Determination of Mogroside V in Luohanguo Extract for Daily Quality Control Operation Using Relative Molar Sensitivity to Single-Reference Caffeine.

Mogroside V is one of the characteristic and effective components of luohanguo extract, a food additive used as a sweetener in Japan as per Japan's Standards and Specifications for Food Additives (JSFA; 9th ed.). JSFA stipulates that the quantitative determination for mogroside V content in luohanguo extract applies HPLC using analytical standard mogroside V. However, no mogroside V reagents with proven purities are commercially available. Therefore the current JSFA determination method is not particularly suited for daily quality control operations involving luohanguo extract. In this study, we applied an alternative quantitative method using a single reference with relative molar sensitivity (RMS). It was possible to calculate the accurate RMS by an offline combination of 1H-quantitative NMR spectroscopy (1H-qNMR) and an HPLC/variable-wavelength detector (VWD). Using the RMS of mogroside V to a commercial certified reference material grade caffeine, the mogroside V contents in luohanguo extracts could be determined using HPLC/VWD without analytical standard mogroside V. There was no significant difference between the mogroside V contents in luohanguo extracts determined using the method employing single-reference caffeine with the RMS and using the JSFA method. The absolute calibration curve for the latter was prepared using an analytical standard mogroside V whose purity was determined by 1H-qNMR. These results demonstrate that our proposed method using a single reference with RMS is suitable for quantitative determination of mogroside V in luohanguo extract and can be used as an alternative method to the current assay method in JSFA.

Caffeine in the Diet: Country-Level Consumption and Guidelines.

Coffee, tea, caffeinated soda, and energy drinks are important sources of caffeine in the diet but each present with other unique nutritional properties. We review how our increased knowledge and concern with regard to caffeine in the diet and its impact on human health has been translated into food-based dietary guidelines (FBDG). Using the Food and Agriculture Organization list of 90 countries with FBDG as a starting point, we found reference to caffeine or caffeine-containing beverages (CCB) in 81 FBDG and CCB consumption data (volume sales) for 56 of these countries. Tea and soda are the leading CCB sold in African and Asian/Pacific countries while coffee and soda are preferred in Europe, North America, Latin America, and the Caribbean. Key themes observed across FBDG include (i) caffeine-intake upper limits to avoid risks, (ii) CCB as replacements for plain water, (iii) CCB as added-sugar sources, and (iv) health benefits of CCB consumption. In summary, FBDG provide an unfavorable view of CCB by noting their potential adverse/unknown effects on special populations and their high sugar content, as well as their diuretic, psycho-stimulating, and nutrient inhibitory properties. Few FBDG balanced these messages with recent data supporting potential benefits of specific beverage types.

Consumption of caffeinated beverages and the awareness of their caffeine content among Dutch students.

The purpose of the current study was to examine the knowledge of caffeine content of a variety of caffeinated beverages among Dutch university students. A pencil-and-paper survey was conducted among N = 800 Dutch students. Most participants (87.8%) reported consuming caffeinated beverages during the past 24 h. Their mean ± SD past 24-h caffeine intake from beverages was 144.2 ± 169.5 mg (2.2 ± 3.0 mg/kg bw). Most prevalent sources of caffeine were coffee beverages (50.8%) and tea (34.8%), followed by energy drink (9.2%), cola (4.7%), and chocolate milk (0.5%). Participants had poor knowledge on the relative caffeine content of caffeinated beverages. That is, they overestimated the caffeine content of energy drinks and cola, and underestimated the caffeine content of coffee beverages. If caffeine consumption is a concern, it is important to inform consumers about the caffeine content of all caffeine containing beverages, including coffee and tea. The current findings support previous research that the most effective way to reduce caffeine intake is to limit the consumption of coffee beverages and tea.

Internal standard application to dried blood spots by spraying: investigation of the internal standard distribution.

BACKGROUND: The scientifically and logistically best way of application of the internal standard (IS) in the analysis of dried blood spots (DBS) analysis is still a matter of debate and investigation. Most commonly the IS is added in the solvent used for extraction of the discs punched from DBS. In this case, the recovery of the non-extracted IS is complete while the recovery of the analyte extracted from DBS is different from the IS. RESULTS: An alternative way for addition of the IS was investigated. A homogeneous distribution and absorption of the test compound across the spots was demonstrated by spraying a solution of a radiolabeled test compound (mimicking an IS solution) onto DBS. CONCLUSION: This spray-on technique is convenient and easily automatable. Spraying of the solution was rapid, precise and reproducible, and therefore seems to be suitable for routine analysis of DBS by offline and online extraction.

