PubMed · 14704460
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Matthew J Powell, Aaron T Timperman. 2004. Proteome analysis.. https://doi.org/10.1385/1-59259-742-4%3A387
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A validated high performance liquid chromatographic method was developed for the determination of chromium picolinate in pharmaceutical dosage forms. The analysis was performed at room temperature using a reversed-phase Supelcosil LC-18 (250 x 4.6 mm, 5 microm) column. The mobile phase consisted of acetonitrile:water (40:60 v/v) at a fl ow rate of 0.8 mL/min. The UV-detector was set at 264 nm. The developed method showed a good linear relationship in the concentration range from 0.125 to 12.5 microg/mL with a correlation coefficient from 0.999. The limit of detection and limit of quanti fi cation were 0.091 and 0.181 microg/mL, respectively.
This chapter enables the reader to carry out the solid-phase synthesis of ribonucleic acid (RNA) using beta-cyanoethyl phosphoramidite chemistry combined with tert-butyldimethylsilyl protection of the ribose 2'-hydroxyl group. Phosphoramidite monomers are activated with 5-benzylmercapto-1H-tetrazole enabling fast and highly efficient coupling to the 5'-hydroxyl group of the support-bound oligonucleotide. On completion of the synthesis, the stepwise deprotection of the nucleobase, phosphate, and ribose protecting groups is carried out using optimized protocols. Subsequently the various high-pressure (performance) liquid chromatography (HPLC) procedures are described enabling the purification and analysis of the RNA. For this purpose anion-exchange and reversed-phase HPLC are used singly or in combination according to the final purity requirement of the RNA.
A protocol for the straightforward preparation of small circular oligodeoxyribonucleotides (2-28 nt) is reported. The assembly of the oligonucleotide chain (standard phosphoramidite chemistry) and cyclization by the phosphotriester method take place on a tailor-made nucleotide-derivatized solid support. Although cyclization yields are moderate, the procedure exploits a synthesis design that allows selective cleavage of the circular oligonucleotide from the support, which facilitates isolation of the target molecule by simple filtration.