Warifteine and methylwarifteine: 1H and 13C assignments by two‐dimensional NMR spectroscopy

The identity and structures of warifteine and methylwarifteine, two tertiary bisbenzylisoquinoline alkaloids of an unusual type, were elucidated and verified by 2D NMR spectroscopy. Complete ¹H and ¹³C NMR spectral assignments were obtained. Copyright © 2003 John Wiley & Sons, Ltd.     

Toxicological evaluation in silico and in vivo of secondary metabolites of Cissampelos sympodialis in Mus musculus mice following inhalation

The ethanolic extract of the leaves of Cissampelos sympodialis showed great pharmacological potential, with inflammatory and immunomodulatory activities, however, it showed some toxicological effects. Therefore, this study aims to verify the toxicological potential of alkaloids of the genus Cissampelos through in silico methodologies, to develop a method in LC-MS/MS verifying the presence of alkaloids in the infusion and to evaluate the toxicity of the infusion of the leaves of C. sympodialis when inhaled by Swiss mice. Results in silico showed that alkaloid 93 presented high toxicological potential along with the products of its metabolism. LC-MS/MS results showed that the infusion of the leaves of this plant contained the alkaloids warifteine and methylwarifteine. Finally, the in vivo toxicological analysis of the C. sympodialis infusion showed results, both in biochemistry, organ weights and histological analysis, that the infusion of C. sympodialis leaves presents a low toxicity.     

High-performance liquid chromatography quantitative analysis of ephedrine alkaloids in Ephedrae Herba on a perfluorooctyl stationary phase

Ephedrae Herba is one of the most commonly used herbal medicines, and it has been shown that most of the clinical efficacy for cold and asthma is exerted by its alkaloidal components. A simple and sensitive high-performance liquid chromatography method was developed using a perfluorooctyl column for the simultaneous determination of five alkaloids (norephedrine, norpseudoephedrine, ephedrine, pseudoephedrine, and methylephedrine) in Ephedrae Herba. The mobile phase comprising acetonitrile and 15 mM ammonium trifluoroacetate was used to elute the targets in isocratic elution mode. The method was validated for linearity (R² > 0.999), repeatability, intraday and interday precision, recoveries with trueness (93.87–110.99%), limits of detection (5.35–5.76 µg/mL), and limits of quantification (20 µg/mL). The quantitative results revealed that the developed method was precise and accurate. Then it was successfully applied to determine the difference in the contents of three batches of Ephedrae Herba from three pharmaceutical companies.     

Development of an HPLC‐DAD method for the determination of five alkaloids in Stephania yunnanensis Lo and in rat plasma after oral dose of Stephania yunnanensis Lo extracts

For the rational utilization and the quantitative quality control of the Stephania yunnanensis Lo, an HPLC‐DAD method was developed for the quantitative and simultaneous determination of five alkaloids in rat plasma (stepharine, sinomenine, palmatine, isocorydine and tetrahydropalmatine), which were the main active chemical constituents of this plant and belong to four kinds of isoquinoline‐type alkaloids (protoberberine, morphine, aporphine and protaporphine alkaloids). The contents of five alkaloids ranged from 0.09 to 2.32% (w/w). The method validation was tested for the linearity (r² > 0.9975), precision (intra‐day RSD < 4.8% and inter‐day RSD < 4.9%), extraction recovery (85.49 ± 2.29% to 99.21 ± 1.48%) and stability (98.5 ± 5.3% to 101.2 ± 3.4%). We developed an HPLC‐DAD method to simultaneously measure these alkaloids in rat plasma after oral administration of the extract of this plant to rats. The results supported the hypothesis that isoquinoline alkaloids were the compounds responsible for the main pharmacological activities for anti‐inflammatory and analgesic.     