Universal method for online enrichment of target compounds in capillary gas chromatography using in-oven cryotrapping.

A method is described that permits automated online enrichment of injected compounds in multidimensional gas chromatography by using a microfluidic heart-cut (H-C) device to direct target compounds into a cryogenically cooled internal trap (cryotrap, CT). By performing multiple injections of a sample, selected compounds or regions of a primary column separation can be collected in the CT. Remobilizing the trapped species allows elution and further resolution on the second column. Using a well-balanced H-C device, compounds can be fully excluded from the collection step or quantitatively transferred to the CT. Peak areas of the remobilized compound correlate well with the number of sample injections. Trapping on various column phases shows the method is suited to quantitative trapping of alkanes of mass greater than about dodecane and fatty acid methyl esters greater than the C8 homologue. Caffeine and menthol standards of concentration 100 µg mL(-1) gave peak area correlation coefficients for 1-10 and 1-50 replicate split injections of 1 µL volume of 0.999 and 0.996, respectively. Peak height correlations were less favorable as a result of peak broadening on the second column, presumably due to overloading at greater collected mass. The method was applied to 0.2% solutions of peppermint oil (menthol; a major component; 44%) and 1.0% lavender oil (α-terpineol and neryl acetate; minor components of 1.05 and 0.42% abundance). The minor components gave good area and height correlations, and good recovery around 90% was observed for menthol compounds recovered from 15 accumulations. Response amplification was further demonstrated for menthol from mint oil headspace sampling using solid phase microextraction. This approach should be a valuable adjunct for improved detection specificity, for detectors of low sensitivity, and when prior sample concentration provides insufficient response of selected target analytes.

Caffeine (1, 3, 7-trimethylxanthine) in foods: a comprehensive review on consumption, functionality, safety, and regulatory matters.

Caffeine ranks as one of the top most commonly consumed dietary ingredients throughout the world. It is naturally found in coffee beans, cacao beans, kola nuts, guarana berries, and tea leaves including yerba mate. The total daily intake, as well as the major source of caffeine varies globally; however, coffee and tea are the 2 most prominent sources. Soft drinks are also a common source of caffeine as well as energy drinks, a category of functional beverages. Moderate caffeine consumption is considered safe and its use as a food ingredient has been approved, within certain limits, by numerous regulatory agencies around the world. Performance benefits attributed to caffeine include physical endurance, reduction of fatigue, and enhancing mental alertness and concentration. Caffeine has also been recently linked to weight loss and consequent reduction of the overall risks for developing the metabolic syndrome. However, the caloric contribution of caffeine-sweetened beverages needs to be considered in the overall energy balance. Despite all these benefits the potential negative effects of excessive caffeine intake should also be considered, particularly in children and pregnant women.

Evaluación de Tecnología Sanitaria: límite máximo recomendable de cafeína contenida en las bebidas energizantes / Health Technology Assessment: recommended maximum limit of caffeine contained in energy drinks

Objetivo: Conocer la evidencia científica y las recomendaciones de organismos expertos en el tema sobre el límite máximo recomendable de cafeína en las bebidas energizantes. Metodología: Se realizó una búsqueda en las bases de datos bibliográficas (Medline, Cochrane, Tripdatabase, LILACS), en buscadores genéricos de Internet como Google, agencias de evaluación de tecnologías sanitarias y agencias nacionales e internacionales reguladoras de alimentos y medicamentos. Conclusiones: La evidencia encontrada en estudios primarios sobre la seguridad de la cafeína contenida en las bebidas energizantes ha sido de baja calidad, con eventos adversos surgidos principalmente de reportes aislados de casos u opiniones de expertos, por lo que no es posible recomendar con precisión un límite de seguridad para el contenido de cafeína en estas bebidas. Los límites recomendados por las diferentes agencias reguladoras varían entre 32­35 mg/100 mL, aunque algunas requieren el rotulado de "alto contendido de cafeína" cuando éste supera los 15­ 18mg/100 mL. Otras agencias fijan además un límite mínimo de cafeína (25 mg/100mL) para considerarse bebida energizante. La mayoría de las recomendaciones derivadas de las agencias reguladoras también se basa en evidencia de baja calidad. Los datos encontrados sugieren una interacción entre la cafeína y el alcohol, que podría depender de la dosis, en relación a una menor percepción de la intoxicación alcohólica por los tomadores de bebidas energizantes, con un patrón de co­consumo entre los jóvenes, tanto en estudios locales como internacionales. No se ha encontrado suficiente evidencia para recomendar un límite máximo de cafeína en las bebidas energizantes

Stereospecific determination of cis- and trans-resveratrol in rat plasma by HPLC: application to pharmacokinetic studies.

