Absence Of Caffeine Research Articles

Metabolomic profiling of in vitro and in situ grown Nilgiris tea reveals unique signatures for breeding decaffeinated varieties

This study was begun by establishing an in vitro culture in UPASI 9, a Nilgiris tea clone (Camellia sinensis) by optimising various factors. Anatomical studies demonstrated that use of lower carbendazim concentration for sterilisation (0.2%) produced viable and healthy explants for callus initiation. To confirm the genetic consistency of the regenerated plants, gene-specific SSR markers were developed and utilised. GC-MS was employed to analyse volatile metabolites extracted from callus, stem, micro shoots, and leaves of the UPASI 9 tea genotype. The results revealed distinct compositions of metabolites in each sample. More interestingly, caffeine was exclusively detected in leaf samples but absent in all other investigated tissues, despite the presence of Tea Caffeine Synthase (TCS) gene-specific SSRs. Thus, this study provided unique information on the absence of caffeine in in vitro grown Nilgiris tea clone, UPASI 9, as decaffeinated tea has a unique niche in the global market.

Extracellular cAMP and MRP4 activity influence in vitro capacitation and fertilizing ability of pig spermatozoa

cAMP has been reported to be an essential driver of sperm capacitation. In bovine sperm cAMP efflux through multidrug resistance-associated protein 4 (MRP4) has been suggested to maintain intracellular cAMP homeostasis and generate extracellular signaling able to regulate capacitation. The aim of this work was to determine whether extracellular cAMP may influence in vitro pig sperm capacitation and acquisition of fertilizing ability and to evaluate the role of MRP4. In vitro sperm capacitation and gamete coincubation were performed in Brackett and Oliphant's medium (BO) in presence of caffeine (Ctr+) or in BO without caffeine (Ctr-) supplemented with 0, 8, 9, 10 mM cAMP. Despite the percentage of capacitated sperm, assayed by immunolocalization of tyrosine-phosphorylated proteins, was significantly lower in Ctr- compared to Ctr+, it increased supplementing 10 mM cAMP to Ctr- reaching values similar to Ctr+. The absence of caffeine during gamete coincubation reduced the fertilization rate compared to Ctr+, while 10 mM cAMP supplementation to Ctr- increased the fertilization rate reaching values similar to Ctr + .The presence of MRP4 in pig spermatozoa was detected for the first time by western blot and immunohistochemistry assays. To evaluate MRP4 role on pig sperm capacitation, in vitro capacitation and gamete coincubation were performed in Ctr + in presence of MK571, a MRP4 selective inhibitor. MK571 reduced the percentage of capacitated cells and the fertilization rate, while cAMP addition fully reversed MRP4 blockade consequences.Present findings suggest that, under our in vitro conditions, extracellular cAMP and MRP4 activity influence pig sperm capacitating events.

Bicarbonate and BSA increase the capacitation pattern and acrosomal exocytosis in boar sperm after 120 min of incubation.

Sperm capacitation is a crucial step towards the acquisition of fertilizing capacity. Despite the attempts to mimic the invivo situation, there is still a lack of standardization invitro techniques. Bicarbonate and serum albumin (BSA) are routinely used, although controversial results are reported regarding the optimal concentration of each compound. In addition, whether caffeine is needed on invitro capacitation media in boar sperm remains to be elucidated. Here, 18 boar commercial artificial insemination doses were used to test different concentrations of bicarbonate (19, 37 or 56 mM) in experiment 1, BSA (1.5, 3, 4.5 mg/mL) in experiment 2 and the presence or absence of caffeine (5.15 mM) experiment 3. We analysed at 0, 30 and 120 min of incubation at 38.5°C, 5% CO2 : Total motility (TMOT), membrane integrity (VIAB), acrosomal exocytosis (rAcro; H33342/PI/PNA), capacitation status (chlortetracycline staining CTC) and mitochondrial membrane potential (JC-1). The higher concentrations of bicarbonate (37 and 56 mM) decreased TM and VIAB (p < .01) but increased rAcro (p < .01) after 120 min of incubation compared to the fresh control. In contrast, only the BSA concentration of 3 mg/mL reduced the VIAB at 120 min, but all the concentrations tested increased the average of JC-1 and decreased TM (p < .01) throughout incubation compared to the fresh control. Finally, in experiment 3, when boar sperm were incubated in the capacitating media with bicarbonate, BSA and with or without caffeine, the capacitated pattern measured by the CTC technique and rAcro increased after 120 min of incubation (p < .01) compared to fresh control, either in the presence or in the absence of caffeine. In summary, our results suggested that the combination of capacitating components, like bicarbonate and BSA, contributed to increasing the proportion of capacitated boar spermatozoa, mitochondrial membrane potential as well as acrosomal exocytosis. However, caffeine did not significantly influence invitro sperm capacitation in this species.

