Chlorogenic acid compounds from coffee are differentially absorbed and metabolized in humans.

Chlorogenic acids (CGA) are abundant phenolic compounds in coffee, with caffeoylquinic (CQA), feruloylquinic (FQA), and dicaffeoylquinic (diCQA) acids being the major subclasses. Despite the potential biopharmacological properties attributed to these compounds, little is known about their bioavailability in humans. In this study, we evaluated the distribution profile of the major CGA isomers and metabolites in plasma and urine of 6 healthy adults for 4 h after brewed coffee consumption. Three CQA isomers and 3 diCQA isomers were identified in the plasma of all subjects after coffee consumption, whereas 2 FQA were identified in only 1 subject. Two plasma concentration peaks were observed, the first at 0.5-1.0 h and the second at 1.5-4.0 h after coffee consumption. The molar ratio CQA:diCQA was 12.2 in the brewed coffee, whereas in plasma it ranged from 0.6-2.9. The molar ratios 5-CQA:3-CQA and 5-CQA:4-CQA were consistently higher in plasma than in the brew. The main CGA metabolites identified in urine after coffee consumption were: dihydrocaffeic, gallic, isoferulic, ferulic, vanillic, caffeic, 5-CQA, sinapic, rho-hydroxybenzoic, and rho-coumaric acids (gallic and dihydrocaffeic acids being the major ones). This study indicates that the major CGA compounds present in coffee are differentially absorbed and/or metabolized in humans, with a large inter-individual variation. Moreover, urine does not appear to be a major excretion pathway of intact CGA compounds in humans.

Chlorogenic acids and lactones in regular and water-decaffeinated arabica coffees.

The market for decaffeinated coffees has been increasingly expanding over the years. Caffeine extraction may result in losses of other compounds such as chlorogenic acids (CGA) and, consequently, their 1,5-gamma-quinolactones (CGL) in roasted coffee. These phenolic compounds are important for flavor formation as well as the health effects of coffee; therefore, losses due to decaffeination need to be investigated. The present study evaluates the impact of decaffeination processing on CGA and CGL levels of green and roasted arabica coffees. Decaffeination produced a 16% average increase in the levels of total CGA in green coffee (dry matter), along with a 237% increase in CGL direct precursors. Different degrees of roasting showed average increments of 5.5-18% in CGL levels of decaffeinated coffee, compared to regular, a change more consistent with observed levels of total CGA than with those of CGL direct precursors in green samples. On the other hand, CGA levels in roasted coffee were 3-9% lower in decaffeinated coffee compared to regular coffee. Although differences in CGA and CGL contents of regular and decaffeinated roasted coffees appear to be relatively small, they may be enough to affect flavor characteristics as well as the biopharmacological properties of the final beverage, suggesting the need for further study.

Effects of the combination of hydrophobic polypeptides, iso-alpha acids, and malto-oligosaccharides on beer foam stability.

The influence of hydrophobic polypeptides concentrated in beer foam, together with the composition of iso-alpha acids and the content of malto-oligosaccharides in beer on foam stability, has been investigated. The objective was to find out whether a shortage of one of these positive contributors to foam stability could be compensated for by an increased presence of another or whether optimum levels of each contributor is necessary. For that purpose, an image analysis method to evaluate beer foam quality was developed. The foam collapse time was the parameter chosen to group beers according to their foam stability. Profiles of hydrophobic polypeptides that concentrate in beer foam, iso-alpha acids, and malto-oligosaccharides of 14 beer brands were acquired by high-performance liquid chromatography. Principal component analysis (PCA) was performed to show the relationship between beer brands and its composition. Beers that contained propylene glycol alginate as a foam enhancer showed high foam stability except for one beer, which had a low content of hydrophobic polypeptides, thereby highlighting the requirement of threshold levels of hydrophobic polypeptides to obtain stable foam. The data of samples that were devoid of a foam additive were subjected to a discriminant statistical analysis. Foam stability declined in proportion to decreases in hydrophobic polypeptides and to a lesser extent to decreases in iso-alpha-acid contents. Apparently, the content of malto-oligosaccharides were found to have no major influence on foam stability. The model of discriminate analysis was found to explain 100% of the variance in data with 85.2% success in classifying all samples according to the model, suggesting that foam stability is mainly governed by the beer constituents evaluated in this study.