A simple, accurate, precise, specific and reproducible high-performance liquid chromatography (HPLC) method was developed for simultaneous determination of resveratrol isomers in rat plasma. Cis-resveratrol was made by exposure of a trans-resveratrol solution to sunlight for 5 days followed by separation by HPLC and identification by mass spectrometry (MS). The assay procedure involved simple liquid-liquid extraction of resveratrol isomers and internal standard (IS, caffeine) from a small plasma volume directly into acetonitrile. The supernatant liquid was added an equal volume of water and injected onto a Hypersil ODS(2) C(18) column (5 microm, 4.6 x 250 mm). Mobile phase consisting of methanol and distilled water was used at a flow rate of 1.0 mL/min for the effective separation of cis-, trans-resveratrol and caffeine (IS). The detection of the analyte peak was achieved by monitoring the eluate using a UV detector set at 303 nm. The ratio of peak area of analyte to IS was used for quantification of plasma samples. Nominal retention times of cis-, trans-resveratrol and IS were 3.2, 4.3 and 6.1 min, respectively. The calibration curve was linear ranging from 0.066 to 6.64 and 0.134 to 13.4 microg/mL with correlation coefficients of 0.9998 and 0.9997 for trans and cis isomers, respectively. The absolute recovery of both isomers was more than 85%. The inter- and intra-day precisions in the measurement of quality control (QC) samples, 0.066, 0.664 and 6.64 microg/mL of trans-resveratrol, were in the range 2.37-6.95% relative standard deviation (RSD) and 0.77-6.97% RSD, respectively. The inter- and intra-day precisions in the measurement of quality control (QC) samples, 0.134, 1.34 and 13.4 microg/mL of cis-resveratrol, were in the range 1.93-3.72% relative standard deviation (RSD) and 1.13-6.57% RSD, respectively. Both analytes and IS were stable in the battery of stability studies and freeze-thaw cycles. Resveratrol isomers were found to be stable for a period of 30 days on storage at -20 degrees C. The application of the assay to determine the pharmacokinetic disposition after a single oral dose to rats is described.

[Current problems in the quality control of pharmaceutical preparations manufactured in pharmacies II. Paracetamol contraining preparations]. / Magisztráilis gyógyszerkészítmények minoségellenorzésének aktuális kérdései II. Paracetamol tartalmú készítmények tartalmi meghatározáisi lehetoségei.

In our previous paper identification methods using chemical reactions and TLC are described for the active ingredients of pharmaceutical preparations in Formula Normales (FoNo VII.). Present paper introduces the development and validation of analytical methods suitable for quantitative determination of paracetamol containing dosage forms in FoNo VII. Titrimetric methods, UV spectroscopy and HPLC are used for assay of paracetamol and accompanying components (e.g. codeine, papaverine, caffeine and acetylsalicylic acid) in these preparations.

[Near infrared determination of the content of caffeine in tea polyphenol].

In the present paper the caffeine in the tea polyphenol was analysed spectrally and quantitatively by using near infrared spectroscopy. From the original absorbance of caffeine in the tea polyphenol an obvious and strong peak can be viewed. By using second derivative, MSC (multiple scatter correction) and correlation analysis the spectral characteristics of caffeine in the near infrared region can be seen very clearly, thus the robust calibration model can be set up easily. The result obtained shows that through this technique the absorptive characteristic of those primary fundamentals of caffeine can be looked through easily, meanwhile, calibration test was performed to quantitatively measure the weight percent of caffeine in the tea polyphenol, and fine precision of the result was obtained in a comparatively very large range of concentration. The SEC(standard error of calibration) is 0.49%, and the correlation coefficient r is 0.993. The result shows that NIR is feasible and superior in analyzing the content of caffeine in tea polyphenol.

Assessment of CYP1A2 activity in clinical practice: why, how, and when?