Postprandial Metabolic Responses to High-fat Feeding in Healthy Adults Following Ingestion of Oolong Tea–Derived Polymerized Polyphenols: A Randomized, Double-blinded, Placebo-controlled Crossover Study

BackgroundPolymerized polyphenols (PP) found in oolong tea can inhibit pancreatic lipase activity in vitro, and pilot work indicates that this may reduce postprandial lipemia. Since tea contains caffeine and catechins, the interactions between these ingredients and PP warrant investigation. ObjectivesTo assess whether PP ingested alone or with caffeine and catechins lowers postprandial lipemia. MethodsFifty healthy adults [mean (SD) age: 26 (7) y; BMI (in kg/m2): 24.0 (2.7); female: n = 16] completed 4 oral lipid tolerance tests in a placebo-controlled randomized, crossover design. Participants ingested 40 g of fat with either 1) placebo, 2) 100 mg PP, 3) 150 mg PP, or 4) 100 mg PP plus 50 mg caffeine and 63 mg catechins (PP + CC). Blood was sampled for 3 h postprandially to assess concentrations of serum and plasma triacylglycerol and plasma markers of lipid (NEFA; glycerol; LDL and HDL cholesterol; and ApoA-I, A-II, B, C-II, C-III, and E) and glucose metabolism (glucose, insulin, and C-peptide). ResultsSerum and plasma triacylglycerol concentrations and lipid metabolism variables generally increased following any test drink ingestion (main effect of time, p < 0.001). Nevertheless, for the lipid metabolism responses, there were no statistically significant condition–time interactions and no statistically significant differences in incremental or total area under the curve between conditions, apart from HDL cholesterol (p = 0.021). Ingesting 100 mg PP + CC lowered peak plasma glucose, insulin, and C-peptide concentrations compared with all other conditions 30 min postingestion (p < 0.001), with persistent alterations in glucose concentrations observed for 90 min compared with placebo and 100 mg PP conditions. ConclusionsPP ingested at doses ≤150 mg does not clearly alter early-phase postprandial triacylglycerol concentrations in healthy adults, irrespective of the presence or absence of caffeine and catechins. Nevertheless, caffeine and catechins added to PP lowered postprandial glucose and insulin concentrations.This trial was registered in ClinicalTrials.gov as NCT03324191 (https://clinicaltrials.gov/ct2/show/NCT03324191).

Caffeine solubility in aqueous solutions of polyethylene glycol dimethyl ether 250: measurement and thermodynamic modeling

ABSTRACT Molar solubility of caffeine in polyethylene glycol dimethyl ether 250 (PEGDME 250) + water mixtures over all mass fractions of PEGDME 250 were determined at 293.15–313.15 K under atmospheric pressure (≈ 85 kPa). The crystal forms of the caffeine in equilibrium with neat solvents of PEGDME 250 and water were analysed using X-ray powder diffraction and the results compared with the original caffeine. Also, the Hansen solubility parameters of caffeine and PEGDME 250 + water mixtures in the absence of caffeine were predicted. The studies of apparent thermodynamic analysis presented an endothermic and enthalpy-driven dissolution of caffeine. Moreover, the solubility of caffeine were correlated using various models; the van’t Hoff, mixture response surface, Jouyban–Acree, Jouyban–Acree–van’t Hoff, the modified version of Jouyban–Acree–van’t Hoff, the CNIBS/R-K, the Buchowski–Ksiazczak, the modified Wilson and modified Wilson–van’t Hoff models, acquiring average relative deviation values that were lower than 14.7%.

Investigation of potential behavioral and physiological effects of caffeine on D. magna.

