The gut-liver axis mechanism of increased susceptibility to non-alcoholic fatty disease in female offspring rats with prenatal caffeine exposure

Caffeine intake during pregnancy is common,while its effect on gut microbiota composition of offspring and the relationship with susceptibility to adult diseases remains unclear.This study aimed to confirm the effects of prenatal caffeine exposure(PCE)on the gut microbiota composition and its metabolites in female offspring rats,and to further elucidate its underlying mechanism and intervention targets in adult non-alcoholic fatty disease(NAFLD).The results showed that the gut microbiota of PCE female offspring at multiple time points from infancy to adolescence were significantly changed with depletion of butyric acid-producing bacteria,leading to a decrease in butyric acid in adulthood.It was also found that PCE female offspring rats were sensitive to NAFLD induced by a postnatal high-fat diet(HFD),which is mainly related to the enhancement of hepatic triglyceride synthesis function.Through mechanism exploration,we found that HFD further reduced the fecal and serum butyric acid levels in the PCE female offspring,which was significantly negatively correlated with hepatic SREBP-1c/FASN mRNA expression and triglyceride level.In vivo and in vitro experiments confirmed that sodium butyrate(NaB)supplementation could reduce hepatic lipid accumulation through MCT1/GPR109A-AMPK,thereby effectively decreasing the susceptibility to NAFLD in the PCE female offspring rats.

The transcription factor CsS40 negatively regulates TCS1 expression and caffeine biosynthesis in connection to leaf senescence in Camellia sinensis

Caffeine is considered as one of the most important bioactive components in the popular plant beverages tea,cacao,and coffee,but as a wide-spread plant secondary metabolite its biosynthetic regulation at transcription level remains largely unclear.Here,we report a novel transcription factor Camellia sinensis Senescnece 40(CsS40)as a caffeine biosynthesis regulator,which was discovered during screening a yeast expression library constructed from tea leaf cDNAs for activation of tea caffeine synthase(TCS1)promoter.Besides multiple hits of the non-self-activation CsS40 clones that bound to and activated TCS1 promoter in yeast-one-hybrid assays,a split-luciferase complementation assay demonstrated that CsS40 acts as a transcription factor to activate the CsTCS1 gene and EMSA assay also demonstrated that CsS40 bound to the TCS1 gene promoter.Consistently,immunofluorescence data indicated that CsS40-GFP fusion was localized in the nuclei of tobacco epidermal cells.The expression pattern of CsS40 in‘Fuding Dabai’developing leaves was opposite to that of TCS1;and knockdown and overexpression of CsS40 in tea leaf calli significantly increased and decreased TCS1 expression levels,respectively.The expression levels of CsS40 were also negatively correlated to caffeine accumulation in developing leaves and transgenic calli of‘Fuding Dabai’.Furthermore,overexpression of CsS40 reduced the accumulation of xanthine and hypoxanthine in tobacco plants,meanwhile,increased their susceptibility to aging.CsS40 expression in tea leaves was also induced by senescence-promoting hormones and environmental factors.Taken together,we showed that a novel senescence-related factor CsS40 negatively regulates TCS1 and represses caffeine accumulation in tea cultivar‘Fuding Dabai’.The study provides new insights into caffeine biosynthesis regulation by a plant-specific senescence regulator in tea plants in connection to leaf senescence and hormone signaling.

A Comparative Analysis of Caffeine Extraction Efficiency from Different Tea Varieties and Its Effect on Human Physiology: A Spectrophotometric Investigation

The current work imitates the trivial method which intricates the abstraction along with classification of caffeine accessible in variability of decoction shrubberies which are expended in two different states of India (Telengana & Uttar Pradesh). As per observation individuals of both the states are identical tender of consumption of tea. Abstraction progression tangled variability of stages which are discoursed in the work. In addition, the outcome of drinking of caffeine or the permitted ingestion of caffeine over tea has also been conferred through the work and linked the data with the literature data. In this study, we used a UV-visible spectrophotometer as well as liquid-liquid extraction method to determine the caffeine content in various tea samples. The significance of this study lies in the fact that accurate determination of caffeine content is essential for quality control and labeling of tea products. The UV-visible spectrophotometer method was found to be simple, reliable, and sensitive for the detection of caffeine in tea samples.

