Chinese Medicine Samples Research Articles

Dual-Readout Immunochromatographic Assay by Utilizing MnO2 Nanoflowers as the Unique Colorimetric/Chemiluminescent Probe.

Manganese dioxide nanoflowers (MnO2 NFs) were synthesized and used as a dual readout probe to develop a novel immunochromatographic test strip (ITS) for detecting pesticide residues using chlorpyrifos as the model analyte. MnO2 NFs-labeled antibody for chlorpyrifos was employed as the signal tracer for conducting the ITS. After 10 min competitive immunoreaction, the tracer antibody was captured by the immobilized immunogen in the test strip, resulting in the captured MnO2 NFs on test line. The captured MnO2 NFs led to the appearance of brown color on the test line, which could be easily observed by the naked eye as a qualitative readout. Due to the very slight colorimetric difference of chlorpyrifos at trace concentrations, the semiquantitative readout by naked eyes could not meet the demand of quantitative analysis. MnO2 NFs showed a significant effect on the luminol-H2O2 chemiluminescent (CL) system, and the CL signal driven by MnO2 NFs were used to detect the trace concentration of chlorpyrifos quantitatively. 1,3-Diphenylisobenzofuran quenching studies and TMB-H2O2 coloration assays were conducted for studying the enhancing mechanism of MnO2 NFs, which was based on the oxidant activity to decompose H2O2 for forming reactive oxygen species. Under optimal conditions, the linear range of chlorpyrifos was 0.1-50 ng/mL with a low detection limit of 0.033 ng/mL (S/N = 3). The reliability of the dual-readout ITS was successfully demonstrated by the application on traditional Chinese medicine and environmental water samples. Due to the simultaneous rapid-qualitative and sensitive-quantitative detection, the dual-readout protocol provides a promising strategy for rapid screening and field assay on various areas such as environmental monitoring and food safety.

Colorimetric and chemiluminescent dual-readout immunochromatographic assay for detection of pesticide residues utilizing g-C3N4/BiFeO3 nanocomposites

Graphitic carbon nitride/bismuth ferrite nanocomposites (g-C3N4/BiFeO3 NCs) were synthesized by a facile one step sol-gel combustion method and employed as a peroxidase-like catalyst. Based on the catalytical activity on the luminol-H2O2 reaction, the nanocomposites were utilized as a colorimetric/chemiluminescent dual-readout immunochromatographic assay (ICA) for the multiplexed detection of pesticide residues by utilizing chlorpyrifos and carbaryl as the model analytes. In the proposed protocol, chlorpyrifos antibody and carbaryl antibody were tagged to g-C3N4/BiFeO3 NCs for developing the spatially-resolved multianalyte ICA. After two competitive immunoreactions completed on the ICA test strip, the tracer antibodies were captured by the immobilized antigens on two test lines. The accumulation of g-C3N4/BiFeO3 NCs led to the appearance of brown color, which were observed as a colorimetric and semi-quantitative signal. Furthermore, the g-C3N4/BiFeO3 NCs-driven generation of CL signal was collected as a sensitively quantitative signal after initiating the luminol-H2O2 reaction on the test lines. Under the optimal conditions, the limits of detection of chlorpyrifos and carbaryl were both 0.033ng/mL. The dual-readout ICA was successfully used to detect chlorpyrifos and carbaryl spiked in environmental water and traditional Chinese medicine samples with acceptable recovery values of 80–119% and 90–118%. Due to many advantages including low cost, time efficiency, high sensitivity and good portability, the novel ICA showed great potential in many areas such as drug safety, environmental monitoring and clinical diagnosis.

In-Syringe Binary-Solvent Liquid-Phase Microextraction for the Preconcentration of Cinnamic Acid Derivatives in Traditional Chinese Medicine Samples

