Caffeine Pathways Research Articles

The relationship between coffee-related factors and cortical and hippocampal structure: a triangulation of evidence approach and Mendelian randomization research.

Existing studies have reported sustained changes in the cortical structure of rats due to coffee-related factors, which are speculated to occur in the human body. However, there is a lack of research on this topic. Additionally, previous observational studies have found the impact of diseases on cortical structure and the potential therapeutic effects of coffee on these diseases. Our aim was to study the causal effects of coffee-related factors on the human brain using SNPs (single nucleotide polymorphisms). We will connect these discovered causal effects to the impact of diseases on the brain. Through triangulating evidence, we will reveal the potential active areas of coffee in preventing diseases. We utilized GWAS data from multiple cohorts and their databases, selecting instrumental variables for genetic prediction of coffee intake and plasma levels of caffeine and its direct metabolites. We applied these instrumental variables to individual data on cortical thickness and surface area, as well as hippocampal volume, from the ENIGMA and CHARGE consortium for Mendelian randomization analysis (MR). Triangular evidence was obtained by integrating existing evidence through a specified retrieval strategy, calculating the overlap between coffee's effects on brain regions and disease-related brain regions to identify potential regions of action. The MR analysis yielded 93 positive results for 9 exposures, among which theobromine, a metabolite in the caffeine pathway, was found to be associated with increased hippocampal volume. For cortical structure, theobromine in the caffeine pathway was associated with a decrease in total surface area, while theobromine and caffeine in the pathway were associated with an increase in total thickness. The overlap rate of triangular evidence showed no difference in both overall and subgroup analyses, indicating a high overlap between the effects of coffee on brain regions and disease. From predicted outcomes from causal effects, coffee intake-related factors may have lasting effects on cortical structure. Additionally, theobromine and theophylline have the greatest impact on certain brain gyri, rather than caffeine. Triangulation evidence indicates that disease and coffee intake-related factors act on the same cortical regions, suggesting the presence of potential shared or antagonistic pathways.

New insights into the mechanism of coupled photocatalysis and Fenton-based processes using Fe surface-modified TiO2 nanotube layers: The case study of caffeine degradation

TiO2 nanotube (TNT) layers are prepared via electrochemical anodization of Ti foil in ethylene glycol-based electrolyte containing fluoride ions. Using spin-coating of Fe(III) precursor solutions of different concentrations (10 µM to 10 mM), iron surface-modified TNT (Fe-TNT) layers are formed after annealing at 450 °C for 2 h in air. The Fe-TNT layers are comprehensively characterized by XRD, SEM, DRS and XPS. To identify and assess the contribution of photocatalysis and Fenton-based processes that are simultaneously triggered by Fe-TNT layers, degradation of caffeine is investigated in the dark and under UVA with and without the presence of 500 µM H2O2. It has been found that in the presence of H2O2 under UVA, 100 µM Fe-TNT can degrade 67 % of caffeine after 3 h, of which 4 % is from photolysis, 23.5 % from photocatalysis and 39.5 % from Fenton-based processes. It is important to note that Fenton-based reactions are due to the iron leaching, while the main reactive oxygen species are identified as hydroxyl radicals. In addition, the main degradation by-products are identified by LC-MS and IC-MS techniques, thus leading to the proposal of different degradation pathways of caffeine. To sum up, the research's findings provide novel insights into the concept of photochemical versatility, which is only obliquely explored in the corpus of previous research.

Genetic determinants of liking and intake of coffee and other bitter foods and beverages

Coffee is a widely consumed beverage that is naturally bitter and contains caffeine. Genome-wide association studies (GWAS) of coffee drinking have identified genetic variants involved in caffeine-related pathways but not in taste perception. The taste of coffee can be altered by addition of milk/sweetener, which has not been accounted for in GWAS. Using UK and US cohorts, we test the hypotheses that genetic variants related to taste are more strongly associated with consumption of black coffee than with consumption of coffee with milk or sweetener and that genetic variants related to caffeine pathways are not differentially associated with the type of coffee consumed independent of caffeine content. Contrary to our hypotheses, genetically inferred caffeine sensitivity was more strongly associated with coffee taste preferences than with genetically inferred bitter taste perception. These findings extended to tea and dark chocolate. Taste preferences and physiological caffeine effects intertwine in a way that is difficult to distinguish for individuals which may represent conditioned taste preferences.

