Weiying Guo

Investigation on the spectrum-effect relationships of EtOAc extract from Radix Isatidis based on HPLC fingerprints and microcalorimetry.

This work investigated the spectrum-effect relationships between HPLC fingerprints and the anti-bacterial activities of EtOAc extracts from Radix Isatidis. Fingerprints of EtOAc extracts of Radix Isatidis from various sources were established by a High-Performance Liquid Chromatography. The process of Escherichia coli (E. coli) growth affected by EtOAc extracts was monitored using a Thermal Activity Monitor (TAM) Air Isothermal Calorimeter by microcalorimetry. By analyzing the power-time curves, quantitative parameters, such as growth rate constant k, maximum heat-production rate P(m), appearance time t and total heat-production Q were obtained to characterize the interactions of E. coli and the EtOAc extracts from Radix Isatidis. The HPLC fingerprints were investigated using hierarchical clustering analysis. The main thermo-kinetic parameters from the power-time curves were analyzed using principal component analysis. The spectrum-effect relationships between the HPLC fingerprints and anti-bacterial activities were analyzed with multivariant correlation analysis. Close correlation existed between the spectrum-effect relationships of the EtOAc extracts. Salicylic acid in the HPLC fingerprints might be one of the anti-bacterial components. This work provides a general model of the combination of HPLC and microcalorimetry to study the spectrum-effect relationships of EtOAc extracts from Radix Isatidis, which can be used to search for principal components of Radix Isatidis on bioactivity.

Multi-class mycotoxins analysis in Angelica sinensis by ultra fast liquid chromatography coupled with tandem mass spectrometry.

An ultra fast liquid chromatography coupled with tandem mass spectrometry (UFLC-MS/MS) method was developed and validated for simultaneous analysis of multi-class mycotoxins including aflatoxins (AFB1, AFB2, AFG1 and AFG2), ochratoxin A (OTA), fumonisins (FB1 and FB2) and zearalanone (ZEN) in 20 batches of Angelica sinensis samples collected from different markets and stores in China. The eight mycotoxins were extracted and cleaned up by using QuEChERS-based procedure, and then were quantified under the multiple reaction monitoring (MRM) together with positive and negative ionization modes. Focusing on the optimization of extraction and clean-up conditions, as well as UFLC separation and MS/MS parameters of targeted analytes, the developed method expressed good linearity for the eight mycotoxins within their respective linear ranges with correlation coefficients all higher than 0.9974. The limits of detection (LODs) and quantification (LOQs) ranged from 0.005 to 0.125 μg/kg and from 0.0625 to 0.25 μg/kg, respectively. Recoveries for spiked A. sinensis sample at three different levels were all above 78.9% with relative standard deviations (RSDs) below 6.36% for all analytes. Analysis of real samples demonstrated that two visibly moldy A. sinensis samples were detected with AFB1 of 2.07 and 2.92 μg/kg, and AFG1 of 2.84 and 1.53 μg/kg. The proposed quantitative method with significant advantages including simple pretreatment, rapid determination and high sensitivity would be the preferred candidate for the determination and quantification of multi-class mycotoxin contaminants in complex matrixes, which well fulfilled the maximum residue limits (MRLs) from various countries.

Microcalorimetric Studies of the Action on Four Organic Acids in Radix Isatidis on the Growth of Microorganisms

The actions of four organic acids in Radix isatidis, a traditional Chinese medicinal (TCM) herb, on Escherichia coli, Staphylococcus aureus and Shigella dysenteriae growth were investigated by microcalorimetry. The four organic acids were syringic acid, 2-amino-benzoic acid, salicylic acid and benzoic acid. The power-time curves of Escherichia coli, Staphylococcus aureus and Shigella dysenteriae growth with and without organic acids were acquired, meanwhile the extent and duration of inhibitory effects on the metabolism were evaluated by growth rate constants (k1, k2), maximum heat-output[0] power (P(m)) and peak time (t(m)). The inhibitory activity varied with different drugs. The sequences of anti-microbial activity of the four organic acids on Escherichia coli, Staphylococcus aureus and Shigella dysenteriae were all: syringic acid > 2-amino-benzoic acid > salicylic acid > benzoic acid. And benzoic acid promoted the growth of Staphylococcus aureus and Shigella dysenteriae. This study provides a basis for the further study on Radix Isatidis.