Modified Caffeine Release System and Its Immunomodulatory Effects on Breast Tumor Cells and Blood Phagocytes

Caffeine is an alkaloid present in a wide variety of plants. Currently the most consumed psychostimulant worldwide, its consumption is associated with several health benefits, including modulation of the innate and adaptive immune response, reduction of oxidative cellular stress, and decreased incidence of some cancers, including breast cancer. Breast cancer is the most common cause of cancer among women and the second leading cause of cancer death in women worldwide. The interaction between biomaterials and drugs has enabled a great advance in science for developing controlled drug delivery systems and has been used to treat numerous pathologies. This work aimed to evaluate the immunomodulatory effects of caffeine associated or not with polyethylene glycol adsorbed in microemulsion (MLP) on MCF-7 cells, phagocytic cells (MN), and coculture. For biological assays, ATCC (American Type Culture Collection, USA) cell lines of breast adenocarcinoma (MCF-7) and phagocytes (MN) obtained from voluntary donors were used. The cells (MN and MCF-7) and coculture were treated with caffeine and MLP and incubated for rheological characterization analyses: flow curve and viscosity, oxidative stress: superoxide anion assay (), and activity of the enzyme superoxide dismutase (Cu-Zn-SOD). Caffeine and MLP increased viscosity and blood and MCF-7 cells and affected the immunomodulation of oxidative stress metabolism of MN and MCF-7 cells treated with caffeine and associated caffeine to the MLP. These data suggest that caffeine is associated or not with MLP-induced immunomodulatory effects on MN phagocytes and MCF-7 cells, demonstrating the antitumor activity via oxidative stress and can be a complementary alternative for treating breast cancer.

CsXDH1 gene promotes caffeine catabolism induced by continuous strong light in tea plant

Tea plant(Camellia sinensis)is an important cash crop with extensive adaptability in the world.However,complex environmental factors force a large variation of tea quality-related components.Caffeine is essential for the formation of bitter and fresh flavors in tea,and is the main compound of tea that improves human alertness.Continuous strong light stimulation was observed to cause caffeine reduction in tea leaves,but the mechanism is not clear.In this study,the response of tea plant to light intensity was analysed mainly by multi-omics association,antisense oligodeoxynucleotide(asODN)silencing technique,and in vitro enzyme activity assay.The results revealed multiple strategies for light intensity adaptation in tea plant,among which the regulation of chloroplasts,photosynthesis,porphyrin metabolism,and resistance to oxidative stress were prominent.Caffeine catabolism was enhanced in continuous strong light,which may be a light-adapted strategy due to strict regulation by xanthine dehydrogenase(XDH).asODN silencing and enzymatic activity assays confirmed that CsXDH1 is a protein induced by light intensity to catalyze the substrate xanthine.CsXDH1 asODN silencing resulted in significant up-regulation of both caffeine and theobromine in in vitro enzyme activity assay,but not in vivo.CsXDH1 may act as a coordinator in light intensity adaptation,thus disrupting this balance of caffeine catabolism.

Characterizing the Impact of Caffeine on Heart Arrhythmias

Caffeine is one of the most commonly consumed stimulants and is found in many items like coffee and energy drinks. Heart arrhythmias are irregular heart rhythms, which can occur when the electrical signals that control the heart’s rhythm are not functioning properly. Due to the stimulant properties of caffeine, it is theorized that caffeine consumption may cause tachycardias-like ventricular arrhythmias. This review article describes the relationship between caffeine intake and heart arrhythmias using a comprehensive Pub-Med search. A comprehensive search was conducted using the search terms “caffeine arrhythmia” which was conducted and a total of 26 search results were obtained. The majority of clinical studies suggest that there are no strong associations between caffeine consumption and arrhythmias. There is little evidence suggesting a direct relationship between caffeine and ventricular arrhythmias (relative Risk 1.00, 95% CI 0.94 - 1.06;13.5%, p = 0.32). Conversely, caffeine consumption has an inverse relationship with the risk of atrial fibrillation (p for overall trend = 0.015;p for nonlinearity = 0.27). Caffeine related deaths are uncommon, but certain groups such as infants, psychiatric patients, and athletes may have an increased risk of arrhythmias following caffeine consumption. Overall, caffeine consumption is not strongly linked to heart arrhythmias and limited studies suggest it may reduce the risk of arrhythmias. Although there is not a strong relationship between caffeine intake and heart arrhythmias, it does cause other cardiovascular problems including high blood pressure and hence should be consumed responsibly (40 - 180 mg/day).