With the ever-increasing consumption of pharmaceutical compounds, their presence in the environment is now an undisputable reality. The majority of these compounds are released into the wild after their therapeutic use, as biotransformation products or in their original form. The presence of this class of compounds in the environment, due to their biological properties, can exert effects on non-target organisms, with adverse consequences. In addition, some bioactive substances, such as stimulants of the central nervous system, are also used by humans as part of their diet. The adverse consequences posed by such chemicals may be permanent or transient, if the exposure to xenobiotics is halted; it is thus of the paramount importance to study effects that result from long-term exposure to toxicants, but also the recovery of organisms previously exposed to such substances, especially if such chemicals may cause some type of addiction. Caffeine (1,3,7-trimethylxanthine) is a naturally occurring alkaloid found in many plants, being one of the most common stimulant/pharmaceutical compounds found in the environment. In addition, it is addictive, and strongly consumed by humans, a factor that contributes also for its continuous presence in the aquatic environment. The aim of this study was to evaluate the effects of environmentally relevant concentrations (0.08; 0.4; 2; 10; and 50μg/L) of caffeine on behavior and physiological parameters (that are proxies of metabolic traits, such as oxygen uptake and glycogen content), in individuals of the freshwater crustacean species Daphnia magna, of distinct ages, and with or without a recovery period in the absence of caffeine. Regarding behavior, the results indicated that caffeine exposure altered the moved distance of the test organisms, but not according to a coherent pattern; low concentrations of caffeine reduced the movement of exposed daphnids, while higher levels did not have any measurable effect on this parameter. In addition, it was possible to identify subtle withdrawal effects (animals exposed to caffeine during 21days and kept in uncontaminated media for 2days). Regarding the other two studied parameters, caffeine exposure did not result in any significant modification in oxygen uptake and glycogen stores/reserves of the test organisms, in animals continuously exposed, or in those subjected to a recovery period, suggesting that despite a behavioral stimulatory effect, this was not followed by any metabolic change, and no addictive effect was possible to infer. The results showed that the presence of caffeine in environmental concentrations can induce mild behavioral effects at low, albeit realistic levels, but not capable of establishing clear biochemical changes.

Fractionated extraction of polyphenols from mate tea leaves using a combination of hydrophobic/ hydrophilic NADES

A new methodology for the selective extraction of antioxidants from mate tea leaves (and decaffeinated mate tea leaves), using different natural deep eutectic systems (NADES), is reported in this paper. A fractionated extraction was carried out and the optimization of the extraction conditions such as solid/liquid ratio, temperature, time, stirring and the use of ultrasound assisted extraction (UAE) technology was performed. The results demonstrate that a sequential extraction using, in a first step, an hydrophobic system Men:Lau (2:1) and, in a second step, an hydrophilic lactic acid-based NADES, leads to two distinct extracts: the first one rich in pigments and the second one rich in polyphenols.NADES systems were able to extract 30% more of the polyphenolic components of the mate tea leaves matrices, when compared with traditional solvents/techniques. Moreover, it has been shown that the incorporation of the extract in the NADES, compared to the same extract in aqueous medium was beneficial for the stabilization of the antioxidants. It maintains their functionality at least for three months, reaching 41% more versus the extracts obtained by traditional solvents/techniques. The absence of caffeine in the extracts did not shown to have any effects on the stability results.

Single versus mixed edge activators in caffeine-loaded transfersomes: physicochemical and cytotoxicity assessment

The performance of transfersomes as nanovesicular systems to enhance the skin permeation of bioactive compounds may be modulated according to their composition of edge activators (EAs). Vesicle size, deformability, and encapsulation efficiency can be altered when using nonionic surfactants as EAs with opposing hydrophilic-lipophilic balance (HLB). Thus, this work aimed to assess the impact on the physicochemical properties and the in vitro cytotoxicity profile of transfersomal formulations composed of single or mixed EAs, with opposing HLB values. Transfersomes made of single or mixed nonionic surfactants (Tween® 80 and/or Span® 80) were prepared in the presence and absence of caffeine, used as a model hydrophilic compound. Single or mixed EAs-containing transfersomes displayed promising physicochemical properties for cutaneous applications and were found to be stable for one month of refrigerated storage. Moreover, all transfersomal formulations were compatible with a keratinocyte cell line upon 24 h treatment. Altogether, these preliminary data suggest that the use of mixed nonionic surfactants as EAs may be further explored to modulate the performance of transfersomes as skin permeation enhancers. Keywords: Vesicular systems, nanotechnology, nonionic surfactants, caffeine

G2/M Checkpoint Abrogation With Selective Inhibitors Results in Increased Chromatid Breaks and Radiosensitization of 82-6 hTERT and RPE Human Cells.