Quantitative and fingerprinting analysis of Atractylodes rhizome based on gas chromatography with flame ionization detection combined with chemometrics.

This study assessed the feasibility of gas chromatography with flame ionization detection fingerprinting combined with chemometrics for quality analysis of Atractylodes rhizome. We extracted essential oils from 20 Atractylodes lancea and Atractylodes koreana samples by hydrodistillation. The variation in extraction yields (1.33-4.06%) suggested that contents of the essential oils differed between species. The volatile components (atractylon, atractydin, and atractylenolide I, II, and III) were quantified by gas chromatography with flame ionization detection and confirmed by gas chromatography with mass spectrometry, and the results demonstrated that the number and content of volatile components differed between A. lancea and A. koreana. We then calculated the relative peak areas of common components and similarities of samples by comparing the chromatograms of A. lancea and A. koreana extracts. Also, we employed several chemometric techniques, including similarity analysis, hierarchical clustering analysis, principal component analysis, and partial least-squares discriminate analysis, to analyze the samples. Results were consistent across analytical methods and showed that samples could be separated according to species. Five volatile components in the essential oils were quantified to further validate the results of the multivariate statistical analysis. The method is simple, stable, accurate, and reproducible. Our results provide a foundation for quality control analysis of A. lancea and A. koreana.

UFLC-MS/MS analysis of four tanshinone components in Salvia miltiorrhizae after ultrasound-assisted extraction.

A sensitive and rapid ultra-fast liquid chromatography with tandem mass spectrometry (UFLC-MS/MS) method was developed for simultaneous qualitative and quantitative of four characteristic tanshione components including tanshinone IIA, cryptotanshinone, tanshinone I and dihydrotanshinone I in Salvia miltiorrhizae after ultrasound-assisted extraction. By using a C18 column, the four analytes were separated by gradient elution with acetonitrile and water both containing 0.1% formic acid at the flow rate of 0.3mL/min. Multiple-reaction monitoring (MRM) was used for quantification, and an information-dependent acquisition (IDA) method was used to trigger enhanced product ion scans (EPI) for supplementary characteristic identification for qualitative research. Calibration curves showed good linearities with correlation coefficients (r) higher than 0.9990. The method showed high sensitivity with limits of detection (LODs) and quantification (LOQs) less than 0.0002ng/mL and 0.0008ng/mL, respectively, as well as good precision and reproducibility. Mean recoveries for four analytes ranged from 92.5% to 106.2% with relative standard deviations (RSDs) lower than 14.59%. Real application of the developed method in 32 batches of S. miltiorrhizae samples demonstrated that the total contents of four analytes in all samples were in the range of 2.258-52.342mg/g. Ultrasound-assisted extraction technique took a small amount of sample and low time but giving high extraction efficiency. Combining with UFLC-MS/MS method in MRM-IDA-EPI mode, more components in other complicated matrices can be simultaneously analyzed for qualitation and quantitation in one run.

Combination of chemical fingerprint and bioactivity evaluation to explore the antibacterial components of Salvia miltiorrhizae