Abstract 13039: Putative Protective Effects Of Caffeine And Its Metabolites On Coronary Heart Disease In The Middle East

Introduction: Protective association between coronary heart disease (CHD) and caffeine intake and its metabolism has been reported. Some of the caffeine’s beneficial actions may be attributed to its direct effect on the endothelial cell, stimulating the production of nitric oxide (NO), as well as its role as an adenosine receptor antagonist. In the Middle East and Gulf region CHD is rampant, and caffeine consumption is also relatively high. The interaction of the two is of interest and will be investigated in this study. Hypothesis: Caffeine is a protective factor for CHD in the Middle East. Methods: The Qatar Cardiovascular Biorepository provided 1,001 CHD patients and the Qatar Biobank provided 2,999 control subjects. Metabolites in the caffeine pathway (KEGG, hsa00232) were quantified and curated via liquid chromatography-mass spectrometry by Metabolon Inc. We then subjected data to standard quality control procedures. Association between metabolite levels and CHD was tested by logistic univariate regression, controlling for age, BMI, and sex. Results: The caffeine KEGG pathway contains 22 compounds. Eleven were present in our cleaned metabolomics panel, and all of them were reduced in CHD patients (Figure 1). Five were significant at P <0.05: caffeine ( P =5.26x10 -3 ), 1,7-dimethyluric acid ( P =2.96x10 -4 ), paraxanthine ( P =8.67x10 -3 ), theobromine ( P =2.33x10 -5 ), and 1-methylxanthine ( P =1.17x10 -2 ). Through pathway enrichment analysis, caffeine metabolism pathway was found to be significantly enriched with an FDR P =4.59x10 -34 and an enrichment ratio of 44.23. Conclusions: The finding of lower levels of caffeine and several of its metabolites in CHD patients in this study suggests a potential protective effect for this stimulant. However, this is a cross-sectional cohort study, so we are observing only association at a slice in time.

The Chromosome-Level Reference Genome of Tea Tree Unveils Recent Bursts of Non-autonomous LTR Retrotransposons in Driving Genome Size Evolution

The Chromosome-Level Reference Genome of Tea Tree Unveils Recent Bursts of Non-autonomous LTR Retrotransposons in Driving Genome Size Evolution

Removal of caffeine from water by combining dielectric barrier discharge (DBD) plasma with goethite

In this research, dielectric barrier discharge plasma was developed to cooperate with goethite for removing caffeine in aqueous solution. Goethite was characterized by X-ray diffraction and scanning electron microscopy. The effects of input power, initial concentration and catalysts concentration on the removal efficiency of caffeine were evaluated. Furthermore, the degradation pathways of caffeine were also discussed preliminarily. In the case of caffeine concentration at 50 mg L−1, the degradation efficiency of caffeine was improved from 41% to 94% after 24 min on the conditions of input power of 75 W by combining goethite catalysts (2.5 g L−1), while the energy efficiency could be enhanced 1.6–2.3 times compared to the single DBD reactor. The reaction mechanism experiments demonstrated that attack by hydroxyl radical and ozone was the main degradation process of caffeine in aqueous solution. These studies also provided a theoretical and practical basis for the application of DBD-goethite in treatment of caffeine from water.