An in-syringe binary-solvent liquid-phase microextraction (in-syringe BSLPME) coupled to high-performance liquid chromatography with diode-array detection (HPLC/DAD) has been established and applied for the determination of four cinnamic acid derivatives (caffeic acid, p-hydroxycinnamic acid, ferulic acid, and cinnamic acid) in traditional Chinese medicine samples. In this method, 70 µL of n-pentanol/n-hexanol (5:5, v:v) mixture as extractant were added to the surface of a solution containing target analytes in a disposable syringe and the solution was manually shaken for a desired time. After the re-aggregation of binary-solvent droplets, the extraction phase was impelled out of the syringe for HPLC analysis. The variables influencing in-syringe BSLPME, such as extraction solvents (types, volume ratios, and volumes), extraction time, sample phase pH, ionic strength, sample volume, and disperser solvent, were investigated. Under the optimized conditions, the limits of detection were between 0.2 and 1.3 ng mL−1 with enrichment factors from 28 to 58. The calibration graphs were linear in the range of 1.0 × 10−2–7.0 µg mL−1 for caffeic acid, 1.3 × 10−3–1.9 µg mL−1 for p-hydroxycinnamic acid, 2.8 × 10−3–4.1 µg mL−1 for ferulic acid, and 2.7 × 10−3–4.1 µg mL−1 for cinnamic acid, with r 2 better than 0.9912. The precision expressed as relative standard deviation varied between 1.4 and 9.6%. In-syringe BSLPME was verified reliable for the determination of cinnamic acid derivatives in Chinese herbal medicines with recovery values from 94.2 to 106.7%. The proposed method was compared with other dispersive liquid-phase microextraction methods. Results showed that the developed method was easy-to-handle, efficient, and rapid for the analysis of active constituents in Chinese medicine samples.

Fibroin/dodecanol floating solidification microextraction for the preconcentration of trace levels of flavonoids in complex matrix samples.

A new fibroin/dodecanol floating solidification microextraction, coupled with high performance liquid chromatography, was developed and applied for enrichment and quantification of the trace flavonoids in traditional Chinese medicine and biological samples. Also, fibroin sensibilization mechanism was described, and influence of sample matrix to enrichment factor was investigated. In this method, a homogeneous fibroin/dodecanol of dispersed solution was employed as microextraction phase to flavonoids (myricetin, quercetin, isorhamnetin, chrysin, kaempferide), the several critical parameters affecting the performance, such as organic extractant, amount of fibroin in organic extractant, volume of extraction phase, dispersant, salt concentration, pH of sample phase, stirring rate, extraction time, and volume of sample phase were tested and optimized. Under the optimized conditions, enrichment factor of flavonoids ranged from 42.4 to 238.1 in different samples, excellent linearities with r2≥ 0.9968 for all analytes were achieved, limits of detection were less than or equal to 5.0ng/mL, average recoveries were 92.5% to 115.0% in different samples. The new procedure is simple, fast, low cost, environmentally friendly and high EF, it can also be applied to the concentration and enrichment of the trace flavonoids in other complex matrixes.

Development of a Matrix Solid-Phase Dispersion Extraction Combined with UPLC/Q-TOF-MS for Determination of Phenolics and Terpenoids from the Euphorbia fischeriana.

A method based on a simplified extraction by matrix solid phase dispersion (MSPD) followed by ultra-performance liquid chromatography coupled with the quadrupole time-of-flight tandem mass spectrometry (UPLC/Q-TOF-MS) determination is validated for analysis of two phenolics and three terpenoids in Euphorbia fischeriana. The optimized experimental parameters of MSPD including dispersing sorbent (silica gel), ratio of sample to dispersing sorbent (1:2), elution solvent (water–ethanol: 30–70) and volume of the elution solvent (10 mL) were examined and set down. The highest extraction yields of chromatogram information and the five compounds were obtained under the optimized conditions. A total of 25 constituents have been identified and five components have been quantified from Euphorbia fischeriana. A linear relationship (r2 ≥ 0.9964) between the concentrations and the peak areas of the mixed standard substances were revealed. The average recovery was between 92.4% and 103.2% with RSD values less than 3.45% (n = 5). The extraction yields of two phenolics and three terpenoids obtained by the MSPD were higher than those of traditional reflux and sonication extraction with reduced requirement on sample, solvent and time. In addition, the optimized method will be applied for analyzing terpenoids in other Chinese herbal medicine samples.

