Methylxanthines analysis

Pentoxifylline is stmcturaHy related to other methylxanthine derivatives such as caffeine [58-02-2] (1,3,7-trimethylxanthine), theobromine [83-67-0] (3,7-dimethylxanthine), and theophylline [58-55-9] (3,7-dihydro-1,3-dimethyl-1 H-piirine-2,6-dione or 1,3-dimethylxanthine), which also show radioprotective activity in some instances, suggesting that methylxanthines as a dmg class may radioprotect through a common mechanism (see Alkaloids). In a retrospective analysis of cervical and endometrial cancer patients receiving primary or adjuvant XRT, no association between caffeine consumption and incidence of acute radiation effects has been found. However, there was a decreased incidence of severe late radiation injury in cervical cancer patients who consumed higher levels of caffeine at the time of thek XRT (121). The observed lack of correlation between caffeine consumption and acute radiation effects is consistent with laboratory investigations using pentoxifylline. 

The analysis of the methylxanthines (caffeine, theobromine, and theophylline) is important in the areas of nutrition and clinical chemistry. These three compounds compose the majority of the alkaloids present in coffee, tea, cocoa, cola nuts, and guarana. 

This chapter on analysis of methylxanthines is divided into three sections historical methods, current analytical methods for foods, and current methods for biological samples which can include plasma, blood, urine, cell extracts, and other potential samples of biological significance. 

The most basic method for the determination of the methylxanthines is ultraviolet (UV) spectroscopy. In fact, many of the HPLC detectors that will be mentioned use spectroscopic methods of detection. The sample must be totally dissolved and particle-free prior to final analysis. Samples containing more than one component can necessitate the use of extensive clean-up procedures, ajudicious choice of wavelength, the use of derivative spectroscopy, or some other mathematical manipulation to arrive at a final analytical measurement. A recent book by Wilson has a chapter on the analysis of foods using UV spectroscopy and can be used as a suitable reference for those interested in learning more about this topic.1... 

As was indicated, there have not been a large number of publications on the CE analysis of methylxanthines in food systems. Analusis published a method using a 20-mm Borate buffer at pH 9.6 and UV detection at 254 nm and +22 kV applied voltage.41 Samples were diluted and prepared for analysis by filtration and analyzed using free solution electro-... 

The various methods that have been outlined in the previous sections are not exclusive and other analytical methods have been used for the determination of methylxanthines in food systems. One of the most widely used methods for food analysis is flow injection analysis (FIA). In a study by Numata,43 a flow injection analysis method for the determination of hypoxanthine in meat was described. 

Analytical studies in clinical chemistry relating to the determination of methylxanthines are concentrated in two areas. The first of these involves the analysis of various ethical pharmaceuticals. The second area involves the analysis of various body fluids for methylxanthines and their metabolites. 

In the clinical area, the largest share of analytical methods development and publication has centered on the determination of theophylline in various body fluids, since theophylline is used as a bronchodilator in asthma. Monitoring serum theophylline levels is much more helpful than monitoring dosage levels.44 Interest in the assay of other methylxanthines and their metabolites has been on the increase, as evidenced by the citations in the literature with a focus on the analysis of various xanthines and methylxanthines. 

As in the case in the analysis of food samples, the introduction of relatively inexpensive MS detectors for GC has had a substantial impact on the determination of methylxanthines by GC. For example, in 1990, Benchekroun published a paper in which a GC-MS method for the quantitation of tri-, di-, and monmethylxanthines and uric acid from hepatocyte incubation media was described.55 The method described allows for the measurement of the concentration of 14 methylxanthines and methyluric acid metabolites of methylxanthines. In other studies, GC-MS has also been used. Two examples from the recent literature are studies by Simek and Lartigue-Mattei, respectively.58 57 In the first case, GC-MS using an ion trap detector was used to provide confirmatory data to support a microbore HPLC technique. TMS derivatives of the compounds of interest were formed and separated on a 25 m DB-% column directly coupled to the ion trap detector. In the second example, allopurinol, oxypurinol, hypoxanthine, and xanthine were assayed simultaneously using GC-MS. 

