Showing papers on "Theobromine published in 2007"

The structure of two N-methyltransferases from the caffeine biosynthetic pathway

Abstract: Caffeine (1,3,7-trimethylxanthine) is a secondary metabolite produced by certain plant species and an important component of coffee (Coffea arabica and Coffea canephora) and tea (Camellia sinensis). Here we describe the structures of two S-adenosyl-l-methionine-dependent N-methyltransferases that mediate caffeine biosynthesis in C. canephora ‘robusta’, xanthosine (XR) methyltransferase (XMT), and 1,7-dimethylxanthine methyltransferase (DXMT). Both were cocrystallized with the demethylated cofactor, S-adenosyl-l-cysteine, and substrate, either xanthosine or theobromine. Our structures reveal several elements that appear critical for substrate selectivity. Serine-316 in XMT appears central to the recognition of XR. Likewise, a change from glutamine-161 in XMT to histidine-160 in DXMT is likely to have catalytic consequences. A phenylalanine-266 to isoleucine-266 change in DXMT is also likely to be crucial for the discrimination between mono and dimethyl transferases in coffee. These key residues are probably functionally important and will guide future studies with implications for the biosynthesis of caffeine and its derivatives in plants.

Influência do método de extração nos teores de metilxantinas em erva-mate (Ilex paraguariensis a. St.-Hil., aquifoliaceae)

Abstract: Ilex paraguariensis A. St. -Hil. is a native species of southern South America. The caffeine content in Ilex paraguariensis leaves has been considered an important quality parameter for mate-derived products. In this work different extraction methods for the determination of these methylxantines are compared. The influence of the extraction conditions on the methylxanthine yields was evaluated. Extraction by decoction with acidic aqueous solution (H2SO4 4N) presented the higher efficiency in the theobromine extraction. The extraction in a Soxhlet with acidic aqueous solution and decoction with acidic aqueous solution showed the highest caffeine yield. For the concomitant theobromine and caffeine quantification, the decoction with acidic aqueous solution is suggested.

Effect of acute administration of hydroalcohol extract of Ilex paraguariensis St Hilaire (Aquifoliaceae) in animal models of Parkinson's disease

Abstract: Ilex paraguariensis St Hilaire (Aquifoliaceae) is a plant widely cultivated in South America and with various reputed medicinal properties that can be attributed to phenolic constituents of the leaves: caffeine, theophylline and theobromine, besides the flavonoids, quercetin and rutin. This study examined the antiparkinsonian activity of the hydroalcohol extract of Ilex paraguariensis in models of protection against cerebral injury induced by MPTP and reversal of the catatonia induced by reserpine in mice. The hydroalcohol extract prevented MPTP-induced hypolocomotion at doses of 250 and 500 mg/kg at the all time points observed and also prevented the reserpine-induced catalepsy at the same doses. The extract potentiated the effect of apomorphine in preventing catatonia, suggesting a non-dopaminergic activity, probably through antagonism of adenosine. In biochemical studies the hydroalcohol extract caused a significant decrease in the NO levels, exhibited a DPPH-scavenging ability and was effective in preventing the oxidation of deoxyribose. The results obtained suggest that the hydroalcohol extract of Ilex paraguariensis may have an antiparkinsonian profile in animal models, probably through its antioxidant activity and antagonist action on adenosine A2A receptors. Copyright © 2007 John Wiley & Sons, Ltd.

Determination of theobromine, theophylline, and caffeine in by-products of cupuacu and cacao seeds by high-performance liquid chromatography.

Abstract: Theobromine, theophylline, and caffeine are determined simultaneously by a rapid and selective reversed-phase high-performance liquid chromatography (HPLC) method with UV detection in by-products of cupuacu and cacao seeds. The determination is carried out in the raw and roasted ground cupuacu seeds and in the corresponding powders obtained after pressure treatment. The by-products of both cupuacu seeds and cacao seeds are obtained under the same technological conditions. The HPLC method uses isocratic elution with a mobile phase of methanol-water-acetic acid (80:19:1) (v/v) at a flow rate of 1 mL/min and UV absorbance detection at 275 nm. Total elution time for these analytes is less than 10 min, and the detection limit for all analytes is 0.1 mg/g. The amounts of theobromine and caffeine found in all the cupuacu samples are one or more orders of magnitude lower than those from cacao. Theophylline is found in all cacao samples except for the roasted ground paste, and it is only found in the roasted ground paste in the cupuacu samples.