Development of a Certified Reference Material from Caffeine Solution for Assuring the Quality of Food and Drug Measurements

Caffeine intake by pregnant women, adults and children can be harmful to the health of all particularly fetuses if the intake exceeds the permissible limits. Therefore, it is of fundamental importance to measure its concentration accurately using certified reference materials (CRMs). In the literature, no scientific details are published about the certification of caffeine standard solutions, and therefore, the present article covers this gap. A batch of caffeine solution was prepared in concentration of 1000 mg/kg and bottled. Homogeneity and stability of the candidate reference material were assessed by HPLC-UV and the results showed that the material is homogenous and stable enough. Characterization of the caffeine reference material was performed by HPLC-UV, LC-MS/MS and UV-VIS-NIR spectrophotometer in three different days and the characterization uncertainty was estimated in accordance with the requirements of ISO GUM. The certified value (999.86 ± 8.57 mg/kg) was derived as a weighted mean from the gravimetry and the three characterization methods and the certified uncertainty was calculated according to ISO Guide 35. The produced CRM is of strong interest to the food and drug analytical laboratories for the validity and credibility of their caffeine measurement results.

Validation of a Method for the Measurement of Caffeine in Water by HPLC-UV

For the production of a reference material from caffeine solution, one of the methods of characterization was HPLC-UV since caffeine is very sensitive to the UV. In this work, a batch solution of caffeine in water reference material of 1000 mg/kg has been gravimetrically prepared using a calibrated analytical balance. A sample of this solution was diluted to 25 mg/kg for measurement by HPLC-UV in the range 10 - 50 mg/kg. The chromatographic separation was carried out by C-18 column and a mobile phase assembled of 75% water and 25% methanol (v:v). The detection was made by the UV detector at 275 nm. The validation of this analytical method was carried out in accordance with requirements of the EURACHEM and ICH guidelines. The selectivity, linearity, accuracy, precision and trueness (recovery and bias) of the method were studied. The validation results proved that the method is fit-for-purpose of measuring the caffeine concentration in water in the range 10 - 50 mg/kg using HPLC-UV.

Development of a Certified Reference Material from Caffeine Solution for Assuring the Quality of Food and Drug Measurements

Caffeine intake by pregnant women, adults and children can be harmful to the health of all particularly fetuses if the intake exceeds the permissible limits. Therefore, it is of fundamental importance to measure its concentration accurately using certified reference materials (CRMs). In the literature, no scientific details are published about the certification of caffeine standard solutions, and therefore, the present article covers this gap. A batch of caffeine solution was prepared in concentration of 1000 mg/kg and bottled. Homogeneity and stability of the candidate reference material were assessed by HPLC-UV and the results showed that the material is homogenous and stable enough. Characterization of the caffeine reference material was performed by HPLC-UV, LC-MS/MS and UV-VIS-NIR spectrophotometer in three different days and the characterization uncertainty was estimated in accordance with the requirements of ISO GUM. The certified value (999.86 ± 8.57 mg/kg) was derived as a weighted mean from the gravimetry and the three characterization methods and the certified uncertainty was calculated according to ISO Guide 35. The produced CRM is of strong interest to the food and drug analytical laboratories for the validity and credibility of their caffeine measurement results.