While technological advances in radiation oncology have led to a more precise delivery of radiation dose and a decreased risk of side effects, there is still a need to better understand the mechanisms underlying DNA damage response (DDR) at the DNA and cytogenetic levels, and to overcome tumor resistance. To maintain genomic stability, cells have developed sophisticated signaling pathways enabling cell cycle arrest to facilitate DNA repair via the DDR-related kinases and their downstream targets, so that DNA damage or DNA replication stress induced by genotoxic therapies can be resolved. ATM, ATR, and Chk1 kinases are key mediators in DDR activation and crucial factors in treatment resistance. It is of importance, therefore, as an alternative to the conventional clonogenic assay, to establish a cytogenetic assay enabling reliable and time-efficient results in evaluating the potency of DDR inhibitors for radiosensitization. Toward this goal, the present study aims at the development and optimization of a chromosomal radiosensitivity assay using the DDR and G2-checkpoint inhibitors as a novel modification compared to the classical G2-assay. Also, it aims at investigating the strengths of this assay for rapid radiosensitivity assessments in cultured cells, and potentially, in tumor cells obtained from biopsies. Specifically, exponentially growing RPE and 82-6 hTERT human cells are irradiated during the G2/M-phase transition in the presence or absence of Caffeine, VE-821, and UCN-1 inhibitors of ATM/ATR, ATR, and Chk1, respectively, and the induced chromatid breaks are used to evaluate cell radiosensitivity and their potency for radiosensitization. The increased yield of chromatid breaks in the presence of DDR inhibitors, which underpins radiosensitization, is similar to that observed in cells from highly radiosensitive AT-patients, and is considered here as 100% radiosensitive internal control. The results highlight the potential of our modified G2-assay using VE-821 to evaluate cell radiosensitivity, the efficacy of DDR inhibitors in radiosensitization, and reinforce the concept that ATM, ATR, and Chk1 represent attractive anticancer drug targets in radiation oncology.

The absence of the caffeine synthase gene is involved in the naturally decaffeinated status of Coffea humblotiana, a wild species from Comoro archipelago

Caffeine is the most consumed alkaloid stimulant in the world. It is synthesized through the activity of three known N-methyltransferase proteins. Here we are reporting on the 422-Mb chromosome-level assembly of the Coffea humblotiana genome, a wild and endangered, naturally caffeine-free, species from the Comoro archipelago. We predicted 32,874 genes and anchored 88.7% of the sequence onto the 11 chromosomes. Comparative analyses with the African Robusta coffee genome (C. canephora) revealed an extensive genome conservation, despite an estimated 11 million years of divergence and a broad diversity of genome sizes within the Coffea genus. In this genome, the absence of caffeine is likely due to the absence of the caffeine synthase gene which converts theobromine into caffeine through an illegitimate recombination mechanism. These findings pave the way for further characterization of caffeine-free species in the Coffea genus and will guide research towards naturally-decaffeinated coffee drinks for consumers.

Impact of ‘free-from’ and ‘healthy choice’ labeled versions of chocolate and coffee on temporal profile (multiple-intake TDS) and liking

Product reformulation for obtaining “free-from” or “healthy choice” versions may confer significant changes in sensory characteristics which could not meet consumer expectations in terms of sensory perception. Therefore, this paper aims to evaluate the influence of formulations of different product categories on their dynamic sensory profile and consumer acceptability. A sensory panel of 23 semi-trained assessors evaluated two product categories, chocolate and milk coffee, using Temporal Dominance of Sensations (TDS) over three consecutive intakes/sips (multiple-intake TDS) on two replications. TDS allowed us to describe the dynamic profile of classic products over the three intakes and to identify differences in the sensory temporal profile compared to their 'healthier choice' reformulated versions. For the chocolate category, increasing the cocoa and the reduction of lactose content mainly influenced the dominance of sweetness, dairy, cocoa, crunchy and bitterness attributes. No sugar addition and absence of caffeine (decaffeinated version) in ready to drink milk coffee beverage modified temporal perception of sweetness, bitterness, coffee flavor and smoothness texture. Higher overall liking averages were associated with the dominant attributes of chocolate with a higher level of cacao and milk coffee decaffeinated. The obtained results of the present work suggest that the multiple-intake TDS technique can be applied for the evaluation of different food categories, helping to develop healthier products and predicting the liking.

