Tobacco alkaloids, analysis

The first group of alkaloids that was subjected to gas chromatographic analysis was the very volatile tobacco alkaloids. They were gas chromatographed by Quin in 1958 on packed columns with polyglycols as stationary phases. Since then a great number of papers have been published of tobacco alkaloids in connection with... 

Bush determined the four most important tobacco alkaloids (nicotine, nornicotine, anabasine and anatabine) using a 10 % DC 550 packed column on Chromosorb 60-80, and using isoquinoline as an internal standard. The alkaloids were extracted with benzene-chloroform (9 1) after treatment of the tobacco sample (1 g) with bariumhydroxide and water. The organic phase was concentrated and used for the gas chromatographic determination. Because of the great differences in the amounts of the minor alkaloids and nicotine in most tobacco samples, two extractions, each with an appropriate amount of internal standard, were required for a complete assay. The precision of the quantitative analysis on tobacco samples of different alkaloids is given in Table 5.6. 

PRECISION OF QUANTITATIVE ALKALOID ANALYSIS ON TOBACCO SAMPLES OF DIFFERENT ALKALOID COMPOSITION15... 

Analysis of the major tobacco alkaloids has been performed on an octadecyl column using a triethylamine - phosphoric acid buffer (pH 7.25) to which 40 methanol was added (Fig.3.2) (24). The method was compared with a GLC method. Several extraction solvents were compared as to their effectiveness. An 0.025 M phosphate buffer (pH 7.8) was found to be suitable. 

A series of reviews describing nicotine metabolism has recently appeared [2], Specific topics covered include the biosynthesis and metabolism of nicotine and related alkaloids [3], an overview of mammalian nicotine metabolism [4], the role of cytochrome P450 in nicotine metabolism [5], nicotine metabolism beyond cotinine [6], N-oxidation, A -methylation, and N-conjugation reactions of nicotine [7], extrahepatic metabolism of nicotine and related compounds [8], metabolism of the minor tobacco alkaloids [9], analysis and levels of nicotine and metabolites in body fluids [10], kinetics of nicotine and its metabolites in animals [11], pharmacokinetics of (S)-nicotine and metabolites in humans [12], and sources of inter-individual variation in nicotine pharmacokinetics [13]. Another recent review described variables which affect nicotine metabolism [14]. Several compilations of studies or reviews on the tobacco-specific A-nitrosamines are available [15-18]... 

Severson, R.F., R.F. Arrendale, K.L. McDuffie, and O.T. Chortyk A rapid GC-2 method for tobacco alkaloids 33rd Tobacco Chemists Research Conference, Program Booklet and Abstracts, Vol. 33, Paper No. 6, 1979, p. 3. Severson, R.F., R.F. Arrendale, K.L. McDuffie, and O.T. Chortyk A modified method for the rapid analysis of long-chained alkanes and neophytadiene from tobacco Beitr. Tabakforsch. Int. 11 (1981) 27-32. 

This chapter is not intended to include a detailed description of all possible methods of alkaloid analysis in tobacco nor provide a comprehensive review of the literature. Certain methods of historical or practical significance have been highlighted with the most pertinent papers cited for each method. The methods of alkaloid analysis discussed are applicable to all classes of tobacco. 

The chemistry of pyridine alkaloids has resulted in many different methods for determining flue-cured tobacco alkaloids qualitatively and quantitatively. Some of these older methods are rather laborious and time-consuming, but may be applicable in the absence of modern techniques. The major alkaloid of interest in flue-cured tobacco is nicotine, which comprises 95% of the total alkaloids in domestic cultivars therefore, it is only necessary to determine total alkaloids as nicotine in most cases. Most laboratories currently use methods for alkaloid analysis which are rapid, sensitive, and provide more dependable data. The introduction of preparative chromatographic techniques and sophisticated spectroscopic instrumentation has led to a dramatic increase in the number of known alkaloids. 

Pyridine alkaloids are high boiling compounds which are stable to high temperatures. Nicotine, the most volatile of the pyridine alkaloids, is readily steam distilled from strong basic solutions however, secondary alkaloids are less volatile than nicotine and are not easily steam distilled quantitatively from basic aqueous solutions. Since nicotine is the predominant pyridine alkaloid present in most tobaccos, this analysis suffices for total alkaloids because secondary alkaloid concentrations are minimal. Tobacco alkaloids removed from basic aqueous solutions by steam distillation are quantified with a spectrophotometer. 