Biosynthetic Pathways of Purine and Pyridine Alkaloids in Coffee Plants

Caffeine (1,3,7-N-trimethylxanthine) and trigonelline (IN-methylnicotinic acid) are major alkaloids in coffee plants. The key enzymes involved-in the biosyntliesis of these compounds are very closely related N-methyltransferases belonging to the motif B' family of methyltransferases. The major biosynthetic pathways of caffeine and trigonelline are summarized in this review, including new evidence obtained from recombinant enzymes. In addition, precursor supply pathways are discussed with newly obtained results. Transgenic plants produced by the modification of the expression of N-methyltransferase genes are also introduced.

Transcriptomic and phytochemical analysis of the biosynthesis of characteristic constituents in tea (Camellia sinensis) compared with oil tea (Camellia oleifera).

BackgroundTea plants (Camellia sinensis) are used to produce one of the most important beverages worldwide. The nutritional value and healthful properties of tea are closely related to the large amounts of three major characteristic constituents including polyphenols (mainly catechins), theanine and caffeine. Although oil tea (Camellia oleifera) belongs to the genus Camellia, this plant lacks these three characteristic constituents. Comparative analysis of tea and oil tea via RNA-Seq would help uncover the genetic components underlying the biosynthesis of characteristic metabolites in tea.ResultsWe found that 3,787 and 3,359 bud genes, as well as 4,042 and 3,302 leaf genes, were up-regulated in tea and oil tea, respectively. High-performance liquid chromatography (HPLC) analysis revealed high levels of all types of catechins, theanine and caffeine in tea compared to those in oil tea. Activation of the genes involved in the biosynthesis of these characteristic compounds was detected by RNA-Seq analysis. In particular, genes encoding enzymes involved in flavonoid, theanine and caffeine pathways exhibited considerably different expression levels in tea compared to oil tea, which were also confirmed by quantitative RT-PCR (qRT-PCR).ConclusionWe assembled 81,826 and 78,863 unigenes for tea and oil tea, respectively, based on their differences at the transcriptomic level. A potential connection was observed between gene expression and content variation for catechins, theanine and caffeine in tea and oil tea. The results demonstrated that the metabolism was activated during the accumulation of characteristic metabolites in tea, which were present at low levels in oil tea. From the molecular biological perspective, our comparison of the transcriptomes and related metabolites revealed differential regulatory mechanisms underlying secondary metabolic pathways in tea versus oil tea.Electronic supplementary materialThe online version of this article (doi:10.1186/s12870-015-0574-6) contains supplementary material, which is available to authorized users.

PharmGKB summary

Departments of Genetics, Bioengineering, Stanford University Medical Center, Stanford, California, USA and Division of Clinical Pharmacology, Karolinska Institute, Stockholm, Sweden Correspondence to Dr Teri E. Klein, PhD, Department of Genetics, Stanford University Medical Center, 300 Pasteur D. Lane L301, Mail Code 5120, Stanford, CA 94305-5120, USA Tel: + 1 650 725 0659; fax: + 1 650 725 3863; e-mail: feedback@pharmgkb.org

Current Trends in Producing Low Levels of Caffeine in Coffee Berry and Processed Coffee Powder

Caffeine is a bioactive compound present in coffee. The physiological effects of caffeine on human and animal models are well documented, and this has led the coffee industry to develop an artificially decaffeinated coffee based on solvent extraction process. An alternative to this approach would be to identify naturally occurring low caffeine lines, breeding for low caffeine coffee trait or to generate transgenic coffee with regulated caffeine pathway. This review is mainly focused on caffeine and its health effects; information on decaffeination processes; latest developments in the understanding of caffeine pathway and development of genetically engineered coffee with low caffeine.

Caffeine synthase and related methyltransferases in plants.