The aim of this study was to explore the possible antibacterial components of Salvia miltiorrhizae on Pseudomonas aeruginosa using a combination of chemical fingerprint and bioactivity evaluation. The chemical fingerprints of 32 batches of S. miltiorrhizae samples from different sources were developed using high-performance liquid chromatography with diode array detection, and then were evaluated by similarity analysis and hierarchical clustering analysis. Anti-P. aeruginosa activity was determined by microcalorimetry. Some crucial thermokinetic parameters obtained from the heat-flow power-time curves of P. aeruginosa growth in the absence or presence of these S. miltiorrhizae samples were evaluated using principal component analysis. Thereafter, multiple linear regression analysis was used to analyze the fingerprint-activity relationship between the chemical fingerprints and anti-P. aeruginosa activity. This established the related equation between the inhibition ratio (I, %) of S. miltiorrhizae samples on P. aeruginosa and the peak areas of the common peaks. The results showed that the 32S. miltiorrhizae samples could be grouped into three clusters according to their chemical fingerprints and anti-P. aeruginosa activities. Protocatechualdehyde, salvianolic acid B, together with three unidentified compounds might be the major components that contributed largely to the antibacterial properties of S. miltiorrhizae and should be the focus of S. miltiorrhizae quality control. Thus, this study provided a preferred way for exploring the bioactive components of medicinal plants.

Rapid gas chromatography with flame photometric detection of multiple organophosphorus pesticides in Salvia miltiorrhizae after ultrasonication assisted one-step extraction

A simple and rapid gas chromatography-flame photometric detection (GC-FPD) method was developed for the determination of 12 organophosphorus pesticides (OPPs) in Salvia miltiorrhizae by using ultrasonication assisted one-step extraction (USAE) without any clean-up steps. Some crucial parameters such as type of extraction solvent were optimized to improve the method performance for trace analysis. Any clean-up steps were negligent as no interferences were detected in the GC-FPD chromatograms for sensitive detection. Under the optimized conditions, limits of detection (LODs) and quantitation (LOQs) for all pesticides were in the range of 0.001-0.002mg/kg and 0.002-0.01mg/kg and 0.002-0.01mg/kg, respectively, which were all below the regulatory maximum residue limits suggested. RSDs for method precision (intra- and inter-day variations) were lower than 6.8% in approval with international regulations. Average recovery rates for all pesticides at three fortification levels (0.5, 1.0 and 5.0mg/kg) were in the range of 71.2-101.0% with relative standard deviations (RSDs) <13%. The developed method was evaluated for its feasibility in the simultaneous pre-concentration and determination of 12 OPPs in 32 batches of real S. miltiorrhizae samples. Only one pesticide (dimethoate) out of the 12 targets was simultaneously detected in four samples at concentrations of 0.016-0.02mg/kg. Dichlorvos and omethoate were found in the same sample from Sichuan province at 0.004 and 0.027mg/kg, respectively. Malathion and monocrotophos were determined in the other two samples at 0.014 and 0.028mg/kg, respectively. All the positive samples were confirmed by LC-MS/MS. The simple, reliable and rapid USAE-GC-FPD method with many advantages over traditional techniques would be preferred for trace analysis of multiple pesticides in more complex matrices.

Differentiation of essential oils in Atractylodes lancea and Atractylodes koreana by gas chromatography with mass spectrometry

Atractylodes rhizome is a valuable traditional Chinese medicinal herb that comprises complex several species whose essential oils are the primary pharmacologically active component. Essential oils of Atractylodes lancea and Atractylodes koreana were extracted by hydrodistillation, and the yield was determined. The average yield of essential oil obtained from A. lancea (2.91%) was higher than that from A. koreana (2.42%). The volatile components of the essential oils were then identified by a gas chromatography with mass spectrometry method that demonstrated good precision. The method showed clear differences in the numbers and contents of volatile components between the two species. 41 and 45 volatile components were identified in A. lancea and A. koreana, respectively. Atractylon (48.68%) was the primary volatile component in A. lancea, while eudesma-4(14)-en-11-ol (11.81%) was major in A. koreana. However, the most significant difference between A. lancea and A. koreana was the major component of atractylon and atractydin. Principal component analysis was utilized to reveal the correlation between volatile components and species, and the analysis was used to successfully discriminate between A. lancea and A. koreana samples. These results suggest that different species of Atractylodes rhizome may yield essential oils that differ significantly in content and composition.