Gas chromatographic analysis of tobacco alkaloids does not require derivation. The general procedures for GC analyses are as follows (1) use the smallest sample applicable for analysis (2) utilize preparations that clean up the sample without loss of alkaloids (3) pre-extract the sample prior to alkaloid extraction with hexane (removes pigments and lipids) (4) extract alkaloids from tobacco with an aqueous acid solution, filter, partition the aqueous acid extract with organic solvent (hexane, methylene chloride, chloroform, etc.) (5) increase pH of the aqueous extract to pH 10 (5N NaOH) and partition the basic solution with chloroform (6) dry the chloroform solution over Na2S04, and (7) concentrate the sample or analyze as is. 

Sadler WW, Chesson RR, Schoenbaum AW (1960) Automated procedure for determining the nicotine content of steam distillates. Tob Sci 4 208-212 Saunders JA, Blume DE (1981) Quantification of major tobacco alkaloids by high-performance liquid chromatography. J Chromatogr 205 147-154 Severson RF, McDuffie KL, Arrendale RF, Gwynn GR, Chaplin JF, Johnson AW (1981) Rapid method for the analysis of tobacco nicotine alkaloids. J Chromatogr 211 111-121... 

Nonvolatile Nitrosamines In Tobacco. A method which we developed several years ago for the analysis of tobacco-specific nitrosamines (TSNA 31) involves extraction of tobacco with buffered ascorbic acid TpH 4.5) followed by partition with ethyl acetate, chromatographic clean-up on silica gel, and analysis by HPLC-TEA (Figure 9). Results obtained with this method for a large spectrum of tobacco products (Table IV), strongly support the concept that the levels of nitrate and alkaloids, and especially the methods for curing and fermentation, determine the yields of TSNA in tobacco products. Recent and as yet preliminary data from snuff analyses indicate that aerobic bacteria play a role in the formation of TSNA during air curing and fermentation. 

NT391 Izman, G. V., N. A. Sherstyannykh, NT402 and L. G. Astaghova. Analysis of the tobacco haploids, produced by culturing anthers in vitro, in regards to their content of alkaloids and volatile acids. 

The laboratory of the Government Chemist in London undertakes the analysis of tobacco smokes in order to produce league tables (Fig. 20) of the tar and nicotine content of all cigarette brands available in the UK, on behalf of the British government. In this context tar is defined as the total particulate matter present in the main-stream smoke (inhaled smoke), adjusted for its content of water and nicotine alkaloids. Nicotine is defined as the total nicotine-type alkaloids present in the mainstream smoke both are expressed in mg per cigarette. The tar and nicotine league tables are published every six months, and include details of approximately 130 brands of cigarette. 

Quin2,3 was the first to apply gas chromatography for the analysis of nicotine alkaloids in tobacco smoke. He used three sets of conditions to analyse the many alkaloids present in a wide boiling range. 

Quantitative analysis of the alkaloids in tobacco was reported by Piade and Hoffmann. ... 

A new photometric determination of volatile bases in tobacco and tobacco smoke in terms of nicotine, which compares quantitatively with mass spectral and g.c. methods, has been developed.27 A colorimetric method for the estimation of nicotine alkaloids in tobacco by reaction with cyanogen bromide and 4,4 -di-aminostilbene-2,2 -disulphonic acid has also been reported.28 Dilute sulphuric acid extraction of nicotine and anabasine from autopsy tissue appears to be a more efficient method than extraction with acidified ethanol, aqueous oxalic acid, and steam distillation.29 Thin-layer chromatography has been effective in the analysis of nicotine and other alkaloids and drugs.30 Two reports on the isolation of anabasine from anabasine-lupinine mixtures have appeared.31 The P a values of some nicotine-type compounds have been determined.32... 

USE Preserving foods, fats, fruit juices, alkaloidal solns, etc manuf benzoates and benzoyl compds, dyes as a mordant in calico printing for curing tobacco. As standard in volumetric and calorimetric analysis. Pharmaceutic aid (antifungal). 

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