Determination of caffeine in coffee products by dynamic complexation with 3,4-dimethoxycinnamate and separation by CZE.

Abstract: A method based on the formation of pi-complexes with chlorogenate-like species was proposed for the determination of caffeine in regular (nondecaffeinated) and decaffeinated coffee. Both caffeate and 3,4-dimethoxycinnamate were able to transform caffeine--a neutral species in aqueous solutions--into an anionic species. The usage of 3,4-dimethoxycinnamate in the running electrolyte is advantageous, because of its greater chemical stability and the improved resolution of the peaks of caffeine, theobromine, and theophylline. Negative peaks were registered with a capacitively coupled contactless conductivity detector when solutions of these alkylxanthines were analyzed with a BGE composed of 20 mmol/L 3,4-dimethoxycinnamic acid and pH adjusted to 8.5 with Tris. This behavior was expected, because the complex is larger and thus should move slower than the free anion. Caffeine was determined in ground and instant coffee with precision and accuracy that meet Brazilian norms about such products. The LOD was estimated as 33 mg/L, which corresponds to 0.8 and 0.3 mg of caffeine per gram of dry instant coffee and ground coffee, respectively. For the case of decaffeinated coffee, ten times preconcentration with dichloromethane was carried out to allow the quantitation of caffeine, which should not exceed the concentration of 1 mg/g in dry matter.

Transdermal and buccal delivery of methylxanthines through human tissue in vitro.

Abstract: We examined the in vitro permeation of central nervous stimulants - caffeine, theophylline, and theobromine across human skin with the aid of six chemical enhancers. It was found that oleic acid was the most potent enhancer for all three methylxanthines. Further optimization studies with different solvents showed that caffeine transport could be enhanced to give flux values up to 585 microg/cm2.hr-1. Theobromine and theophylline delivery rates proved insufficient. An additional study involving a buccal tissue equivalent showed that this membrane was more permeable than skin for all model actives tested and would offer an alternate way of delivery.

Topical application of plant extracts containing xanthine derivatives can prevent UV-induced wrinkle formation in hairless mice.

Abstract: Background: Plants are the source of important products with nutritional and therapeutic value. Topical or oral administration of some plant extracts has been shown to reduce photodamage. Cacao bean and cola nut are popular edible plants that contain polyphenols and xanthine derivatives. These plant extracts possess protective effects against UV-induced erythema when taken orally, and an H2O2-scavenging effect. Methods: Plant extracts containing xanthine derivatives and three xanthine derivatives were topically applied to the dorsal skin of hairless mice, and the mice were exposed to a resemblance of solar ultraviolet irradiation at a dose of 13.0 J/cm2 (UVA) for 15 weeks, five times a week on weekdays. After the final irradiation, histological, and analytical studies were performed. Results: Topical application of plant extracts (cacao beans, cola nuts) and caffeine, theobromine, and theophylline markedly prevented photodamage including wrinkle formation and histological alterations. A significant increase in total hydroxyproline content caused by UV irradiation was observed. In contrast, topical application of plant extracts and xanthine derivatives reduced total hydroxyproline and pepsin-resistant hydroxyproline content in comparison with that of the control (vehicle, UV-irradiation group). Moreover, naphthol AS-D chloroacetate esterase staining and diaminobenzidine staining suggested that leukocytes including neutrophils increased in the UV-exposed skin. In contrast, weak staining was observed in skin treated with xanthine derivatives. Conclusion: Topical application of plant extracts and xanthine derivatives suppressed wrinkle formation, dermal connective alteration, and collagen accumulation. It is suggested that xanthine derivatives prevented neutrophil infiltration caused by UV-irradiation.