Acute cognitive, mood and cardiovascular effects of green and black tea

AbstractTea is often described as having “refreshing” properties. Indeed, components of both green and black tea have been shown to acutely improve aspects of cognition and mood and to affect cardiovascular function. There is, however, surprising little research directly comparing these effects between different types of tea. This study therefore examined acute cognitive, mood, and cardiovascular effects of green tea and black tea.A double blind, randomised, placebo-controlled,balanced, three-way cross-over study was conducted. Fourteen healthy young adults (M = 23.8, SD = 3 years) consumed placebo (water), green tea and black tea in balanced order at least 24 h apart.The drinks were prepared according to standardised procedures and consumed under double-blind conditions (in opaque containers with lemon cordial added for taste-masking). Mood and cognition were measured at baseline then 90 min post-treatment, cardiovascular function was assessed post-dose only. Mood was evaluated using a series of visual analogue scales, cognitive performance was assessed using a suite of standardised tests from the Computerised Mental Performance Assessment System (COMPASS), gauging performance on aspects of attention, memory and executive function.Peripheral and central systolic and diastolic pressure,augmentation pressure, augmentation index,and blood flow velocity were measured via Sphygmocor pulse wave analysis and Doppler ultrasound, respectively. There were a number of significant effects associated with the active treatments. Compared with placebo, black tea and green tea was associated with significantly improved digit vigilance accuracy (p &lt; .01), while green tea was associated with higher alertness (p &lt; .05).Regarding cardiovascular outcomes, black tea and green tea significantly increased central diastolic pressure, and peripheral diastolic pressure in comparison to placebo (P &lt; .01). Black tea also significantly increased central systolic pressure compared to green tea and placebo (p &lt; .05). Analysis of treatment guessing suggested that the masking procedure was highly effective. In conclusion, the present study shows that green tea improves alertness. Since black tea contains higher levels of caffeine, this effect is likely underpinned by non-caffeine mechanisms or caffeine/non-caffeine compound interactions.Improved vigilance is consistent with caffeine effects which may be modulated by cardiovascular effects. Interestingly the cardiovascular effects are opposite to those reported for chronic tea interventions. These results suggest that acute cognitive and cardiovascular components of tea merit further investigation both in the presence and absence of caffeine.

Examination of some toxicological parameters of dimethylamylamine when consumed alone or with caffeine

Dimethylamylamine (DMAA) is a bodybuilding supplement with fat-burner or performance-enhancing properties. DMAA is often combined with caffeine to enhance its effectiveness and this can have serious adverse effects on health. In this study, we examined for the first time the cytotoxic, oxidative and genotoxic effects of DMAA in the presence or absence of caffeine in lymphocytes cultured from human blood, and its vascular irritant effects in a hen's chorioallantoic membrane egg test. Cytotoxic effects were observed by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT), lactate dehydrogenase release (LDH), which serves as a measure of cell membrane damage, changes in the mitotic index (MI) and proliferative rate index (PRI) assays. Oxidative changes were evaluated by the total antioxidant activity and the total oxidative status assay. Genotoxic damage was analyzed by chromosomal aberration and micronucleus assays. DMAA and its combination with caffeine (cDMAA) had no genotoxic effects. DMAA (1000 mg/L) and cDMAA (500 and 1000 mg/L) decreased cell viability while significantly increasing LDH activity, MI and the oxidative level. DMAA and cDMAA caused weak and moderate vascular irritant effects, respectively. In conclusion, DMAA exhibits cytotoxic effects via membrane dysfunction and mitotic disturbance following increased oxidative stress in a dose- and caffeine-dependent manner.

Caffeine attenuates contraction-induced diminutions of the intracellular calcium transient in mouse lumbrical muscle ex vivo

The amount of calcium released from the sarcoplasmic reticulum in skeletal muscle rapidly declines during repeated twitch contractions. In this study, we test the hypothesis that caffeine can mitigate these contraction-induced declines in calcium release. Lumbrical muscles were isolated from male C57BL/6 mice and loaded with the calcium-sensitive indicator, AM-furaptra. Muscles were then stimulated at 8 Hz for 2.0 s in the presence or absence of 0.5 mM caffeine, at either 30 °C or 37 °C. The amplitude and area of the furaptra-based intracellular calcium transients and force produced during twitch contractions were calculated. For each of these measures, the values for twitch 16 relative to twitch 1 were higher in the presence of caffeine than in the absence of caffeine at both temperatures. We conclude that caffeine can attenuate contraction-induced diminutions of calcium release during repeated twitch contractions, thereby contributing to the inotropic effects of caffeine.