Contribution of chlorogenic acids to the iron-reducing activity of coffee beverages.

The iron-reducing activity of coffee beverages was determined by the ferric reducing antioxidant power (FRAP) assay. The influence on FRAP due to the degree of roasting (light, medium, and dark), species (Coffea arabica and Coffea robusta), and caffeine content (regular and decaffeinated) was investigated using ground and soluble coffee samples. The concentration of specific chlorogenic acids and caffeine in the beverages was determined by high-performance liquid chromatography and related to FRAP using Pearson correlation coefficients. All measurements were expressed per unit of soluble solids. Beverages prepared with ground coffee had, on average, 27% higher FRAP values than those prepared with soluble coffee (p < 0.05). In the former beverages, FRAP of C. robusta samples was significantly higher (on average, 50.3%) when compared to that of C. arabica samples, and FRAP values decreased with increasing degree of roasting (p < 0.05). A strong correlation (r > 0.91) was found between FRAP and the total content of chlorogenic acids, particularly that of the caffeoylquinic acid isomers. The iron-reducing activity of coffee beverages was not influenced by caffeine.

Effect of roasting on the formation of chlorogenic acid lactones in coffee.

Of all plant constituents, coffee has one of the highest concentrations of chlorogenic acids. When roasting coffee, some of these are transformed into chlorogenic acid lactones (CGL). We have studied the formation of CGL during the roasting of coffee beans in Coffea arabica cv. Bourbon; C. arabicacv. Longberry; and C. canephora cv. Robusta. Individual CGL levels were determined by comparison of HPLC peaks with those of synthetic CGL standards. Seven CGL were identified: 3-caffeoylquinic-1,5-lactone (3-CQL), 4- caffeoylquinic-1,5-lactone (4-CQL), 3-coumaroylquinic-1,5-lactone (3-pCoQL), 4-coumaroylquinic-1,5-lactone (4-pCoQL), 3-feruloylquinic-1,5-lactone (3-FQL), 4-feruloylquinic-1,5-lactone (4-FQL), and 3,4-dicaffeoylquinic-1,5-lactone (3,4-diCQL). 3-CQL was the most abundant lactone in C. arabica and C. canephora, reaching peak values of 230 +/- 9 and 254 +/- 4 mg/100 g (dry weight), respectively, at light medium roast ( approximately 14% weight loss). 4-CQL was the second most abundant lactone (116 +/- 3 and 139 +/- 2 mg/100 g, respectively. The maximum amount of CGL represents approximately 30% of the available precursors. The relative levels of 3-CQL and 4-CQL in roasted coffee were reverse to those of their precursors in green coffee. This suggests that roasting causes isomerization of chlorogenic acids prior to the formation of lactones and that the levels of lactones in roasted coffee do not reflect the levels of precursors in green coffee.

Beta-carotene storage, conversion to retinoic acid, and induction of the lipocyte phenotype in hepatic stellate cells.

Hepatic stellate cells (HSCs) are the major site of retinol (ROH) metabolism and storage. GRX is a permanent murine myofibroblastic cell line, derived from HSCs, which can be induced to display the fat-storing phenotype by treatment with retinoids. Little is known about hepatic or serum homeostasis of beta-carotene and retinoic acid (RA), although the direct biogenesis of RA from beta-carotene has been described in enterocytes. The aim of this study was to identify the uptake, metabolism, storage, and release of beta-carotene in HSCs. GRX cells were plated in 25 cm(2) tissue culture flasks, treated during 10 days with 3 micromol/L beta-carotene and subsequently transferred into the standard culture medium. beta-Carotene induced a full cell conversion into the fat-storing phenotype after 10 days. The total cell extracts, cell fractions, and culture medium were analyzed by reverse phase high-performance liquid chromatography for beta-carotene and retinoids. Cells accumulated 27.48 +/- 6.5 pmol/L beta-carotene/10(6) cells, but could not convert it to ROH nor produced retinyl esters (RE). beta-Carotene was directly converted to RA, which was found in total cell extracts and in the nuclear fraction (10.15 +/- 1.23 pmol/L/10(6) cells), promoting the phenotype conversion. After 24-h chase, cells contained 20.15 +/- 1.12 pmol/L beta-carotene/10(6) cells and steadily released beta-carotene into the medium (6.69 +/- 1.75 pmol/ml). We conclude that HSC are the site of the liver beta-carotene storage and release, which can be used for RA production as well as for maintenance of the homeostasis of circulating carotenoids in periods of low dietary uptake.