Selective liquid CO2 extraction of purine alkaloids in different Ilex paraguariensis progenies grown under environmental influences

Abstract: In this investigation, liquid carbon dioxide at 20 degrees C and 150 bar was used for the selective extraction of caffeine and theobromine from dry leaves of mate. A comparison between the chromatograms from CO2 extraction and traditional solvent extraction supports the selectivity of carbon dioxide for these purine alkaloids. The advantages of selective liquid CO2 extraction in terms of speed and resolution of UV/HPLC is also evidenced. A randomized block design of experiments was proposed to investigate the influence of 16 progenies of Ilex paraguariensis grown in 3 diverse sites on the contents of caffeine and theobromine in liquors of mate leaves obtained by extraction with compressed CO2. A significant effect of both these factors on the parameters investigated was observed by involving the F distribution in the statistical analysis. A cluster analysis based on the experimental uncertainties in the contents of these two methylxanthines has identified from four to six different groups of mate progenies.

Degradation Kinetics of Caffeine and Related Methylxanthines by Induced Cells of Pseudomonas sp.

Abstract: In this study, the kinetics of degradation of caffeine and related methylxanthines by induced cells of Pseudomonas sp. was performed. The kinetics data showed that degradation of caffeine, theobromine, and 7-methylxanthine followed Michealis–Menten kinetics. The values of Km are low for caffeine and 7-methylxanthine and high for theobromine. Degradation of caffeine and theobromine was enhanced in the presence of NADH and NADPH, whereas the degradation of 7-methylxanthine was unaffected. Among the various metal ions tested, Fe2+ was found to enhance the rate of degradation for all three substrates, whereas Zn2+ and Cu2+ inhibited the degradation of caffeine and theobromine but not 7-methylxanthine. The differences in kinetic parameters and cofactor requirement suggest the possibility of the involvement of more than one N-demethylases in the caffeine catabolic pathway in Pseudomonas sp. The induced cells can serve as effective biocatalysts for the development of biodecaffeination techniques.

A Kinetic Comparison on the Inhibition of Adenosine Deaminase by Purine Drugs

Abstract: The effects of allopurinol, acyclovir and theophylline on the activity of adenosine deaminase (ADA) were studied in 50 mM sodium phosphate buffer pH 7.5 at 27 °C, using a UV– Vis spectrophotometer. Adenosine deaminase is inhibited by these ligands, via different types of inhibition. Allopurinol, as a transition state analog of xanthine oxidase, and acyclovir competitively inhibit the catalytic activity of ADA. Inhibition constant values are 285 and 231 µM for allopurinol and acyclovir, respectively. Theophylline acts as a non-competitive inhibitor for ADA, which shows different affinity binding sites at various drug concentrations. There were two different types of inhibition constant, one of them due to a low concentration of the drug (K i = 56 μM) and the other appearing at higher concentrations of theophylline (K i = 201 μM). Thermodynamic parameters also show that ADA has two binding sites for theophylline. The comparison of inhibition constant for inosine (K i = 143 μM) and acyclovir (K i = 231 μM) elucidates the critical role of the ribose ring within the inosine structure, relative to the open ring of acyclovir. Comparison of the inhibition constant of theobromine (K i = 311 μM) with inosine (K i = 143 μM) shows the critical binding role of N 7 position within the purine ring. Interestingly, the N 7 position in allopurinol is replaced by a CH 2 group, which demonstrates the lower inhibiting potency of allopurinol (K i = 285 μM) relative to inosine (K i = 143 μM). In a structural sense, a comparison made between the structure of theophylline and theobromine besides a comparison between the inhibition constant of theophylline (K i = 56 μM at low and 201 μM at higher concentrations) and caffeine (K i = 342 μM) indicate that substitution of a bulky group in N 1 and N 7 positions of purine has a critical role in the binding affinity of the above- mentioned inhibitors to the enzyme.