A biochemometrics strategy combining quantitative determination,bioactivity evaluation and relationship analysis for identification of analgesic alkaloids of raw and vinegar‐processed Corydalis turtschaninovii

A biochemometrics strategy combining quantitative determination, bioactivity evaluation, and relationship analysis was proposed for identification of analgesic components of herbs. First, a robust liquid chromatography tandem mass spectrometry method was developed for simultaneous determination of nine major alkaloids in crude and vinegar‐processed Corydalis turtschaninovii. Nine alkaloids were separated on a BEH C₁₈ column with a mobile phase consisting of acetonitrile and water spiked with 0.1% formic acid and then detected by multiple reactions monitoring in the positive ion mode. Nitidine chloride was employed as the internal standard. The method displayed good linearity and the precisions of intra‐day and inter‐day were all within 3.0%. The recovery rates of each alkaloid ranged from 97.1 to 102.9%. The method was successfully applied for quantitative analysis of nine alkaloids in ten batches of crude and vinegar‐processed Corydalis turtschaninovii. Second, the analgesic effects of crude and vinegar‐processed Corydalis turtschaninovii were evaluated in mice. Third, principle component analysis, canonical correlation analysis, and partial least squares regression were used to analysis the relationship between the contents of nine major alkaloids and the analgesic effect of different crude and vinegar‐processed samples. Tetrahydropalmatine, coptisine, and dehydrocorydaline have a close positive correlation with the analgesic effect.     

Nonaqueous CE for Rapid and Sensitive Determination of Matrine and Oxymatrine in Sophora flavescens and Its Medicinal Preparations

A simple, rapid, and sensitive non-aqueous capillary electrophoresis procedure for the quantitative determination of matrine and oxymatrine is established. Optimum separation conditions were obtained when the sample was injected under pressure for 3 s at 50 mbar and separated with the buffer containing 70 mM ammonium acetate, 7.0% (v/v) acetic acid, and 10% (v/v) acetonitrile in methanol medium at 25 kV applied voltage. The analytes were detected at 205 nm. The two alkaloids can be separated within 12 min and quantified with high sensitivity. The method was validated in terms of reproducibility, linearity, and accuracy when applied to the analysis of matrine and oxymatrine in Sophora flavescens and its medicinal preparations.     

Nonaqueous CE ESI‐IT‐MS analysis of Amaryllidaceae alkaloids

The Amaryllidaceae are widely distributed medical plants. Lycorine, lycoramine, lycoremine, and lycobetaine are the major active alkaloids in Amaryllidaceae plants. A nonaqueous CE ESI‐IT‐MS method for separation, identification, and quantification of the Amaryllidaceae alkaloids has been developed. The MS^1–3 behavior has been studied and the fragmentation pathways of main fragment ions have been proposed. The effects of several factors such as composition and concentration of buffer, applied voltage, composition, and flow rate of the sheath liquid, nebulizing gas pressure, flow rate, and temperature of drying gas were investigated. Under the optimal conditions, the linear concentration range of these compounds was wide with the correlation coefficient (R²) >0.99. RSDs of migration time and peak areas were <10%. The LODs were <240 ng/mL. The proposed method can be successfully applied to the determination of the related alkaloids in the Lycoris radiata roots.     

Quantitative Determination of Three Protoberberine Alkaloids in Jin-Guo-Lan by HPLC-DAD

An accurate and simple HPLC method for the simultaneous determination of three protoberberine alkaloids (columbamine, jatrorrhizine, and palmatine) contained in Chinese medicine Jin-Guo-Lan (Tinospora sagittata Oliv. and Tinospora capillipes Gagnep) is presented in this study. The herb samples from six main origins and three herb markets were investigated. The separation was performed on a YMC-C₁₈ ODS column at 30°C with a gradient elution program. Acetonitrile and phosphate buffer (0.02 mol L⁻¹ sodium dihydrogen phosphate and 0.01 mol L⁻¹ triethylamine, pH 3) were used as mobile phases and the flow rate was set at 1 mL min⁻¹. The recovery of the method was in the range of 99.43–100.96%, and all the alkaloids showed good linearity (r ² > 0.9997) in the relatively wide concentration ranges. The developed method was applied to the determination of these alkaloids in the collected herb samples, and the results showed that the contents of these components in Jin-Guo-Lan varied greatly from habitat to habitat. It was demonstrated that the proposed method was helpful for the quality evaluation of Chinese medicine Jin-Guo-Lan.     