Chocolate milk samples prepared from sweetened cocoa powders averaged 58 mg per serving of theobromine and 5 mg per serving of caffeine.28 Analysis of a "home-style" recipe resulted in higher methylxanthine values — 94 mg theobromine and 10 mg caffeine per serving. However, the authors noted that this recipe also had a stronger chocolate flavor. The lower values reported by Zoumas et al. and Blauch and Tarka compared to others was attributed to the inability of older methods to separate theobromine and caffeine, and the lack of precision and accuracy of the older methods. A compendium of theobromine and caffeine values reported for chocolate beverages from both published and unpublished studies has been compiled in Table 8. 

It is necessary to determine the methylxanthine content of chocolate foods, as well as beverages, in order to obtain an accurate assessment of the total amount of theobromine and caffeine that is ingested via the diet. This area of analysis has received little attention, and only scant published data exist on the methylxanthine content of chocolate foods. 

Chemical analysis of the finished food product is a more accurate determination of the methylxanthine content. In studies performed at Hershey Foods Corporation, the methylxanthine content of a large variety of commercially available chocolate foods was measured by HPLC methods.38 These results have been compiled together with other literature values in Table 11. Large methylxanthine variations can be seen among the chocolate foods, as well as within different brands of the same item. 

Stein, M., Krasowski, M. Leventhal, B., Phillips, W. and Bender, B., Behavioral and cognitive effects of methylxanthines A meta-analysis of theophyliine and caffeine. Arch Pediatr Adolesc Med 150(3), 284-288, 1996. 

Barr RG, Rowe BH, Canargo CA Jr. Methylxanthines for exacerbations of chronic obstructive pulmonary disease meta-analysis of randomised trials. BMJ 2003 20 327 643. 

Fig. 2 HPLC analysis of eight methylxanthines (a) with isocratic elution (b) with gradient elution.

To improve the theophylline analysis, Bailey et al. used 3-isobutyl-l-methylxanthine as... 

Fig. 11.8. Analysis of theophylline in the presence of caffeine and its metabolites1 Precolumn Lichrosorb RP2 (45x2.0 mm ID), column Ultrasphere ODS 5 urn (150x4.6 mm ID), mobile phase 0.01 M sodium acetate and 0.005 M tetrabutylammonium hydrogen sulfate in water (pH 4.75 with 0.1 M acetic acid or sodium hydroxide) to which 12.5% methanol is added, flow rate 1.5 ml/min, detection UV 274 nm. Peaks 1, uric acid 2, 3-methyl xanthine 3, 1-methylxanthine ...

Extubation from mechanical ventilation as soon as possible is essential to minimize the toxicity of pressure and oxygen to lungs. Methyixanthines have been shown to facilitate successful extubation. In a meta-analysis of the benefits of methyixanthines, neonates weighing less than 1000 g were able to remain extubated more often if a methylxanthine was used concurrently with CPAP. ... 

Blanchard, J. Weber, C.W. Shearer, L.E. HPLC analysis of methylxanthines in human breast milk. J.Chromatogr.Sci., 1990, 28, 640-642... 

The major analytes of coffee include caffeine, chlorogenic acids, and flavor and volatile aromatic components. The major analytes in cocoa are me-thylxanthines, mainly theobromine and trace amounts of caffeine, cocoa fat, and lipids. Other analytes of interest in cocoa are tannins, pigments, and aroma components. The major analytes of tea are the methylxanthine alkaloids, including caffeine and theophylline, polyphenols (catechins, tannins, and related flavanols), and volatile and aromatic components. Analysis of black tea would also include theaflavins and thearubigens, which are oxidation and condensation products of polyphenols. 

With regard to alkaloids analysis, theobromine was the main compound found in cocoa extracts. This is in agreement with this compound being the most important methylxanthine found in products from the cocoa tree, Theobroma cocao [22], Theobromine was also detected at higher concentrations than caffeine. These were around 42 and 3.5 mg/g lyophilized cocoa extracts (beans, nibs, liquor, and powder) [28],... 

Laboratory evaluation. The stomach cxmtenls, serum, or urine can be analyzed for the presence alkaloids. The alkaline methylxanthines partition into the acidic stomach, so even washings of an empty stomach may be useful for analysis. 

Santos, J. R. and A. O. S. S. Rangel. 2012. Development of a chromatographic low pressure flow injection system Application to the analysis of methylxanthines in coffee. 

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