利用差异转录组定位Paraburkholderia caffeinilytica CF1咖啡因降解功能基因

实验室前期从茶园土壤中筛选到一株高效咖啡因降解菌CF1。为了寻找菌株CF1降解咖啡因的功能基因,对有无咖啡因添加条件下菌株CF1的转录组进行测序,并进行了差异转录组分析。结果显示,共得到4 871条Unigene序列,以菌株CF1的全基因组为模板,96.96%的序列可得到注释。组间差异基因分析结果显示,添加咖啡因条件下上调基因为123条,而下调基因为2 669条。在上调基因中,112条基因编码在染色体上,11条基因编码在巨型质粒上。通过Blast比对和关键酶酶活测定,证明了这11条编码在质粒上的基因与咖啡因降解功能直接相关,且聚集排布形成一条咖啡因降解基因簇。

在inside-out膜片上caffeine对猪冠状动脉平滑肌细胞钙激活钾通道的调节作用及ryanodine的影响

目的:利用膜片钳技术,研究咖啡因对钙激活钾通道(calcium-activated potassium channel,KCa)的作用机制,及咖啡因存在时兰尼定(ryanodine)对KCa的影响以阐明冠状动脉平滑肌的调控机制。方法:采用急性酶分离方法,应用膜片钳单通道电流记录技术记录猪冠状动脉平滑肌钿胞上KCa电流活动。电流信号经放大、滤波及A/D、D/A转换后输入微机进行采样和储存。应用PCLAMP9.0软件系统进行数据采集及分析。结果:在内面向外式(inside-out)膜片下,咖啡因(0.1、0.5、1.0、5.0mmol/L)可浓度依赖性地增加通道开放概率,而对电流幅值无明显影响,开放概率增加是通过明显缩短平均关闭时间实现的(n=8,P<0.01);洗去药物后通道活性可以恢复到对照水平;5.0mmol/L咖啡因对KCa激活作用最大(P<0.01)。在细胞贴附式(cell-attached)膜片上,咖啡因激活KCa后,ryanodine(10-40μmol/L)浓度依赖性地抑制通道开放概率,开放时间缩短,关闭时间延长,对电流幅度无明显影响。结论:在inside-out膜片下,咖啡因能够直接激活猪冠状动脉平滑肌细胞KCa通道。在cell-atta-ched构型上,ryanodine可通过胞内一定的信号通路浓度依赖性间接抑制咖啡因对KCa激活。

Influence of Electric Field Direction on Enhanced Transdermal Delivery of Caffeine by Electroporation

Aim To study the influence of electric field direction on the in vitro enhanced transdermal delivery of caffeine by eleetroporation. Methods Using side-by-side compartment diffusion cells method and Ag-Ag/AgCl electrodes, the transport of caffeine through human cadaver skin by electroporation （exponentially decaying pulse, pulse voltage = 350 V, pulse frequency = 4 pulses· min^-1, capacity = 22 μF, pulse length = 7 ms, 25 pulses） with different electric field directions was carried out and compared with passive diffusion and iontophoresis （0.25 mA·cm^ - 2, lasted for 4 h）. Results （i） The cumulative quantity and flux of caffeine through human skin were increased significantly by eleetroporation or iontophoresis. （ii） The transport of caffeine by positive iontophoresis （ with electric field from donor to receptor compartment） was significantly greater than that by negative iontophoresis （with electric field from receptor to donor compartment）. （iii） The transport of caffeine by positive eleetroporation （with electric field from donor to receptor compartment） was similar to that by negative eleetroporation （with electric field from receptor to donor compartment）. （iv） The enhancing effect of positive iontophoresis on the transdermal delivery of caffeine was significantly greater than that of electroporation （positive or negative）. Conclusion Electric field direction significantly influences the enhancing effect of iontophoresis on the transdermal delivery of caffeine, but does not influence the enhancing effect of eleetroporation.

新法合成Cu_2(Benzoate)_4(Caffeine)_4及其性质

新法合成了铜()、咖啡因、苯甲酸的三元配合物,并用红外光谱、元素分析、X-单晶衍射和热重分析进行了表征,配合物与ct-DNA作用的紫外-可见吸收光谱表明配合物能以嵌入的方式与ct-DNA作用,结合常数Kb=1.5×104mol/L。