LC Separation and Determination of Five Diester-Diterpenoid Alkaloids in the Unprocessed and Processed Aconite Roots

A reliable liquid chromatographic method with photodiode array detector (DAD) was developed and validated for simultaneous separation and determination of five diester-diterpenoid alkaloids in the aconite roots. The separation was successfully performed on a Zorbax Extend-C₁₈ column with a mobile phase gradient prepared from methanol and ammonia solution at a flow rate of 1.0 mL min⁻¹. Good linearity (r > 0.999) was observed over the concentration ranges investigated, and intra-day and inter-day precision were high. The mean recoveries of five components ranged from 90.45 to 102.63% and relative standard deviations were always <5%. The validated method was successfully used for simultaneous determination of the five diester-diterpenoid alkaloids of unprocessed and processed aconite roots. The quantitative method provided a scientific basis for safety assurance and clinical application of aconite roots.

     

Development of thermogravimetric method to quantitative determination of mebendazole

The objective of this work was to develop and validate a fast and reproducible method able to determine the concentration of mebendazole in raw materials and tablets. The samples were analyzed by dynamic thermogravimetry, in the heating rates of 10, 20, 40, 60 and 80°C min^–1, in the atmospheres of nitrogen and nitrogen with synthetic air. Obtained data were used in the equations of Antoine and Langmuir, with the purpose to get the pressure curves of those. Vapor pressure curves of drug and tablet of mebendazole were evaluated using the mathematical indexes of difference factor, f ₁, and similarity factor, f ₂, to compare its profiles. The data showed that there is no significant difference between the vapor pressure profiles of drug and tablet of mebendazole in both environmental conditions studied, what confirms that the process is really vaporization. The concentration of mebendazole was determined in the raw material and tablets with the drug.     

Development of thermogravimetric method for quantitative determination of metronidazole

The objective of this work was to develop and validate a fast and reproducible method to determine the concentration of metronidazole in drug substance and tablets. The samples were analyzed by dynamic thermogravimetry, using 10, 20, 40, 60 and 80°C min^–1 heating rates in nitrogen and in nitrogen with synthetic air. Obtained data were used in the Antoine and Langmuir equations in order to have the pressure curves. Vapor pressure curves of drug and tablet of metronidazole were evaluated using the mathematical indexes of difference factor, f ₁, and similarity factor, f ₂, to compare their profiles. The data showed that there is no significant difference between the vapor pressure profiles of drug and tablet of metronidazole in both environmental conditions, which confirms that the process is really vaporization. The concentration of metronidazole was determined in the raw material and tablets of the drug.     

Characterization of some essential oils and their key components: Thermoanalytical techniques

The present study was aimed at determining the kinetics of evaporation and establishing vapor pressure curves for both single and multi-component systems by thermogravimetry (TG) and differential scanning calorimetry (DSC). Essential oils (e.g. lavender oil, orange oil, clove oil and eucalyptus oil, etc.) are typically multi-component systems consisting of various volatile pure components (e.g. linalyl acetate, limonene, cinnamaldehyde, etc.) which resemble single component systems. In this study linalyl acetate was taken as the calibration compound for TG. The vapor pressure curves for the pure substances were plotted using TG and vapor pressure plots for clove oil and eucalyptus oil were constructed using DSC. The thermodynamic and kinetic parameters of the pure compounds were compared to that of the multi-component systems to quantitatively and qualitatively measure the influence of different compounds on each other. The k-value from the vapor pressure data for linalyl acetate was calculated as 112006 Pa kg^0.5mol^0.5s^-1 m^-2 K^-0.5. The vapor pressure values were used to determine the Antoine constants using the SPSS 10.0 software.This revised version was published online in November 2005 with corrections to the Cover Date.     

LC Separation and Determination of Five Diester-Diterpenoid Alkaloids in the Unprocessed and Processed Aconite Roots

A reliable liquid chromatographic method with photodiode array detector (DAD) was developed and validated for simultaneous separation and determination of five diester-diterpenoid alkaloids in the aconite roots. The separation was successfully performed on a Zorbax Extend-C₁₈ column with a mobile phase gradient prepared from methanol and ammonia solution at a flow rate of 1.0 mL min^?1. Good linearity (r > 0.999) was observed over the concentration ranges investigated, and intra-day and inter-day precision were high. The mean recoveries of five components ranged from 90.45 to 102.63% and relative standard deviations were always <5%. The validated method was successfully used for simultaneous determination of the five diester-diterpenoid alkaloids of unprocessed and processed aconite roots. The quantitative method provided a scientific basis for safety assurance and clinical application of aconite roots.     