Caffeine (1,3,7-trimethylxanthine) is a purine alkaloid present in high concentrations in tea and coffee and it is also found in a number of beverages such as coca cola. It is necessary to elucidate the caffeine biosynthetic pathway and to clone the genes related to the production of caffeine not only to determine the metabolism of the purine alkaloid but also to control the content of caffeine in tea and coffee. The available data support the operation of a xanthosine-->7-methylxanthosine-->7-methylxanthine-->theobromine-->caffeine pathway as the major route to caffeine. Since the caffeine biosynthetic pathway contains three S-adenosyl-L-methionine (SAM) dependent methylation steps, N-methyltransferases play important roles. This review focuses on the enzymes and genes involved in the methylation of purine ring. Caffeine synthase, the SAM-dependent methyltransferase involved in the last two steps of caffeine biosynthesis, was originally purified from young tea leaves (Camellia sinensis). The isolated cDNA, termed TCS1, consists of 1,483 base pairs and encodes a protein of 369 amino acids. Subsequently, the homologous genes that encode caffeine biosynthetic enzymes from coffee (Coffea arabica) were isolated. The recombinant proteins are classified into the three types on the basis of their substrate specificity i.e. 7-methylxanthosine synthase, theobromine synthase and caffeine synthase. The predicted amino acid sequences of caffeine biosynthetic enzymes derived from C. arabica exhibit more than 80% homology with those of the clones and but show only 40% homology with TCS1 derived from C. sinensis. In addition, they share 40% homology with the amino acid sequences of salicylic carboxyl methyltransferase, benzoic acid carboxyl methyltransferase and jasmonic acid carboxyl methyltransferase which belong to a family of motif B' methyltransferases which are novel plant methyltransferases with motif B' instead of motif B as the conserved region.

Induction and accumulation of caffeine in young, actively growing leaves of cocoa (Theobroma cacao L.) by wounding or infection with Crinipellis perniciosa

Crinipellis perniciosa, the causal agent of witches' broom disease, attacks young actively growing shoots, flowers and developing fruits of cocoa (Theobroma cacao L.). Infected shoots become fasciated and a profusion of hypertrophic shoots with small leaves develop from the lateral buds. This is followed by desiccation and death of the infected stem and leaves. Infected stem tissue contains significant amounts of caffeine (109±35μg g−1), approximately seven to eight times greater than healthy stems (14±1μg g−1). Wounding young actively growing leaves induces the production of considerable amounts of caffeine within 1day of treatment, whereas the level of caffeine in older leaves is unaffected by wounding. Caffeine production is stimulated by treatment of young actively growing leaves with salicylic acid (SA) and its synthetic analog, benzothiadiazole, compounds that induce pathogen defense responses in plants. Growth of C. perniciosa is also significantly inhibited on caffeine-containing media. The results suggest that in cocoa, the caffeine pathway is inducible in young actively growing leaves by pathogen attack, wounding and SA, and may be part of the defense response of this species to herbivory and infection.

A new caffeine biosynthetic pathway in tea leaves: utilisation of adenosine released from the S-adenosyl- L-methionine cycle

The four-step caffeine biosynthetic pathway includes three methylation steps that utilise S-adenosyl- L-methionine (SAM) as the methyl donor. In the process SAM is converted to S-adenosyl- L-homocysteine (SAH) which in turn is hydrolysed to L-homocysteine and adenosine. Significant amounts of radioactivity from [methyl- 14C]methionine and [methyl- 14C]SAM were incorporated into theobromine and caffeine in young tea leaf segments, and very high SAH hydrolase activity was found in cell-free extracts from young tea leaves. Substantial amounts of radioactivity from [adenosyl- 14C]SAH were also recovered as theobromine and caffeine in tea leaf segments, indicating that adenosine derived from SAH is utilised for the synthesis of the purine ring of caffeine. From the profiles of activity of related enzymes in tea leaf extracts, it is proposed that the major route from SAM to caffeine is a SAM→SAH→adenosine→adenine→AMP→IMP→XMP→xanthosine→7-methylxanthosine→7-methylxanthine→theobromine→caffeine pathway. In addition, direct adenosine kinase-catalysed formation of AMP from adenosine may participate as an alternative minor route. The activity of two of the three N-methyltransferase activities involved in caffeine biosynthesis and part of the activities of SAH hydrolase, adenosine nucleosidase, adenine phosphoribosyltransferase and adenosine kinase were located in tea chloroplasts. In contrast, no detectable activity of SAM synthetase was associated with the purified chloroplast fraction. This is a first demonstration that the purine skeleton of caffeine is synthesised from adenosine released from the SAM cycle.