Highly sensitive determination of paeonol using porous alumina microfibers modified electrode

A kind of porous alumina microfibers was prepared via hydrothermal reaction, and then used to modify the carbon paste electrode (CPE). Compared with the unmodified CPE, the alumina microfibers modified CPE is more active for the oxidation of paeonol, and greatly increases the oxidation signals of paeonol. The oxidation mechanism of paeonol on the alumina microfibers modified CPE was studied. The oxidation process involves one electron and one proton. Based on the remarkable signal enhancement effects of alumina microfibers, a highly sensitive electrochemical method was developed for the determination of paeonol. The linear range is from 10 to 1000nM, and the detection limit is 6.7nM. It was used in the analysis of traditional Chinese medicine samples. The results obtained by this method and high-performance liquid chromatography are in good agreement.

Trace determination of five triazole fungicide residues in traditional Chinese medicine samples by dispersive solid-phase extraction combined with ultrasound-assisted dispersive liquid-liquid microextraction and UHPLC-MS/MS.

A novel and reliable method for determination of five triazole fungicide residues (triadimenol, tebuconazole, diniconazole, flutriafol, and hexaconazol) in traditional Chinese medicine samples was developed using dispersive solid-phase extraction combined with ultrasound-assisted dispersive liquid-liquid microextraction before ultra-high performance liquid chromatography with tandem mass spectrometry. The clean up of the extract was conducted using dispersive solid-phase extraction by directly adding sorbents into the extraction solution, followed by shaking and centrifugation. After that, a mixture of 400μL trichloromethane (extraction solvent) and 0.5mL of the above supernatant was injected rapidly into water for the dispersive liquid-liquid microextraction procedure. The factors affecting the extraction efficiency were optimized. Under the optimum conditions, the calibration curves showed good linearity in the range of 2.0-400 (tebuconazole, diniconazole, and hexaconazole) and 4.0-800ng/g (triadimenol and flutriafol) with the regression coefficients higher than 0.9958. The limit of detection and limit of quantification for the present method were 0.5-1.1 and 1.8-4.0ng/g, respectively. The recoveries of the target analytes ranged from 80.2 to 103.2%. The proposed method has been successfully applied to the analysis of five triazole fungicides in traditional Chinese medicine samples, and satisfactory results were obtained.

Selective and sensitive determination of As(III) and tAs in Chinese herbal medicine samples using L-cysteine modified carbon paste electrode-based electrolytic hydride generation and AFS analysis

A novel non-chromatographic speciation technique for ultra-trace arsenite [As(III)] and total arsenic (tAs) in Chinese herbal medicine (CHM) is developed and validated by electrolytic hydride generation (EHG) coupled with atomic fluorescence spectrometry (AFS). The studies show that As(III) can be converted efficiently to AsH3 on an L-cysteine modified carbon paste electrode (CMCPE), which has never been reported before. Significantly, other arsenic species such as arsenate [As(V)], monomethylarsonic acid (MMA) and dimethylarsinic acid (DMA) do not form any or only less volatile hydrides at low applied current mode (<1.0 A). The results also demonstrate that L-cysteine and graphite powder play different roles in the electrolytic generation of AsH3. Comparing with the traditional graphite electrode, CMCPE has better stability, sensitivity and interference tolerance. Under the optimal conditions, the limit of detection (LOD) of tAs and As(III) for this method are 0.087µgL−1 and 0.095µgL−1 respectively. The accuracy of the method is verified through the analysis of reference materials (CRM 08231 and SRM1568a), and the proposed method has been applied satisfactorily to the determination of As(III) and tAs in several CHM samples.

Multiplexed immunochromatographic test strip for time-resolved chemiluminescent detection of pesticide residues using a bifunctional antibody

A novel bifunctional antibody (BfAb) that could recognize methyl parathion and imidacloprid simultaneously was prepared by a hybrid hybridomas technique. Using the BfAb as the sole recognition reagent, a multiplexed immunochromatographic test strip based on a time-resolved chemiluminescence (CL) strategy was developed for quantitative detection of pesticide residues. Horseradish peroxidase (HRP) and alkaline phosphatase (ALP) were used as the CL probes to label the haptens of methyl parathion and imidacloprid, respectively. After the labeled haptens competed with methyl parathion and imidacloprid to bind with the BfAb immobilized on the test strip, the two CL reactions catalyzed by the enzymes were triggered simultaneously by coreactants injection. Due to the distinct CL kinetics characteristics of HRP and ALP, the signals for methyl parathion and imidacloprid detections were collected at 2.5s and 300s, respectively. The linear ranges for methyl parathion and imidacloprid were both 0.1–250ngmL−1, with detection limits of 0.058ngmL−1 (S/N=3). The whole assay process could be accomplished within 22min. The detection results for spiked traditional Chinese medicine samples demonstrated its application potential. The proposed method provided a low-cost, facile and rapid tool for multiplexed screening of pesticide residues using single antibody.