Easy preparative scale syntheses of labelled xanthines: caffeine, theophylline and theobromine

Abstract: Several easy preparative scale (0.5–1.5 g) syntheses of deuterium labelled caffeine, theophylline and theobromine are described. Some new selective syntheses of theophylline and theobromine have been developed. Labelled xanthines are of great interest in qualitative or quantitative isotope dilution-mass spectrometry, coupled with gas or liquid chromatography, currently performed in anti-doping and forensic laboratories. Copyright © 2007 John Wiley & Sons, Ltd.

Biotransformation of Caffeine to Value Added Products

Abstract: Caffeine is a naturally occurring molecule belonging to the xanthine alkaloid family and is present in tea, coffee over 60 other plant species. It is also available as a by-product of decaffeination process, coffee and tea processing wastes and by chemical synthesis. The world production of coffee is 6.795 million tons and the world decaffeinated coffee market is about 15% of the total coffee market. About 2 lakh tons of caffeine is generated per year from decaffeination of tea and coffee. Besides this, lots of caffeine is available from unutilized coffee processing wastes such as coffee pulp and hull. Every ton of coffee, when processed, generates 40% processing wastes in the form of coffee pulp and coffee hull. These processing wastes contain caffeine ranging from 1.1-2.2%, which can be extracted efficiently and used as substrates for useful biotransformation where caffeine is converted into potent therapeutic molecules leading to value addition. Biotransformation of caffeine and bioconversion of the wastes to safe products is an effective solution to the above problems. Caffeine extracted from these wastes can be biotransformed by using suitable caffeine biotransforming microbial cultures to valuable methyl xanthines such as theophylline, theobromine and paraxanthine, which are potent therapeutic molecules. Production of theophylline by biotransformation of caffeine will obviate the disadvantages of chemically synthesized theophylline and lead to utilization of abundantly available caffeine and value addition. The processing wastes of caffeine such as coffee pulp and coffee hull are considered as major agro-industrial wastes, as they are associated with several anti-physiological and inhibitory factors like high content of caffeine, tannins and polyphenols. Efficient methods of utilization or disposal of these wastes are still not available and are therefore disposed by the processing units and left unused, causing pollution of the nearby water bodies and soil. Effective utilization of these processing wastes is required not only to save the environment, but also utilize it for the benefit of the producers, which would generate revenues in the lean season and generate extra employment. Further, microbial cultures can also be used for the decaffeination of the processing wastes and their efficient utilization. The removal or minimization of the anti nutritional factors in these processing wastes makes them suitable to have alternative applications in animal feed, organic fertilizer, as a substrate in solid state fermentation, biogas production, edible mushroom production and vermiculture. Work reported in the thesis involves the isolation, screening and characterization of caffeine utilizing fungal cultures. The most potent fungal culture, which could biotransform caffeine to theophylline, was characterized as Penicillium citrinum and used for the production of theophylline. The caffeine biotransformation pathway of P. citrinum was elucidated and the enzymes responsible for biotransformation of caffeine were identified. Optimization of parameters for caffeine biotransformation by the selected microorganism was done. Work was also carried out towards the efficient utilization of coffee processing wastes and spent coffee. Proposed objectives of the thesis: 1. Isolation, screening and characterization of microbial cultures for biotransformation. 2. Identification and characterization of biotransformed products. 3. Studies on the enzymes involved in the biotransformation of caffeine. 4. Optimization of parameters for growth and caffeine biotransformation by selected microorganism. 5. Utilization of coffee processing wastes and spent coffee.