Hepatic stellate cells uptake of retinol associated with retinol-binding protein or with bovine serum albumin.

Retinol is stored in liver, and the dynamic balance between its accumulation and mobilization is regulated by hepatic stellate cells (HSC). Representing less than 1% total liver protein, HSC can reach a very high intracellular retinoid (vitamin-A and its metabolites) concentration, which elicits their conversion from the myofibroblast to the fat-storing lipocyte phenotype. Circulating retinol is associated with plasma retinol-binding protein (RBP) or bovine serum albumin (BSA). Here we have used the in vitro model of GRX cells to compare incorporation and metabolism of BSA versus RBP associated [(3)H]retinol in HSC. We have found that lipocytes, but not myofibroblasts, expressed a high-affinity membrane receptor for RBP-retinol complex (KD = 4.93 nM), and both cell types expressed a low-affinity one (KD = 234 nM). The RBP-retinol complex, but not the BSA-delivered retinol, could be dislodged from membranes by treatments that specifically disturb protein-protein interactions (high RBP concentrations). Under both conditions, treatments that disturb the membrane lipid layer (detergent, cyclodextrin) released the membrane-bound retinol. RBP-delivered retinol was found in cytosol, microsomal fraction and, as retinyl esters, in lipid droplets, while albumin-delivered retinol was mainly associated with membranes. Disturbing the clathrin-mediated endocytosis did not interfere with retinol uptake. Retinol derived from the holo-RBP complex was differentially incorporated in lipocytes and preferentially reached esterification sites close to lipid droplets through a specific intracellular traffic route. This direct influx pathway facilitates the retinol uptake into HSC against the concentration gradients, and possibly protects cell membranes from undesirable and potentially noxious high retinol concentrations.

Use of oral contraceptives blunts the calciuric effect of caffeine in young adult women.

Caffeine consumption increases the urinary excretion of calcium and other minerals. Factors that affect caffeine metabolism such as steroid hormones may modify this effect. The purpose of this study was to evaluate the influence of oral contraceptive (OC) use on the 4-h urinary excretion of calcium, phosphorus, magnesium, zinc, sodium, potassium and caffeine metabolites in response to a high caffeine dose given as coffee beverage. Adult women, 20-29 y, users (+OC, n = 15) and nonusers (-OC, n = 15) of oral contraceptives, with calcium intake approximately 500 mg/d, participated in two tests, caffeine load (5 mg/kg body weight) and no-caffeine control, in a randomized crossover design. The net increase (caffeine load corrected by no caffeine) in urinary excretion of most minerals was significantly higher in -OC than in +OC (P < 0.05), with the larger group difference for calcium (ninefold) followed by magnesium (twofold), zinc (onefold) and potassium (onefold). Net increases in urinary excretion of 1-methylurate and paraxanthine were about three- and fivefold higher, respectively, in -OC than in +OC (P < 0.05) whereas net increases in urinary excretion of 5-acetylamino-6-formylamino-3-methyluracil (AFMU) and 1,7-dimethylurate were over twofold higher in the +OC than in -OC (P < 0.05). Following the caffeine load, most urinary minerals showed negative correlation with urinary 1-methylurate in -OC (R

Changes of sphingolipid species in the phenotype conversion from myofibroblasts to lipocytes in hepatic stellate cells.