The cytochrome P450 enzyme CYP1A2 mediates the rate-limiting step in the metabolism of many drugs including theophylline, clozapine, and tacrine as well as in the bioactivation of procarcinogens. CYP1A2 activity shows both pronounced intra- and interindividual variability, which is, among other factors, related to smoking causing enzyme induction, to drug intake and to dietary factors which may result in induction or inhibition. In contrast to these exogenous factors, genetic influences on enzyme activity seem to be less pronounced. Therefore, phenotyping of CYP1A2, i.e. the determination of the actual activity of the enzyme in vivo, represents a useful approach both for scientific and clinical applications. CYP1A2 is almost exclusively expressed in the liver. Since liver tissue cannot be obtained for direct phenotyping, a probe drug which is metabolized by CYP1A2 has to be given. Proposed probe drugs include caffeine, theophylline, and melatonin. Caffeine is most often used because of the predominating role of CYP1A2 in its overall metabolism and the excellent tolerability. Various urinary, plasma, saliva, and breath based CYP1A2 caffeine metrics have been applied. While caffeine clearance is considered as the gold standard, the salivary or plasma ratio of paraxanthine to caffeine in a sample taken approximately 6 hr after a defined dose of caffeine is a more convenient, less expensive but also fully validated CYP1A2 phenotyping metric. CYP1A2 phenotyping is applied frequently in epidemiologic and drug-drug interaction studies, but its clinical use and usefulness remains to be established.

[Ratio spectrophotometry and its application to assay of compound pharmacy preparations].

The present paper is to study ratio spectrophotometry for its application to the assay of compound pharmacy preparations. According to the feature of ratio spectra, the wavelength pair was selected at the peak point or valley point of ratio spectra to establish ratio spectrophotometry for the determination of Co-Chlorzoxazone tablet and injection of caffeine and sodium benzoate. The average recoveries and relative standard deviations for Chlorzoxazone and paracetamol were 100.0% and 1.28%, and 100.0% and 0.84%, respectively; the average recoveries and relative standard deviations for caffeine and sodium benzoate were 99.98% and 0.70%, and 99.91% and 0.78%, respectively. It is concluded that as long as the ratio spectra exhibit at least one peak and one valley, the results are satisfactory.

Correction of accurate mass measurement for target compound verification by quadrupole time-of-flight mass spectrometry.

The aim of this work is to evaluate quadrupole/time-of-flight (QTOF) mass spectrometry for simultaneous measurement of accurate mass and quantification of a target by using a stable isotopically labeled internal standard. Mixtures of caffeine and (13)C(3)-caffeine (internal standard) at different concentration ratios were analyzed by capillary HPLC/QTOF. A calibration plot for quantification is linear over a factor of 20. Without invoking any correction scheme, the mass accuracy seriously degraded when the ratio of the mass standard to the test compound was not unity. The accuracy could be restored to approximately 2 ppm by using a quadratic function to correct the measured mass as a function of the measured signal ratio of target and internal calibrant.

In vitro predictions of skin absorption of caffeine, testosterone, and benzoic acid: a multi-centre comparison study.

To obtain better insight into the robustness of in vitro percutaneous absorption methodology, the intra- and inter-laboratory variation in this type of study was investigated in 10 European laboratories. To this purpose, the in vitro absorption of three compounds through human skin (9 laboratories) and rat skin (1 laboratory) was determined. The test materials were benzoic acid, caffeine, and testosterone, representing a range of different physico-chemical properties. All laboratories performed their studies according to a detailed protocol in which all experimental details were described and each laboratory performed at least three independent experiments for each test chemical. All laboratories assigned the absorption of benzoic acid through human skin, the highest ranking of the three compounds (overall mean flux of 16.54+/-11.87 microg/cm(2)/h). The absorption of caffeine and testosterone through human skin was similar, having overall mean maximum absorption rates of 2.24+/-1.43 microg/cm(2)/h and 1.63+/-1.94 microg/cm(2)/h, respectively. In 7 out of 9 laboratories, the maximum absorption rates of caffeine were ranked higher than testosterone. No differences were observed between the mean absorption through human skin and the one rat study for benzoic acid and testosterone. For caffeine the maximum absorption rate and the total penetration through rat skin were clearly higher than the mean value for human skin. When evaluating all data, it appeared that no consistent relation existed between the diffusion cell type and the absorption of the test compounds. Skin thickness only slightly influenced the absorption of benzoic acid and caffeine. In contrast, the maximum absorption rate of testosterone was clearly higher in the laboratories using thin, dermatomed skin membranes. Testosterone is the most lipophilic compound and showed also a higher presence in the skin membrane after 24 h than the two other compounds. The results of this study indicate that the in vitro methodology for assessing skin absorption is relatively robust. A major effort was made to standardize the study performance, but, unlike in a formal validation study, not all variables were controlled. The variation observed may be largely attributed to human variability in dermal absorption and the skin source. For the most lipophilic compound, testosterone, skin thickness proved to be a critical variable.