Organic,inorganic and total mercury determination in fish by chemical vapor generation with collection on a gold gauze and electrothermal atomic absorption spectrometry

A method for organic, inorganic and total mercury determination in fish tissue has been developed using chemical vapor generation and collection of mercury vapor on a gold gauze inside a graphite tube and further atomization by electrothermal atomic absorption spectrometry. After drying and cryogenic grinding, potassium bromide and hydrochloric acid solution (1 mol L^− 1 KBr in 6 mol L^− 1 HCl) was added to the samples. After centrifugation, total mercury was determined in the supernatant. Organomercury compounds were selectively extracted from KBr solution using chloroform and the resultant solution was back extracted with 1% m/v L-cysteine. This solution was used for organic Hg determination. Inorganic Hg remaining in KBr solution was directly determined by chemical vapor generation electrothermal atomic absorption spectrometry. Mercury vapor generation from extracts was performed using 1 mol L^− 1 HCl and 2.5% m/v NaBH₄ solutions and a batch chemical vapor generation system. Mercury vapor was collected on the gold gauze heated resistively at 80 °C and the atomization temperature was set at 650 °C. The selectivity of extraction was evaluated using liquid chromatography coupled to chemical vapor generation and determination by inductively coupled plasma mass spectrometry. The proposed method was applied for mercury analysis in shark, croaker and tuna fish tissues. Certified reference materials were used to check accuracy and the agreement was better than 95%. The characteristic mass was 60 pg and method limits of detection were 5, 1 and 1 ng g^− 1 for organic, inorganic and total mercury, respectively. With the proposed method it was possible to analyze up to 2, 2 and 6 samples per hour for organic, inorganic and total Hg determination, respectively.     

An HPLC method for simultaneous quantitative determination of seven secoiridoid glucosides separated from the roots of Ilex pubescens

A simple and specific high‐performance liquid chromatographic method has been developed and validated to simultaneously determine seven secoiridoid glucosides for the first time. Three of them were separated from the ethanolic extract of the roots of Ilex pubescens for the first time, namely nuezhenide A, ligusides B and oleonuezhenide. In quantitative analysis, all of the calibration curves showed good linear regression (r > 0.999) within the tested ranges, and the mean recoveries of three different concentrations ranged from 97.6 to 101.2%. The limit of detection and limit of quantification were <4.18 and 11.63 ng mL⁻¹, respectively. The relative standard deviation for repeatability and the precision of seven analytes were <3.4 and 1.9%, respectively. The established method was successfully applied to simultaneous determination of seven secoiridoid glucosides in 11 batches of samples collected from different habitats in China.     

An optode membrane for determination of gold using a simple light-emitting diode-based device

Abstract  

A selective optode based on immobilization of 5-(p-dimethylaminobenzylidene)rhodanine on a triacetylcellulose membrane was developed for quantitative determination of Au(III). The determination procedure was performed using a simple light-emitting diode (LED)-based device as a new effort to overcome low reproducibility and repeatability problems which usually accompany optode-based determinations. The results obtained were compared with those of conventional spectrophotometric methods. The response characteristics of the sensor including dynamic range, reproducibility, response time, and lifetime are discussed in detail. This sensor was used for the determination of Au(III) in ore and electroplating liquid effluent samples with satisfactory results in comparison with flame atomic absorption spectroscopy as standard method. Under the optimum conditions, the calibration plot was linear in the concentration range of 0.3–6.0 μg cm⁻³. The relative standard deviations for five replicate determinations of 1.0 μg cm⁻³ Au(III) and the corresponding limits of detection were 1.76% and 0.12 μg cm⁻³, respectively.     

Thermodynamic study on the sublimation of diphenyl and triphenyl substituted acetic and propanoic acids

Knudsen mass-loss effusion technique was used for measuring the vapor pressures at different temperatures of the following crystalline compounds: diphenylacetic acid, between 357.27 and 379.08 K; triphenylacetic acid, between 418.98 and 436.97 K; 2,2-diphenylpropanoic acid, between 366.08 and 386.00 K; 3,3-diphenylpropanoic acid, between 366.09 and 386.03 K; 3,3,3-triphenylpropanoic acid, between 402.17 and 420.10 K. From the temperature dependence of the vapor pressure of each crystalline compound, the standard (p ⁰ = 10⁵ Pa) molar enthalpies and Gibbs energies of sublimation, at T = 298.15 K, were derived. The measured thermodynamic properties are compared with literature results for phenylacetic and phenylpropanoic acids and correlations for estimation of the vapor pressures from the enthalpy of sublimation and the temperature of fusion of these and other compounds are presented.     

