Barbiturates, identification

Figure 11.19 SPME-CE analysis of urine samples (a) blank urine (a) directly injected and extracted for (b) 5 (c) 10 and (d) 30 min (b) Urine spiked with barbiturates, extracted for (e) 30 and (f, g) 5 min. Peak identification is as follows 1, pentobaitibal 2, butabarbital 3, secobarbital 4, amobarbital 5, aprobarbital 6, mephobarbital 7, butalbital 8, thiopental. Concenti ations used are 0.15-1.0 ppm (e, f) and 0.05-0.3 ppm (g). Reprinted from Analytical Chemistry, 69, S. Li and S. G. Weber, Determination of barbiturates by solid-phase microexti action and capillary electrophoresis, pp. 1217-1222, copyright 1997, with permission from the American Chemical Society.

Narhi LO, AJ Fulco (1987) Identification and characterization of two functional domains in cytochrome P-450g, j.3, a catalytically self-sufficient monooxygenase induced by barbiturates in Bacillus megaterium. J Biol Chem 262 6683-6690. 

B27. Broughton, P. M. G., A rapid ultraviolet spectrophotometric method for the detection, estimation and identification of barbiturates in biological material. Biochem. J. 63, 207-215 (1956). 

S2S. Street, H. V., Gas-liquid chromatography of submicrogram amounts of drugs. IV. Identification of barbiturates, hydantoins, amides, imides, carbamates, phenylbutazone, carboxylic acids and hydrazine derivatives by direct derivative fonnation within the gas chromatograph. J. Chromaiogr. 41, 358-366 (1969). 

Spectrophotometric methods have traditionally been used for the analysis of barbiturates, but these procedures are time consuming, do not identify the barbiturate in question, and it is difficult to quantify accurately unless this identity is known. For these reasons, the development of thin layer and gas chromatographic methods for barbiturate analysis has been occurring. Jain and Cravey (32) have reviewed all the different types of methods being used for the identification of barbiturates from biological specimens. 

Finkle et al. (46) have established a GC/MS reference data system for the identification of drugs of abuse. The data include phenethylamine derivatives, opiate and synthetic narcotics, barbiturates, and urinary metabolites. These data have been established for use with the gas chromatographic retention time index previously developed. 

R. Gill, A. H. Stead, and A. C. Moffat, Analytical aspects of barbiturate abuse Identification of drugs by the effective combination of gas-liquid, high-performance liquid and thin-layer chromatographic techniques, J. Chromatogr., 204 215 (1981). 

H. H. Maurer, Identification and differentiation of barbiturates, other sedative-hypnotics and their metabolites in urine integrated in a general screening procedure using computerized gas chroma-tography-mass spectrometry, J. Chromatogr., 530 307 (1990). 

Butyl derivatives make possible the resolution of compounds the methylation of which lead to the same derivatives [518]. They were prepared in an analogous manner, by injection of substrates with a 25% methanolic solution of tetra-n-butylammonium hydroxide into the injection port heated at 270°C. The analysis can be performed on 3% OV-17 at 220°C Fig. 5.31 illustrates an example of the separation of several barbiturates in the form of methyl and butyl derivatives. Mephobarbital and phenobarbital, which being methylated give rise to the same compound, could be resolved after their conversion into butyl derivatives in addition, the different retention times of the derivatives could be utilized for the identification of barbiturates. 

Narhi, L. O., and Fulco, A. J. 1987. Identification and Characterization of 2 Functional Domains in Cytochrome-P-450bm-3, a Catalytically Self-Sufficient Monooxygenase Induced by Barbiturates in Bacillus megaterium. J. Biol. Chem., 262, 6683-6690. 

Chemical derivatives may be used to improve the chromatographic characteristics of polar compounds, e.g. acids may be converted to their methyl esters and barbiturates to their methyl derivatives. This technique may also provide information concerning the number of active sites in a molecule by noting the increase in molecular weight on methylation. In many cases the mass spectrum of tiie derivative may be of more use for identification purposes. 

An example of the use of NMR in forensic science is the identification of four 5-ethyl-5-alkyl substituted barbiturates, butobarbitone, pentobarbitone, secbutobarbitone, and amylobarbitone. The proton spectra of all four barbiturates show a very broad signal at around 5 8 ppm to 5 10 ppm which is characteristic of amido protons. The useful region of the spectrum for the differentiation of these barbiturates lies between 5... 

To understand the identification and quantification techniques used to analyse barbiturates and benzodiazepines. 

By employing this system, it is possible to analyse and identify barbiturates in casework samples, as illusttated in Figure 9.1 for the analysis of phenobarbitone. If the retention time and mass specttal data obtained for the standard and the components of the drugs sample are the same (as shown in this case), then an identification can be called. 

R. Gill, A. C. Moffat, R. M. Smith, and T. G. Hurdley, A collaborative study to investigate the retention reproducibility of barbiturates in HPLC with a view to establishing databases for drug identification, 7. Chromatogr. Sci. 24 (1986), 153-159. 

Barbiturates, rotenone and piericidin block electron transfer from the FeS centres to ubiquinone-10, but do not block the reduction of artificial acceptors by NADH in Type II NADH dehydrogenase. Both rotenone [282,286] and piericidin [307] bind to the enzyme with an apparent 1 1 stoicheiometry to the content of FMN [39]. The binding of these inhibitors is non-covalent which has so far prevented identification of the binding site(s). All EPR- or optically detectable redox centres (possibly with the exception of centre N-la) are reducible by NADH in the presence of these inhibitors [308,309]. The structural analogy between piericidin and Q-10 provides further evidence that these inhibitors interact at the point where the enzyme delivers reducing equivalents to ubiquinone (Fig. 3.14). 

HPLC has found considerable application within the pharmaceutical laboratory. For example, Roos [248] described a HPLC method for the separation, identification and quantitative determination of barbiturates in pharmaceutical dosage forms. The separation of 16 barbiturates was described and the effects of varying experimental parameters discussed. The successful separation of two cinchona alkaloids, four opium alkaloids, and heroin-related narcotics with measurements at nanogram to microgram levels has been achieved [249]. Quantitation and identification of morphine in opium in three samples was described by Wittwer [250]. 

Elisabeth, I. Rene, S. Dieter, J. Screening for drugs in clinical toxicology hy high-performance liquid chromatography Identification of barbiturates by post-column ionization and detection by a multiplex photodiode array spectrophotometer. J. Chromatogr. 1988, 428, 369. 

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