Extractionless method for the simultaneous high-performance liquid chromatographic determination of urinary caffeine metabolites for N-acetyltransferase 2, cytochrome P450 1A2 and xanthine oxidase activity assessment.

Urinary metabolic ratios of caffeine are used in humans to assess the enzymatic activities of cytochrome P450 isoenzyme 1A2 (CYP1A2), xanthine oxidase (XO) and for phenotyping individuals for the bimodal N-acetyltransferase 2 (NAT2), all of them involved in the activation or detoxification of various xenobiotic compounds. Most reported analytical procedures for the measurement of the urinary metabolites of caffeine include a liquid-liquid extraction of urine samples prior to their analysis by reversed-phase HPLC. At neutral to basic pH however, 5-acetylamino-6-formylamino-3-methyluracil (AFMU), a metabolite of caffeine, spontaneously decomposes to 5-acetylamino-6-amino-3-methyluracil (AAMU). Since AAMU is not extracted in most organic solvents, the extent of AFMU decomposition cannot be precisely assessed. Although the decomposition reaction can be minimized by immediate acidification of the urine, accurate results can only be obtained when both AAMU and AFMU are monitored, or alternatively, if AAMU is measured after complete transformation of AFMU into AAMU in basic conditions. We report a liquid chromatographic method for the simultaneous quantitative analysis of the five urinary metabolites of caffeine used for the CYP1A2, XO and NAT2 phenotyping studies: AAMU, AFMU, 1-methylxanthine, 1-methyluric acid and 1,7-dimethyluric acid. These metabolites are satisfactory separated from all other known caffeine metabolites as well as endogenous urinary constituents. Sample treatment does not require any liquid-liquid extraction procedure. Urine samples are diluted and centrifuged before being injected (10 microl) onto a YMC-Pack Polyamine II (250x4.6 mm) column. A step-wise gradient elution program is applied using acetonitrile-0.75% (v/v) formic acid: (91:9) at 0 min-->(75:25) at 25 min-->(65:35) at 35 min-->(65:35) at 45 min, followed by a re-equilibration step to the initial solvent composition. The flow-rate is 1.0 ml/min and the separations are monitored by UV absorbance at 260 and 280 nm. The procedure described here represents a substantial improvement over previous methods: a single analysis and a minimal urine sample treatment enables the simultaneous quantitation of five caffeine metabolites, notably AFMU and AAMU, used for the determination of CYP450 1A2, XO and NAT2 enzyme activity. Importantly enough, phenotyping individuals for the bimodal NAT2 is made possible without the uncertainty associated with the deformylation of AFMU, which is likely to happen at all steps prior to the analysis, during sample storage and even in the bladder of the subjects.

Caffeine and health

Today, huge numbers of people everywhere consume caffeine to change or enhance the way they feel and act. Given its universal appeal consumers ought to become more familiar with its history and health consequences. What are the origins of caffeine? Are there any health benefits from its ingestion? Is there a down side? This issue of Nyam News provides information on the roots of this stimulating ingredient and its relationship to health.

[HPC purity tests for caffeine, theophylline and theobromine]. / Koffein, teofillin, teobromin tisztaságvizsgálata HPLC módszerrel.

After giving a survey of the literature of the HPLC methods for the determination of caffeine, theophylline and theobromine two methods are presented for their purity test, i.e. their detection in each other at the trace level. In the reversed-phase system the stationary phase was C18-silica and the 2:3 mixture of methanol and water was used as the eluent at a detection wavelength of 254 nm. The k' values for theobromine, theophylline and caffeine are 0,25, 0,62 and 1,12, respectively. This system enables the detection of 0,2% of theobromine and 0,1% of theophylline in caffeine and 0,2% of caffeine in theobromine or theophylline. In the normal phase system silica was used as the stationary phase while the eluent was 85:15:0,05 mixture of chloroform, dioxane and formic acid. The wavelength of the detection was 273 nm. The k' values of caffeine, theobromine and theophylline are 1,5 4,0 and 5,6, respectively. This system also enables the detection of 0,1% of theophylline and theobromine in caffeine or 0,2% of caffeine in the former. Theophyline and theobromine can be detected in each other at the 0,1% level.