-(lacto-JV-difucohexaose I) antibody immunoglobulin Purification of membrane glycoproteins from human-blood platelets... 

A New Method to Quantitate Lipid Contents in Human Platelets by Thin-Layer Chromatography with Flame Ionization Detector Tokai J. Exp. Clin. Med. 5(2) 177-186 (1980) CA 93 128147r... 

The Study of the Fatty Acids and Aldehydes of Human Platelet Phos-phatides by Gas Chromatography... 

The applications surveyed in Tables 4 through 9 illustrate the general principles of vitamin E assays outlined in I.E. 1. Specifically, each matrix puts a different emphasis on the E vitamers to be determined. Thus, in serum/plasma a-tocoph-erol is clearly the main compound of interest. Accordingly, leversed-phase chromatography with UV detection is the indicated technique for this purpose. In addition, as part of the assessment of the antioxidant status of humans, tocopher-ols are determined concurrently with retinoids (retinol, retinyl palmitate) and carotenoids (particularly p-carotene). a-Tocopherol is also the principal target compound in erythrocytes and platelets but here, predictably, the quantitation of a-tocopherolquinone may also be meaningful as an indicator of oxidative stress (7). The need to assay this minor constituent in turn justifies coulometric detection. The analysis of tissues is complementary to that of plasma and red blood cells and mainly concerns the determination of a-tocopherol as well as retinoids, carotenoids, and ubiquinones (Table 6). 

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