Sphingolipids play a relevant role in cell-cell interaction, communication, and migration. We studied the sphingolipid content in the murine hepatic stellate cell line GRX, which expresses the myofibroblast phenotype, and can be induced in vitro to display the fat-storing phenotype. Lipid modifications along this induction were investigated by labeling sphingolipids with [(14)C]galactose, [(14)C]serine, or [(14)C]choline, and determination of fatty acid composition of sphingomyelin. The total ganglioside content and the GM2 synthase activity were lower in myofibroblasts. Both phenotypes presented similar gangliosides of the a-pathway: GM2, GM1, and GD1a as well as their precursor GM3. Sphingomyelin and all the gangliosides were expressed as doublets; the upper/lower band ratio increased in lipocytes, containing more long-chain fatty acids in retinol-induced lipocytes as compared to the insulin/indomethacin induced ones. Time-course experiments indicated a transfer of metabolic precursors from phosphatidylcholine to sphingomyelin in the two phenotypes. Taken together, these results indicate that myofibroblast and lipocytes can use distinct ceramide pools for sphingolipid synthesis. Differential ganglioside expression and presence of the long-chain saturated fatty acids suggested that they may participate in formation of distinct membrane microdomains or rafts with specific functions on the two phenotypes of GRX-cells.

Mathematical method for the prediction of retention times of fatty acid methyl esters in temperature-programmed capillary gas chromatography.

An accurate method for identification of fatty acids in complex mixtures analyzed by temperature-programmed capillary gas chromatography is described. The method is based on a mathematical approach using regression curves obtained by plotting the relative retention times of fatty acid methyl esters (FAMEs) analyzed in isothermal and gradient temperature conditions. The method was applied to a complex biological sample (human milk), and it was possible to identify 64 fatty acids, including branched-chain and other fatty acids for which reference standards were not readily available. The identities of the majority of the peaks were confirmed by mass spectrometry. The relative residuals and the relative differences between estimated and measured relative retention times of individual FAMEs varied from 0.03 to 3.15% and from 0.0 to 2.9%, respectively. The method is useful for identification of fatty acids in routine analysis.

Acyl-CoA: retinol acyltransferase (ARAT) and lecithin:retinol acyltransferase (LRAT) activation during the lipocyte phenotype induction in hepatic stellate cells.

We have examined retinol esterification in the established GRX cell line, representative of hepatic stellate cells, and in primary cultures of ex vivo purified murine hepatic stellate cells. The metabolism of [3H]retinol was compared in cells expressing the myofibroblast or the lipocyte phenotype, under the physiological retinol concentrations. Retinyl esters were the major metabolites, whose production was dependent upon both acyl-CoA:retinol acyltransferase (ARAT) and lecithin:retinol acyltransferase (LRAT). Lipocytes had a significantly higher esterification capacity than myofibroblasts. In order to distinguish the intrinsic enzyme activity from modulation of retinol uptake and CRBP-retinol content of the cytosol in the studied cells, we monitored enzyme kinetics in the purified microsomal fraction. We found that both LRAT and ARAT activities were induced during the conversion of myofibroblasts to lipocytes. LRAT induction was dependent upon retinoic acid, while that of ARAT was dependent upon the overall induction of the fat storing phenotype. The fatty acid composition of retinyl-esters suggested a preferential inclusion of exogenous fatty acids into retinyl esters. We conclude that both LRAT and ARAT participate in retinol esterification in hepatic stellate cells: LRAT's activity correlates with the vitamin A status, while ARAT depends upon the availability of fatty acyl-CoA and the overall lipid metabolism in hepatic stellate cells.

Quality control of beer hopped with reduced isomerised products

The traditional method for measuring bitterness in beer by UV absorbance (ASBC9.6) remains a viable quality control method for normally hopped beer. However, after reduction of isoalpha acids specific absorptivity of active components change and HPLC analysis showns to be a reliable method for quality control of these materials. This work is intended to adapt mathematically the ASBC(American Society of Brewing Chemist) spectrophotometrical method and to evaluate an HPLC method to control composition of different commercial reduced isomerised products added to unhopped wort and partially kettle hopped beer. The results we obtained showed that the HPLC method is reliable for quality control of reduced isomerised products and that the traditional spectrophotometrical method is viable for quality control of beer, also using reduced isomerised products as far as an adequate factor is used.