Caffeine metabolism in a group of 67 patients with primary biliary cirrhosis.

To evaluate the polygenic regulated caffeine metabolism in a group of 67 patients with a documented primary biliary cirrhosis (PBC) classified according to the histologic stage proposed by Scheuer. Over a 14-year period, drug liver metabolism, using caffeine as a probe drug, has been systematically carried out in addition to the usual clinical, histological and biochemical investigations performed in patients with PBC. The "Caffeine test" consisted of a 200 mg caffeine oral intake. Urines were collected over 24 hours: caffeine (137X), 1-7-dimethylxanthine (17X), 1-3-dimethylxanthine (13X), 1-3-dimethylurate (13U), 3-7-dimethylxanthine (37X), 1-7-dimethylurate (17U), 1-methylxanthine (1X), 1-methylurate (1U), 7-methylxanthine (7X), 3-methylxanthine (3X), and 5-acetylamino-6-formylamino-3-methyluracyl (AFMU) were analyzed by high performance liquid chromatography (HPLC). Total and individual metabolite urinary elimination rates were expressed in micromol/24 hours. Enzyme activities were evaluated from the following urinary metabolite ratios: (AFMU+1U+1X)/17U for CYP1A2, 17U/17X for CYP2A6, AFMU/(AFMU+U+ 1X) for NAT-2, 1U/1X for XO. Compared to healthy subjects, patients with PBC presented a reduced metabolism of caffeine due to a decreased CYP1A2 activity, all the more important since the patients had an advanced histological stage. This picture was nearly identical to the observed picture in chronic liver diseases from various origins. PBC affected the various metabolic pathways of caffeine in a differential manner. CYP1A2 activity was decreased but XO and mainly CYP2A6 activities were increased as shown by the raised urinary ratio 17U/total metabolite elimination. In contrast to the described loss of bimodality of the NAT-2 index distribution in patients with alcoholic cirrhosis, we found a clear-cut, bimodal distribution in patients with PBC, without a high incidence of slow acetylator status. Metabolism of caffeine is strongly and differentially disturbed in patients with PBC and apparently not exactly in the same way as that in alcoholic cirrhosis which is more often taken as an index of chronic liver disease. This suggests the need for caution with medicines whose metabolism is under polygenic regulation. Because of the relationships between caffeine metabolism modifications and histological stages, the caffeine test might be used along with the usual tests to safely follow-up the evolution of the disease.

Relationship between the severity of alcoholic liver cirrhosis and the metabolism of caffeine in 226 patients.

To evaluate the polygenic regulated caffeine metabolism in a group of 226 patients with liver alcoholic cirrhosis classified according to the Child score. Over a 14-year period an hepatic function test, using caffeine as probe drug, has been systematically associated to the usual clinical and biochemical investigations performed in patients with liver alcoholic cirrhosis. "Caffeine test" consisted in a 200 mg caffeine oral intake. Urines were collected over 24 hours: caffeine (137X), 1-7 dimethylxanthine (17X), 1-3 dimethylxanthine (13X), 1-3 dimethylurate (13U), 3-7 dimethylxanthine (37X), 1-7 dimethylurate (17U), 1-methylxanthine (1X), 1-methylurate (1U), 7-methylxanthine (7X), 3-methylxanthine (3X), and 5-acetylamino-6-formylamino-3-methyluracyl (AFMU) were analyzed by high performance liquid chromatography (HPLC). Total and individual metabolite urinary elimination rates were expressed in micromol/24 hours. Enzyme activities were evaluated from the following urinary metabolites ratios: (AFMU+1U+1X)/17U for CYPIA2, 17U/17X for CYP2A6, AFMU/(AFMU+ 1U+1X) for NAT-2, 1U/1X for XO. Compared to healthy subjects, whatever the Child score, caffeine metabolism was reduced by half in patients with alcoholic cirrhosis. The main cause was the decreased CYP1A2 activity. On the other hand, XO and CYP2A6 activities were increased and NAT-2 activity remained unchanged in slow acetylators (SA) and decreased in rapid acetylators (RA) Child B and C. Bimodality of NAT-2 distribution was unclear, but a right assignment of RA and SA phenotype in cirrhotic patients, confirmed by comparison with genotype, was obtained, using the antimode value of NAT-2 distribution used in healthy subjects. At last, there was an interindividual variability in caffeine metabolism as great as in the usual laboratory parameters. Metabolism of caffeine is decreased in patients with alcoholic liver cirrhosis. This decrease paralleled the modifications of the usual laboratory tests and does not bring additional information on the severity of the disease. But the equilibrium between the various metabolic pathways of caffeine is impaired. Beyond the changes of a specific enzymatic activity, this must be taken into account particularly for drugs whose metabolism is of the polygenic regulation type.