Genotype characterization of cocoa into genetic groups through caffeine and theobromine content predicted by near infra red spectroscopy

Abstract: Purine compounds such as caffeine and theobromine are involved in cocoa flavour development. Measurement of these compounds is used to determine the percentage of cocoa mass in chocolate. Additionally the relative level of these two compounds may be associated with the genetic and/or geographical origin of the cocoa. The present work was undertaken to assess the potential of NIRS, as a rapid and non-destructive method for measuring these compounds. The study was carried out on 284 cocoa samples over 5 production years (1999 to 2003). The samples came from Ivory Coast, Venezuela and Trinidad, and belonged to genotypes of the Forastero, Criollo and Trinitario genetic groups. The sampling protocol ensured good representation of the spectral and chemical variability of fermented and dried cocoa. Spectra acquisition was perform on 3 g of shelled ground and sieved beans in diffuse reflectance using a FOSS Nirsystem 6500. Partial Least Square models (PLS) were used to establish quantitative relations between NIR spectral data and caffeine and theobromine contents. Models were developed which fitted the data with coefficients of determination of 0.95 for caffeine content and 0.89 for theobromine content. The SECV errors were 0.04% for caffeine and 0.07% for theobromine. These values are close to the repeatability of the reference method. The ratios of SD to SECV were 3.5 and 2.6 for caffeine and theobromine respectively. The theobromine to caffeine ratio separated Ivorian Forastero cocoa from the Trinitario cocoa from Trinidad and the Criollo cocoa from Venezuela. Given the performance of the equations developed to represent the range of samples over spectral variability, NIRS can be considered to provide a routine analysis method for purine compounds. Whilst this study did not lead to definitive conclusions on discrimination between genetic groups according to their purine compositions, it did confirm earlier observations. With a tool such as NIRS that enables determination of purine composition, it shall be possible to make progress in that direction in the foreseeable future. [Resume d'auteur]

Determination of caffeine and caffeine-related metabolites in ephedra-containing standard reference materials using liquid chromatography with absorbance detection and tandem mass spectrometry.

Abstract: The concentrations of caffeine and caffeine-related compounds in 2 ephedra-containing reference materials have been determined by 3 independent methods with measurements performed by the National Institute of Standards and Technology (NIST) and a collaborating laboratory. Results from the 3 methods were used for value assignment of caffeine, theobromine, and theophylline in these Standard Reference Materials (SRMs). The methods used at NIST to determine the concentration levels of caffeine, theobromine, and theophylline in SRM 3243 Ephedra-Containing Solid Oral Dosage Form and SRM 3244 Ephedra-Containing Protein Powder used reversed-phase liquid chromatography with absorbance detection and tandem mass spectrometry. These reference materials are part of the first suite in a series of NIST SRMs that provide concentration values for multiple components in dietary supplements. These SRMs are primarily intended for method validation and for use as control materials to support the analysis of dietary supplements and similar materials.

Using composition containing cAMP (cyclic adenosine monophosphate), nicotinic acid, prostaglandin, gamma linolenic acid and xanthines in weight loss and/or reducing elevated triglyceride

Abstract: An using composition containing cAMP (Cyclic Adenosine Monophosphate), Nicotinic Acid, Prostaglandin, Gamma Linolenic Acid as effective ingredient in management of weight reduction or hypertriglyceridemia, furthermore disclose of certain natural products are essentially contain of said ingredients are to be used for such purpose. For incidence, coffee is the source of caffeine, tea is the source of theophylline, and cocoa is the source of theobromine. Zizyphus jujube mill contains rich amount of cAMP, Radix Angelica Sinensis, Sunflower, Matrimony vine ( Lycium chinense Mill), Edible fig ( Ficus carica ) are sources of Nicotinic Acid, Evening primrose, Starflower contain Gamma Linolenic Acid which are indirect sources of Prostaglandin. This invention discloses the random combination use of the above composition is useful and effective in the management of weight reduction or reducing elevated triglyceride.

한국산 보성 덖음 녹차의 가공 및 저장중의 카테킨류의 변화

Abstract: This study investigated changes in the catechin and alkaloid contents of Bosung green tea during different manufacturing processes and storage periods, using HPLC and UV-VIS spectrophotometer analysis. For changes in the catechin and alkaloid contents by roasting technique, we found that EGCG, CG and GCG decreased just slightly by processes performed before roasting, rather than after roasting. In addition, theobromine, caffeine, and ECG changed minimally throughout all the processes. For changes in the catechin and purine alkaloid contents of the green tea leaves during storage, EGCG, ECG, and CAF decreased considerably in the green tea stored at temperatures of 5℃ and 25℃, and they decreased by 20～30% after storage for 1 year. However, a quantitative difference was hardly observed in the catechin and alkaloid contents regardless of storage temperature.