Effect of Caffeine extracted from Coffee on the Growth of Bacteria Isolated from Vellore

Caffeine is a chemical compound belonging to the class methylxanthines which acts as stimulant. It is a widely consumed stimulant which helps to prevent drowziness and enhances alertness by its action on the central nervous system. The effect of caffeine is well known among humans and it is commonly consumed in the form of coffee. Coffee beans have a bitter taste which acts a as a deterrent against certain animals and insects. However, in this study we will discuss about the effect of caffeine on growth of bacteria isolated from Vellore. Caffeine can be extracted from various sources like coffee beans, tea, Cacao beans, Kola nut etc. In our study, we have extracted caffeine from commercially available coffee powder and bacteria from soil. Then, by using spread plate technique, we cultivated the bacteria to observe its growth in the presence and the absence of caffeine [7, 10]. We also used various stains to observe its morphological changes.

A Unified Model of Performance for Predicting the Effects of Sleep and Caffeine.

Existing mathematical models of neurobehavioral performance cannot predict the beneficial effects of caffeine across the spectrum of sleep loss conditions, limiting their practical utility. Here, we closed this research gap by integrating a model of caffeine effects with the recently validated unified model of performance (UMP) into a single, unified modeling framework. We then assessed the accuracy of this new UMP in predicting performance across multiple studies. We hypothesized that the pharmacodynamics of caffeine vary similarly during both wakefulness and sleep, and that caffeine has a multiplicative effect on performance. Accordingly, to represent the effects of caffeine in the UMP, we multiplied a dose-dependent caffeine factor (which accounts for the pharmacokinetics and pharmacodynamics of caffeine) to the performance estimated in the absence of caffeine. We assessed the UMP predictions in 14 distinct laboratory- and field-study conditions, including 7 different sleep-loss schedules (from 5 h of sleep per night to continuous sleep loss for 85 h) and 6 different caffeine doses (from placebo to repeated 200 mg doses to a single dose of 600 mg). The UMP accurately predicted group-average psychomotor vigilance task performance data across the different sleep loss and caffeine conditions (6% < error < 27%), yielding greater accuracy for mild and moderate sleep loss conditions than for more severe cases. Overall, accounting for the effects of caffeine resulted in improved predictions (after caffeine consumption) by up to 70%. The UMP provides the first comprehensive tool for accurate selection of combinations of sleep schedules and caffeine countermeasure strategies to optimize neurobehavioral performance.

Longitudinal bone growth is impaired by direct involvement of caffeine with chondrocyte differentiation in the growth plate.

We showed previously that caffeine adversely affects longitudinal bone growth and disrupts the histomorphometry of the growth plate during the pubertal growth spurt. However, little attention has been paid to the direct effects of caffeine on chondrocytes. Here, we investigated the direct effects of caffeine on chondrocytes of the growth plate invivo and invitro using a rapidly growing young rat model, and determined whether they were related to the adenosine receptor signaling pathway. A total of 15 male rats (21days old) were divided randomly into three groups: a control group and two groups fed caffeine via gavage with 120 and 180mgkg-1 day-1 for 4weeks. After sacrifice, the tibia processed for the analysis of the long bone growth and proliferation of chondrocytes using tetracycline and BrdU incorporation, respectively. Caffeine-fed animals showed decreases in matrix mineralization and proliferation rate of growth plate chondrocytes compared with the control. To evaluate whether caffeine directly affects chondrocyte proliferation and chondrogenic differentiation, primary rat chondrocytes were isolated from the growth plates and cultured in either the presence or absence of caffeine at concentrations of 0.1-1mm, followed by determination of the cellular proliferation or expression profiles of cellular differentiation markers. Caffeine caused significant decreases in extracellular matrix production, mineralization, and alkaline phosphatase activity, accompanied with decreases in gene expression of the cartilage-specific matrix proteins such as aggrecan, type II collagen and type X. Our results clearly demonstrate that caffeine is capable of interfering with cartilage induction by directly inhibiting the synthetic activity and orderly expression of marker genes relevant to chondrocyte maturation. In addition, we found that the adenosine type 1 receptor signaling pathway may be partly involved in the detrimental effects of caffeine on chondrogenic differentiation, specifically matrix production and mineralization, as evidenced by attenuation of the inhibitory effects of caffeine by blockade of this receptor. Thus, our study provides novel information on the integration of caffeine and adenosine receptor signaling during chondrocyte maturation, extending our understanding of the effect of caffeine at a cellular level on chondrocytes of the growth plate.

