DNA, hydrolysates

Figure 2. Chromatographic profile of DNA hydrolysate from rat liver 12 h after application of 133 mg/18.5 mCi/kg l l-[2f3- H]-nitrosodipropylamine. Key O, A , CPAf. Conditions included column, 30-cm i rBondapak-C18 eluant, 3% methanol/0.05M ammonium formate (pH 3.5) flow rate, 1 mL/min and sample, 10 mg hydrolyzed DNA containing unlabeled 7-propylguarune (41 fig/mg DNA) and 74sopropylguanine (34 fig/mg DNA), which was injected five times in five...

Erhlich and Erhlich (El) have reported on the separation of uracil, 5-methylcytosine, cytosine, guanine, and thymine using reversed-phase ion pairing (Fig. 5) and also, on the separation of adenine and N -methyladenine from the five DNA bases listed above. Although the separation was reported using only reference compounds, it has potential for the analysis of DNA hydrolysates. 

In biological samples, the frequency of drug-induced base modification is generally very low. When the immunoassay uses polymeric DNA, it is particularly important that cross-reaction with unmodified DNA is minimal, because it is not possible to chromatographically separate modified from unmodified nucleotides before the analysis. This can be done when working with DNA hydrolysates... 

Roberts et al. [76] reported the detection of etheno-dC adducts in erode DNA hydrolysates on the order to 5 adducts in 10 nucleobases using 100 pg DNA. The use of on-line affinity LC (Ch. 1.4.2) allowed a 100-fold improvement in detection limits compared to a conventional Cig-based SPE approach. 

I. Hoes, E.L. Esmans, Analysis of melphalan adducts of 2 -deoxynucleotides in calf thymus DNA hydrolysates by capillary LC-ESI-MS-MS, J. Chromatogr. B, 736 (1999)43. 

DNA hydrolyses promoted by di- and multi-nuclear metal complexes 04CCR(248)147. 

Need to verify that radioactivity is not due to contamination or metaboUsm, i.e., to avoid possible artifacts, HPLC of DNA hydrolysates and digestion with proteases and ribonucleases can be used. 

Figure 1. About 5 mg of human liver DNA hydrolysates was fractionated by reverse phase HPLC system. Each fraction was screened with respect, to the r ctivity.with specific monocloial antibodies against 0°-MetG, 0°-EtG, O -MstT or O -Et-T. Ihe arrows indicate the position of the corresponding authentic...

Hydrolyzed DNA Synonyms DNA hydrolysate Definition Hydrolysate of DNA derived by acid, enzyme or other method of hydrolysis Uses Biological additive, skin conditioner, hair conditioner in cosmetics Hydrolyzed elastin... 

Immediately prior to derivatization, the dried DNA hydrolysate is treated (in the original hydrolysis tube) with 10/rl of dry pyridine and dried in vacuo to remove any trace of water by azeotrope formation. The derivatizing reagent (10 fil of a 1 1 solution of dry pyridine and MTBSTFA containing 1% TBDMCS) is added to the dried sample and the tube is flame-sealed and heated for... 

Depending on the modification, physical separation and detection methods for modified bases/nucleosides/nucleotides in DNA hydrolysates can already or may eventually provide greater specificity and sensitivity than immxmological methods. However, it seems that antibodies recognizing adducts in polymeric DNA provide the only foreseeable basis for specific quantification of certain types of DNA modifications in individual cells and for isolation of DNA fiag-ments carrying those modifications. 

PEI-cellulose has been extensively studied and used in the separation of nucleic acid bases, nucleosides, and nucleotides, with good separation and resolution. It is also used for the separation of RNA and DNA hydrolysates, and large scale preparations among other applications. We feel this remains the most versatile layer for separations of dNMPs. 

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