Untersuchungen zur Pharmakokinetik der Methylxanthine Theophyllin und Theobromin hinsichtlich der Dopingrelevanz beim Pferd

Abstract: Aim of this study was to optimize and validate a suitable method for the analysis detection and quantification of theophylline, theobromine and the metabolites paraxanthine and caffeine in blood and urine samples of horses and to investigate the pharmacokinetic behaviour of caffeine and theobromine. Validation and measurement of samples was made by using a high performance liquid chromatograph (HPLC) with connected tandem-mass-spectrometer. Validation contained the criteria selectivity, linearity, accuracy, precision, stability and recovery. The limit of quantification for theophylline, theobromine, paraxanthine and caffeine was fixed at 100 ng/ml in plasma and at 50 ng/ml in urine for theophylline, paraxanthine and caffeine. For theobromine it was fixed at 100 ng/ml. The limit of detection was found at 13 ng/ml for theophylline, at 15 ng/ml for theobromine, at 14 ng/ml for caffeine and at 29 ng/ml for paraxanthine in plasma. In urine the limit of detection was found at 20 ng/ml for theophylline, at 26 ng/ml for theobromine, at 33 ng/ml for caffeine and at 32 ng/ml for paraxanthine. Theophylline was administered to six horses at a single intravenous dose of 4 mg/kg body weight, theophylline at a single oral dose of 4 mg/kg body weight, or theobromine at a single intravenous dose of 4 mg/kg body weight. Maximum concentrations of theophylline in plasma after intravenous administration ranged between 5165 and 8834 ng/ml. Maximum theophylline concentrations in urine ranged between 21636 and 93373 ng/ml. 60 hours after administration urine concentrations of all six horses were not below limit of quantification. In plasma and urine the metabolites paraxanthine and caffeine were detectable. But caffeine concentrations were mainly around limit of detection. Theobromine was not detectable. Maximum concentrations of theophylline in plasma after oral administration ranged between 1791 and 3483 ng/ml. Maximum concentrations of theophylline in urine ranged between 16365 and 41256 ng/ml. 108 hours after administration the urine concentration of only one horse was below limit of quantification. In plasma and urine the metabolite paraxanthine was detectable. After administration of theobromine its maximum concentrations in plasma ranged between 5715 and 12402 ng/ml and in urine between 65771 and 94889 ng/ml. 168 hours after administration all urine concentrations of the six horses were still above limit of detection. No metabolites could be detected in plasma and urine. Using the pattern of metabolites the parent substance can be concluded. If there is detected almost sole theobromine in plasma or urine, application of theobromine can be submitted, and if there is detected theophylline and paraxanthine in plasma or urine, application of theophylline can be submitted. Afterwards the results were integrated into a pharmacokinetic/pharmacodynamic approach of TOUTAIN and LASSOURD (2002) to calculate the irrelevant plasma and urine concentrations where caffeine has no effect on the organism anymore. Calculation of effective plasma concentration, irrelevant plasma concentration and irrelevant urine concentration by the pharmacokinetic/pharmacodynamic model of TOUTAIN and LASSOURD (2002) showed that for theophylline the irrelevant plasma concentration (at 14 ng/ml) and urine concentration (at 40 ng/ml) at a dosing interval of 12 hours are below respectively at limit of detection. Therefore it is concluded that any detection of theophylline in plasma or urine by this method has to be considered as a positive result of dopingtest. That is why an establishment of a selected cut-off value for therapeutics seems not to be acceptable for theophylline. In addition a considerable variance of measured data demonstrates the interindividual variability. It must be concluded that a prediction about excretion times of theophylline and theobromine based on this data by a small number of probands is not reliably possible. Any detection of theophylline in plasma or urine samples should be considered as relevant for doping and should be punished appropriately.