Metabolism of Caffeine and Related Purine Alkaloids in Leaves of Tea (Camellia sinensis L.)

Purine alkaloid catabolism pathways in young, mature and aged leaves of tea (Camellia sinensis L.) were investigated by incubating leaf sections with 14C-labelled theobromine, caffeine, theophylline and xanthine. Incorporation of label into CO2 was determined and methanol-soluble metabolites were analysed by high-performance liquid chromatography-radiocounting and thin layer chro-matography. The data obtained demonstrate that theobromine is the immediate precursor of caffeine, which accumulates in tea leaves because its conversion to theophylline is the rate limiting step in the purine alkaloid catabolism pathway. The main fate of [8-14C]theophylline incubated with mature and aged leaves, and to a lesser extent young leaves, is conversion to 3-methylxanthine and onto xanthine which is degraded to 14CO2 via the purine catabolism pathway. However, with young leaves, sizable amounts of [8-14C]-theophylline were salvaged for the synthesis of caffeine via a 3-methylxanthine → theobromine → caffeine pathway. Trace amounts of [2-14C]xanthine were also salvaged for caffeine biosynthesis in young leaves, by conversion to 3-methylxanthine, and this was enhanced in the presence of 5 mM allopurinol which inhibits purine catabolism. Feeds of [2-14C]xanthine to young leaves also indicated that 3-methylxanthine, as well as being salvaged for theobromine and caffeine production, is also converted, via N-l-methyla-tion, to theophylline.

Caffeine biosynthesis in young leaves of Camellia sinensis: In vitro studies on N‐methyltransferase activity involved in the conversion of xanthosine to caffeine

The aim of this study was to investigate the S‐adenosylmethionine dependent N‐methyltransferase(s) (NMT) associated with the three methylation steps in the caffeine biosynthesis pathway in tea (Camellia sinensis L.). NMT activity in cell‐free preparations from young leaves was purified by anion‐exchange and gel‐filtration column chromatography. In both systems, a single zone of NMT activity, with broad substrate specificity was detected. The N‐3 position of dimethylxanthine and monomethylxanthines was methylated more readily than N‐1 while comparatively little substitution occurred at the N‐7 locus. When xanthosine was used as a substrate only the N‐7 position was methylated. These results indicate that a single NMT may participate in the conversion of xanthosine to caffeine. The apparent Mr of the NMT, estimated by gel filtration chromatography, was 61 000. The substrate specificity of the NMT is compatible with the operation of a xanthosine → 7‐methylxanthosine → 7‐methylxanthine → theobromine → caffeine pathway as the main biosynthetic route to caffeine in young tea leaves. The data also indicate that the conversion of 7‐methylxanthine → paraxanthine → caffeine may function as one of a number of minor pathways that also contribute to the production of caffeine.

Biosynthesis of Caffeine in Leaves of Coffee.

