Rapid extraction protocols

FIGURE1.4 A comparison of three extraction protocols for the analysis of rat plasma samples for propranolol. (Reprinted from Mallet, C.R. et al., Rapid Commun. Mass Spectrom., 17, 163, 2003. With permission.)... 

Comparison between studies is limited by the use of different extraction protocols, and by differences in signal assignments. Chemical shifts are modified by the chemical environment around the phosphorus nuclei (e.g. sample pH), but two recent studies provide reference data for future research (Makarov et al., 2002 Turner et al., 2003a). They also show that concentrations of phosphate diesters can be underestimated due to the rapid degradation of some compounds, especially RNA and some phospholipids, to phosphate monoesters in alkaline extracts. Further details on solution... 

Standard protocols for rapid extraction of recombinant plasmid DNA from bacterial cultures (6), omitting further purification by equilibrium centrifugation in CsCl-ethidium bromide gradients, give DNA substrates of sufficient purity (see Chapter 9). 

Molinie R, Kwiecien RA, Silvestre V, Robins RJ (2009) Determination of nitrogen-15 isotope fractionation in tropine evaluation of extraction protocols for isotope ratio measurement by isotope ratio mass spectrometry. Rapid Commun Mass Spectrom 23 (Copyright (C) 2012 American Chemical Society (ACS). All Rights Reserved.) 4031 1037. doi 10.1002/ rcm.4344... 

This unit describes procedures for extraction, purification, and identification by MALDI-MS of fiavonol glycosides from a plant source. The extraction and purification protocols are not meant to be comprehensive, but rather to offer guidelines for sample preparation prior to a MALDI-MS analysis. The MALDI-MS technique is suggested as a complement to other analytical methods such as HPLC or NMR. Its strength lies in the ability to rapidly screen a number of samples for the presence of fiavonol glycosides, which can be identified on the basis of their molecular weights. 

A protocol for in vitro transcription used in our laboratory is given below. Rapid plasmid minipreparations made by alkaline lysis without RNase treatment are usually sufficient to prepare DNA templates. DNA samples linearized by restriction digestion are extracted with chloroform, and precipitated with ethanol, and 0.2 to 0.5 /ig of DNA dissolved in 1 pil of distilled water is used per assay. Contamination with RNases must be avoided during the following steps. The reaction mixture for eight reactions is prepared at room temperature to avoid precipitates. 

---