Nrf2 and Snail-1 in the prevention of experimental liver fibrosis by caffeine

AIM:To determine the molecular mechanisms involved in experimental hepatic fibrosis prevention by caffeine(CFA).METHODS:Liver fibrosis was induced in Wistar rats by intraperitoneal thioacetamide or bile duct ligation and they were concomitantly treated with CFA(15 mg/kg per day).Fibrosis and inflammatory cell infiltrate were evaluated and classified by Knodell index.Inflammatory infiltrate was quantified by immunohistochemistry(anti-CD11b).Gene expression was analyzed by quantitative reverse transcription-polymerase chain reaction for collagenⅠ?(Col-1),connective tissue growth factor(CTGF),transforming growth factorβ1(TGF-β1),tumor necrosis factor alpha(TNF-α),interleukin-1(IL-1),IL-6,superoxide dismutase(SOD)and catalase(CAT).Activation of Nrf2 and Snail-1 was analyzed by Westernblot.TNF-αexpression was proved by enzyme-linked immunosorbant assay,CAT activity was performed by zymography.RESULTS:CFA treatment diminished fibrosis index in treated animals.The Knodell index showed both lower fibrosis and necroinflammation.Expression of profibrogenic genes CTGF,Col-1 and TGF-β1 and proinflammatory genes TNF-α,IL-6 and IL-1 was substantially diminished with CFA treatment with less CD11b positive areas.Significantly lower values of transcriptional factor Snail-1 were detected in CFA treated rats compared with cirrhotic rats without treatment;in contrast Nrf2was increased in the presence of CFA.Expression of SOD and CAT was greater in animals treated with CFA showing a strong correlation between mRNA expression and enzyme activity.CONCLUSION:Our results suggest that CFA inhibits the transcriptional factor Snail-1,down-regulating profibrogenic genes,and activates Nrf2 inducing antioxidant enzymes system,preventing inflammation and fibrosis.

Caffeine therapy in preterm infants

Caffeine is the most commonly used medication for treatment of apnea of prematurity. Its effect has been well established in reducing the frequency of apnea, intermittent hypoxemia, and extubation failure in mechanically ventilated preterm infants. Evidence for additional short-term benefits on reducing the incidence of bronchopulmonary dysplasia and patent ductus arteriosus has also been suggested. Controversies existamong various neonatal intensive care units in terms of drug efficacy compared to other methylxanthines, dosage regimen, time of initiation, duration of therapy, drug safety and value of therapeutic drug monitoring. In the current review, we will summarize the available evidence for the best practice in using caffeine therapy in preterm infants.

Effects of Theophylline on Plasma Levels of Interleukin-4,Cyclic Nucleotides and Pulmonary Functions in Patients with Chronic Obstructive Pulmonary Disease

Summary: In order to measure the plasma levels of interleukin 4 (IL 4), cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP) in patients with chronic obstructive pulmonary disease (COPD) and observe the effects of oral theophylline on them, we divided 28 COPD patients into COPD experimental group and COPD control group. Plasma levels of IL 4, cAMP and cGMP as well as parameters of pulmonary function tests were measured in these 2 groups before and after 2 weeks of treatment with oral theophylline (Protheo, 400 mg, qd) or placebo. Plasma levels of IL 4 and cGMP were significantly elevated in patients with COPD as compared with normal controls ( P <0.05), while cAMP and cAMP/cGMP were significantly lower than those in controls ( P <0.01). Plasma level of IL 4 was inversely correlated with forced expiratory volume at the first second (FEV 1) and with maximum expiratory flow rate at 50 % of forced vital capacity (V 50 ) (both r =-0.46, P <0.05) while it was directly correlated with the scores of the clinical manifestations ( r =0.57, P <0.05) in COPD patients. Two weeks after treatment with theophylline, IL 4 and cGMP in COPD experimental group were decreased significantly while cAMP and cAMP/cGMP increased significantly ( P <0.05). The change of IL 4 was inversely correlated with the changes of FEV 1 and V 50 ( r =-0.53 and -0.54, respectively, P <0.05). Two weeks after placebo treatment, the COPD control group did not show such changes. We are led to conclude that IL 4 might play a role in the pathoge nesis of the airway inflammation and air flow limitation in COPD patients and the mechanisms of theophylline's therapeutic effects of attenuating air flow limitation may partially depend on its anti inflammatory effects on the airways which, in turn, is dependent on its inhibitory effects on some inflammatory mediators such as IL 4.

Caffeine Crystallization Induction Time Measurements Using Laser Scattering Technique and Correlation to Surface Tension in Water and Ethanol

Caffeine nucleation induction times were measured at 30 °C and 40 °C in water and ethanol solvents employing laser light absorption technique. Supersaturation concentrations and liquid/solid phase surface tensions were calculated from crystallization induction times using classic homogeneous nucleation theory. Induction time and surface tension decreased at higher temperature.