Development and Validation of a Stability-Indicating LC Method for Determination of Ebastine in Tablet and Syrup

A simple stability-indicating reversed-phase liquid chromatographic method with diode-array detection was developed and validated for the quantitative determination of ebastine in tablets and syrup. The LC method was carried out on a C₁₈ column with acetonitrile:phosphoric acid 0.1% pH 3.0 (55:45, v/v) as mobile phase, at a flow rate of 1.2 mL min⁻¹. Ultraviolet detection of ebastine was at 254 nm. A linear response (r = 0.9999) was observed in the range of 10–80 μg mL⁻¹. The RSD values for intra- and inter-day precision studies showed good results (RSD < 2%) and accuracy was greater than 98%. Validation parameters such as specificity and robustness were also determined. The method was found to be stability-indicating and can be applied to quantitative determination of ebastine in tablets and syrup.     

Anti-fungal effect of berberine on <Emphasis Type="Italic">Candida albicans</Emphasis> by microcalorimetry with correspondence analysis

Using a LKB-2277 bioactivity monitor, stop-flow mode, the power–time curves of Candida albicans growth at 37 °C affected by berberine were measured. The check experiments were studied based on agar cup method to observe the inhibitory diameter and serial dilution method to determine the minimal inhibitory concentration (MIC) of berberine on C. albicans growth. By analyzing the quantitative thermogenic parameters taken from the power–time curves using correspondence analysis (CA), we could find that berberine at a low concentration (5.0 μg mL⁻¹) began to inhibit the growth of C. albicans and at a high concentration (75.0 μg mL⁻¹) completely inhibited C. albicans growth. The anti-fungal activity of berberine could also be expressed as half-inhibitory concentration IC₅₀, i.e., 50% effective in this inhibition. The value of IC₅₀ of berberine on C. albicans was 34.52 μg mL⁻¹. The inhibitory diameters all exceeded 10 mm in test range and the MIC was 500 μg mL⁻¹. Berberine had strong anti-fungal effect on C. albicans growth. This work provided an important idea of the combination of microcalorimetry and CA for the study on anti-fungal effect of berberine and other compounds. Compared with the agar cup method and serial dilution method, microcalorimetry not only offered a useful way for evaluating the bioactivity of drugs, but also provides more information about the microbial growth and all this information was significant for the synthesis and searching of antibiotics.     

Simultaneous analysis of ten low‐molecular‐mass organic acids in the tricarboxylic acid cycle and photorespiration pathway in Thalassiosira pseudonana at different growth stages

A method using high‐performance liquid chromatography coupled with tandem mass spectrometry was developed for the simultaneous determination of organic acids in microalgae. o‐Benzylhydroxylamine was used to derivatize the analytes, and stable isotope‐labeled compounds were used as internal standards for precise quantification. The proposed method was evaluated in terms of linearity, recovery, matrix effect, sensitivity, and precision. Linear calibration curves with correlation coefficients >0.99 were obtained over the concentration range of 0.4–40 ng/mL for glycolic acid, 0.1–10 ng/mL for malic acid and oxaloacetic acid, 0.02–2 ng/mL for succinic acid and glyoxylic acid, 4–400 ng/mL for fumaric acid, 20–2000 ng/mL for isocitric acid, 2–200 ng mL⁻¹ for citric acid, 100–10000 ng mL⁻¹ for cis‐aconitic acid, and 1–100 ng mL⁻¹ for α‐ketoglutaric acid. Analyte recoveries were between 80.2 and 115.1%, and the matrix effect was minimal. Low limits of detection (0.003–1 ng/mL) and limits of quantification (0.01–5 ng/mL) were obtained except cis‐aconitic acid. Variations in reproducibility for standard solution at three different concentrations levels were <9%. This is the first report of the simultaneous analysis of ten organic acids in microalgae, which promotes better understanding of their growth state and provides reference value for high‐yield microalgae cultures.