Efeito da maltagem e da extrusäo termoplástica sobre a qualidade tecnológica da cultivar de soja Br 16 / Effect of malting and thermoplastic extrusion on the technologic quality of the derived product of soybean

Sementes de soja cultivar BR 16 foram submetidas à maltagem por 24 e 48 horas e à extrusao obtendo-se flocos, os quais foram moídos e utilizados na fabricaçao de biscoitos. Flocos e biscoitos foram analisados sensorialmente. O efeito da maltagem e da extrusao foram também avaliados em relaçao às características químico-nutricionais. A composiçao em macro nutrientes nao apresentou alteraçoes acentuadas mostrando pequeno decréscimo nos teores de carboidratos totais e discreto aumento relativo nos teores de carboidratos totais e discreto aumento relativo nos teores de lipídeos no produto maltado. Dentre os fatores antinutricionais, os inibidores de tripsina apresentaram queda de 50 por cento após 24h de maltagem e desaparecimento após a extrusao. Os teores de ácido fítico nao sofreram variaçao expressiva, enquanto que os alfa-galactosídeos apresentaram decréscimo de até 80 por cento com os processos combinados de maltagem e extrusao.

Application of high performance liquid chromatography to the analysis of some non-volatile coffee components

High performance liquid chromatography (HPLC) was applied to the analysis of caffeine, trigonelline, nicotinic acid and sucrose in Arabica an Robusta coffee. Green and roasted coffee samples were used in this study and the degradation of sucrose and trigonelline, with the formation of nicotinic acid, was followed during roasting. Caffeine did not undergo significant degradation with only 5.4% being lost under severe roasting. Sucrose was degraded rapidly during processing with light roasting producing a 97% loss and dark roasting degrading it completely. Loss of trigonelline was strongly dependent on the degree of roasting being higher in the Robusta coffee. Trigonelline degradation was associated with nicotinic acid formation both in the Arabica and Robusta coffees as a consequence of the roasting process. Trigonelline and sucrose were determined simultaneously by partition chromatography and detection with the mass detector. Determination of caffeine was carried out using reversed phase chromatography and nicotinic acid by ion-pair reversed phase chromatography. Detection in both cases was achieved using an ultraviolet detector at 272nm or 254nm, respectively. HPLC showed adequate precision and accuracy for routine analyses. In addition, the methods used were more rapid and simple than traditional procedures. HPLC appears to be a suitable technique for quality control in the coffee industry, and for fundamental investigation on the mechanisms involved in the roasting process (AU) s

Efeito da torrefaçäo no perfil cromatográfico obtido por filtraçäo em gel de extratos de café Arábico / Effect of roasting on the chromatographic profile obtained by gel filtration of Arabian coffee extract

As modificaciones que ocorrem durante a torrefaçäo do café, em relaçäo aos pesos moleculares de seus componentes, foram estudadas, usando-se cromatografia de filtraçäo em gel com colunas do tipo TSK-pW400, sob alta pressäo. Os extratos obtidos de amostras de café verde e torrado em diferentes intensidades foram cromatografados e monitorizados usando-se detectors de índice de refraçäo diferencial (IRD) e de ultra violeta (UV) a 280, 325 e 410nm. Marcantes diferenças foram verificadas nos cromatogramas dos extratos, usando-se o detector IRD, em relaçäo ao café verde e ao café torrado em diferentes intensidades. O uso do detector de UV a 280nm e 325nm mostraram similaridades no perfil cromatogrâfico, indicando que a formaçäo de componentes de alto peso molecular, durante a torrefaçäo, envolve a participaçäo de ligaçöes entre proteínas e compostos fenólicos. A formaçäo de pigmentos pode ser claramente acompanhada durante a torrefaçäo com detecçäo a 420 nm. O uso de cromatografia líquida de alta resoluçäo provou ser um método extremamente rápido, em comparaçäo a filtraçäo em gel tradicional, mostrando-se útil para estudos mais detalhados das modificaçöes de pesos moleculares que ocorrem durante o processo de torrefaçäo do café