Detection and determination of undeclared synthetic caffeine in weight loss formulations using HPLC-DAD and UHPLC-MS/MS

Caffeine is present in products marketed for weight loss, with the purpose of increasing thermogenesis and lipid metabolism. The dosage declared by the product manufacturer, or even its presence, is not always correctly described on the label. This work aimed to investigate the undeclared synthetic caffeine in weight loss formulations by a high-performance liquid chromatography with diode array detection(HPLC-DAD) method. From one hundred products purchased through Brazilian e-commerce, seventeen contained caffeine, either naturally or synthetically added to formulation. The caffeine-containing samples were confirmed by an ultra-high performance liquid chromatography-tandem mass spectrometry(UHPLC-MS/MS) method, and adulteration was clearly proven in five products. The content highest caffeine contained 448.8 mg per dose. Other irregularities were also found; nevertheless, the most serious was the addition of synthetic drugs without asking the consumers. Additional drugs expose the consumer to more possible side-effects as well as deleterious drug interactions. Intentional adulteration with any unlabeled substance is typically motivated by a desire to increase or alter the claimed effect of the marketed product to gain a commercial advantage.

Quantification of theophylline or paracetamol in milk matrices by high-performance liquid chromatography

A simple, accurate and sensitive high-performance liquid chromatography （HPLC） method was developed, validated and applied to the determination of either theophylline or paracetamol in milk-based samples. The method allowed drug quantification in fresh and powdered milk with a relatively short run time of analysis and it was also successfully applied to the quantification of the drugs in solid dosage forms intended for pediatric use. Moreover, the main significant advantages over other published works are the simplicity of the sample preparation, reduced assay time and sample loss. The method meets the International Conference on Harmonization guideline for analytical methods validation regarding specificity, linearity, accuracy, precision, specificity and robustness as required by health authorities and applied by industry while designing and marketing new drug products. The technique encompasses the separation of the analytes with a reverse phase C18 column under isocrafic conditions and UV detection at 272 nm and 243 nm, respectively, for theophylline and paraeetamol. The lower limit of quantification for both drugs was determined as 0.2 μg/mL and the between-batch accuracy was 99.7%. This HPLC method allows quantification of theophylline and paracetamol in milk matrices and it can be applied in the design, development and production of milk-based pediatric dosage forms.

Simultaneous Determination of Catechins and Caffeine in Green Tea-Based Beverages and Foods for Specified Health Uses

Catechins in green tea have various useful features including antioxidant activity and preventive effects on metabolic syndrome. Various beverages that are enriched with tea catechins are marketed as Foods for Specified Health Uses (FOSHU) in Japan. However, recent reports have indicated that excessive consumption of green tea extracts as a dietary supplement are associated with adverse health effects such as liver disorders. Various catechins and caffeine are constituents of FOSHU tea-based beverages. The amount of catechins in FOSHU products is displayed on labels as total catechin content, but the content of individual catechins are not provided. Although health hazards of FOSHU products have rarely been reported, precise information about the content and types of catechins in FOSHU products is needed to ensure safety. We used high-performance liquid chromatography with a photodiode array (HPLC/PDA) to simultaneously identify and quantify catechins and caffeine in green tea-based popular beverages and FOSHU beverages. This technique allowed simultaneous quantitation of five types of catechins and caffeine in green tea without complicated sample preparation. Epigallocatechin gallate (EGCG) and epigallocatechin EGC were the main catechins in various FOSHU beverages and the concentrations of almost all catechins were higher in FOSHU, than in popular green tea-based beverages. The concentrations of EGCG in green tea-based popular beverages and in FOSHU beverages were 5.4 - 7.3 and 10.2 - 41.9 mg/100mL, respectively, with the highest concentration being in a product named Healthya (approximately 147 mg/bottle). The simultaneous determination of compounds such as catechins and caffeine in FOSHU beverages can help to estimate beneficial and adverse effects to prevent deleterious effects on health and the excessive consumption of FOSHU beverages containing high concentrations of tea catechins should be avoided.