Antioxidative potential of different coffee substitute brews affected by milk addition

Due to the specific sensory properties, a rich bioactive composition, attributed to different ingredients used for their production, as well as the absence of caffeine, coffee substitutes are becoming increasingly popular alternatives to usual coffee brews. The objective of this study was to determine the content of polyphenolic compounds and antioxidant capacity of 10 coffee substitute brews with different composition and the effect of milk addition to coffee substitute brews on their antioxidant properties. The content of total phenols and flavonoids, flavan-3-ols and proanthocyanidins, as well as the antioxidant capacity of coffee substitute brews, was determined using UV/Vis spectrophotometric methods. Consumer analysis and principal component analysis were conducted in order to further characterize the sensory properties and antioxidant parameters of coffee substitute brews. Total phenol content of coffee substitute brews prepared with water varied between 345.45 and 2190.91 mg gallic acid equivalents/L and decreased, as well as the antioxidant capacity, proportionally with the addition of milk. The antioxidant potential of coffee substitute brews was significantly affected by their composition. The combination of roasted chicory and barley provided the highest antioxidant potential, while coffee substitutes composed of several different ingredients exhibited the best sensory properties.

Effects of adolescent caffeine consumption on cocaine sensitivity.

Caffeine is the most commonly used psychoactive substance, and consumption by adolescents has risen markedly in recent years. We identified the effects of adolescent caffeine consumption on cocaine sensitivity and determined neurobiological changes within the nucleus accumbens (NAc) that may underlie caffeine-induced hypersensitivity to cocaine. Male Sprague-Dawley rats consumed caffeine (0.3 g/l) or water for 28 days during adolescence (postnatal day 28-55; P28-P55) or adulthood (P67-P94). Testing occurred in the absence of caffeine during adulthood (P62-82 or P101-121). Cocaine-induced and quinpirole (D2 receptor agonist)-induced locomotion was enhanced in rats that consumed caffeine during adolescence. Adolescent consumption of caffeine also enhanced the development of a conditioned place preference at a sub-threshold dose of cocaine (7.5 mg/kg, i.p.). These behavioral changes were not observed in adults consuming caffeine for an equivalent period of time. Sucrose preferences were not altered in rats that consumed caffeine during adolescence, suggesting there are no differences in natural reward. Caffeine consumption during adolescence reduced basal dopamine levels and augmented dopamine release in the NAc in response to cocaine (5 mg/kg, i.p.). Caffeine consumption during adolescence also increased the expression of the dopamine D2 receptor, dopamine transporter, and adenosine A1 receptor and decreased adenosine A2A receptor expression in the NAc. Consumption of caffeine during adulthood increased adenosine A1 receptor expression in the NAc, but no other protein expression changes were observed. Together these findings suggest that caffeine consumption during adolescence produced changes in the NAc that are evident in adulthood and may contribute to increases in cocaine-mediated behaviors.

P-32 * EFFECTS OF INTESTINAL MOTILITY ON ETHANOL ABSORPTION IN SMALL INTESTINE

Administration of caffeine and Ryokucha Saponin markedly decreases the metabolic rate of ethanol. The effect of caffeine and Ryokucha Saponin on ethanol absorption in small intestine was investigated. Sample animals were used male Wistar rats (6 week). The rate of intestinal absorption of ethanol was measured using the small intestine of rats according to Creine and Wilson method ( J Appl Physiol 12 : 145–146, 1958). The solution which transmitted small intestine was measured for ethanol levels at 0 to 30 minutes in presence or absence of caffeine or Ryokucha Saponin with ethanol solution. Contractile activity of the intestine was investigated by Magnus apparatus. A small intestine was removed from rats and were placed in a Tyrode solution. The opposite end of the tissue was secured to a sensitive strain gauge and tension measurement were recorded on a kymographion. The rate of intestinal absorption of ethanol was found that the increase in the ethanol absorption is depending on time. In the presence of caffeine and Ryokucha Saponin reduced ethanol absorption about over 10% of control. Caffeine and Ryokucha Saponin inhibits intestinal motility. Based on these results, we speculate that intestinal motility by increase and decrease were augment and suppres s alcohol absorption.