The levels of endogenous caffeine and theobromine were much higher in buds and young leaves of Coffea arabica L. cv Kent than in fully developed leaves. Biosynthesis of caffeine from 14C-labeled adenine, guanine, xanthosine, and theobromine was observed, whereas other studies (H. Ashihara, A.M. Monteiro, T. Moritz, F.M. Gillies, A. Crozier [1996] Planta 198: 334-339) have indicated that there is no detectable incorporation of label into caffeine when theophylline and xanthine are used as substrates for in vivo feeds with leaves of C. arabica. The capacity for caffeine biosynthesis, especially from guanine and xanthosine, was reduced markedly in both fully developed mature and aged leaves. Data obtained in pulse-chase experiments with young leaves indicate the operation of an AMP -> IMP -> xanthosine 5[prime]-monophosphate (or GMP -> guanosine) -> xanthosine -> 7-methylxanthosine -> 7-methylxanthine -> theobromine -> caffeine pathway. The data obtained provide strong evidence against proposals by G.M. Nazario and C.J. Lovatt ([1993] Plant Physiol 103: 1203-1210) concerning the independence of caffeine and theobromine biosynthesis pathways and the role of xanthine as a key intermediate in caffeine biosynthesis.

Oxidation of caffeine and related methylxanthines in ascorbate and polyphenol-driven Fenton-type oxidations.

Caffeine and related methylxanthines were subjected to free radical mediated oxidation by incubation with Fe(3+)-EDTA/ascorbate and Fe(3+)-EDTA/polyphenolics. The reaction mixtures were analysed by reverse-phase HPLC, revealing the corresponding C-8 hydroxylated analogues as the major products of hydroxyl radical mediated attack. Further oxidation products of caffeine, analysed by liquid chromatography-mass spectrometry (LC-MS), were the N1-, N3- and N7-demethylated methylxanthine analogues theobromine, paraxanthine and theophylline, respectively. Isolable amounts of the imidazole ring operated 6-amino-5-(N-formylmethyl-amino)-1,3-dimethyl-uracil (1,3,7-DAU) derivative were also detected, which was characterised by 1H NMR and mass spectroscopy. The identified products indicate that the pertinent chemical reactions, i.e. C-8 hydroxylation, demethylations, and C8-N9 bond scission, are comparable to the primary metabolic pathways of caffeine in humans. The influence of pH, transition metals, hydrogen peroxide, free radical scavengers and metal chelators on caffeine oxidation was studied. This report illustrates that natural food-borne reactants can aid in identifying specific chemical markers of free radical induced damage. Furthermore, potentially anti-and pro-oxidative reactions can be elucidated which may be important in assessing the impact of nutrient additives and supplements on the shelf life and stability of foods and beverages.

Effect of pregnandiol on caffeine metabolism in female rats.

Three groups of six 5-week-old Sprague Dawley female rats received i.p. injections of pregnandiol, 1.25, 2.50 or 5 mg/kg, respectively, in triolein daily for 7 days. Caffeine metabolism was studied in liver slices on day 8 by HPLC. Only primary metabolites were formed. N-1 demethylation was the most important pathway (theobromine represented 51% of total dimethylxanthines). Unlike in human in vitro or in vivo, 1,3,7-DAU (6-amino-5-(N-formylmethylamino)-1,3-dimethyluracil) was an important metabolite (9.7% of total caffeine metabolites). Pregnandiol inhibited N-1, N-3 and N-7 demethylation in vitro (-33%, -33% and -28%, respectively, at 5 mg/kg/day), but it had no effect on N-1 demethylation at 1.25 or 2.50 mg/kg/day. Pregnandiol at all doses had no effect on 1,3,7-trimethyluric acid and 1,3,7-DAU formation. These results are consistent with the hypothesis that C-8 hydroxylation and demethylation of caffeine are mediated by different isoenzymes. They indicate that pregnandiol is a potent inhibitor of microsomal drug metabolism, specifically of cytochrome P450 IA, which could explain the immaturity of some metabolic pathways of caffeine in neonates.