An ultra-facile and label-free immunoassay strategy for detection of copper (II) utilizing chemiluminescence self-enhancement of Cu (II)-ethylenediaminetetraacetate chelate

The establishment of facile, rapid, sensitive and cost-effective protocols for the detection of heavy metals is of great significance for human health and environmental monitoring. Hereby, an ultra-facile and label-free immunoassay strategy was designed for detecting heavy metal ion by using Cu (II) as the model analyte. Cu (II) reacted previously with ethylenediaminetetraacetate (EDTA) was captured by immobilized monoclonal antibody for Cu (II)-EDTA chelate. Then Cu (II) was detected based on the self-enhancing effect of Cu (II)-EDTA chelate to luminol-H2O2 chemiluminescence reaction. The CL intensity is linear relative with Cu (II) concentration in a very wide range of 1.0–1000ng/mL, with a detection limit of 0.33ng/mL (S/N=3). Since the specificity of this proposed strategy relied on both the specificity of monoclonal antibody and the specificity of luminol-H2O2 system, it could avoid interference from most common ions. The proposed method was used successfully to detect Cu (II) in traditional Chinese medicine and environmental water samples with acceptable recovery values of 82–113%. This proof-of-principle work demonstrated the feasibility of the label-free immunoassay for heavy metal ions, and opened a new avenue for rapid screening and field assay for drug safety, environmental monitoring and clinical diagnosis.

Exploring bitterness of traditional Chinese medicine samples by potentiometric electronic tongue and by capillary electrophoresis and liquid chromatography coupled to UV detection

Instrumental bitterness assessment of traditional Chinese herbal medicine (TCM) preparations was addressed in this study. Three different approaches were evaluated, high-performance liquid chromatography coupled to UV detector (HPLC), capillary electrophoresis coupled to UV detector (CE) and a potentiometric multisensor system – electronic tongue (ET). Most studies involving HPLC and CE separations use these as selective instruments for quantification of individual substances. However we employed these techniques to provide chromatographic or electrophoretic sample profiles. These profiles are somewhat analogous to the profiles produced by the ET. Profiles from all instruments were then related to professional sensory panel evaluations using projections on latent structures (PLS) regression. It was found that all three methods allow for bitterness assessment in TCM samples in terms of human sensory panel with root mean squared errors of prediction ca. 0.9 within bitterness scale from 0 (no bitterness) to 6 (maximal bitterness).

Integration of hydride generation and photochemical vapor generation for multi-element analysis of traditional Chinese medicine by ICP-OES

A dual-mode chemical vapor generation (DM-CVG) sample introduction system that integrates hydride generation (HG) and photochemical vapor generation (PVG) was coupled to a commercial inductively coupled plasma optical emission spectrometer (ICP-OES) for simultaneous multi-element analysis of traditional Chinese medicine samples for trace hydride-forming elements and non–hydride-forming elements. Three hydride-forming elements (Cd, As, and Ge) and two non–hydride-forming but photochemical vapor–generating elements (Co and Ni) were selected as target elements to validate the proposed method. Optimum experimental conditions for DM-CVG-ICP-OES were carefully investigated, and satisfactory limits of detection (LODs), 0.2–1.4μgL−1, and relative standard deviations (RSDs), 1.3%–4.6%, were obtained for these target elements. A certified reference celery sample (GBW10048) was used to investigate the feasibility of the proposed method for real analysis. Strong suppression on the PVG efficiency occurred either when little nitric acid residue or some unknown coexisting materials remained in the sample solutions. Complete acid-evaporation/catching and moderate dilution of the digests could effectively alleviate this negative effect. Four different acid-evaporation/catching methods were performed and compared. Catching acid by adding an appropriate volume of formic acid gave the best signal response and the results agreed well with the certified values. Satisfactory analytical results were also obtained in real sample analysis of TCM Curcuma wenyujin Y.H. Chen et C. Ling.

Microscopic Images Restoration of Chinese Herbal Medicine Using the TV Model

Chinese herbal medicine (CHM) samples defiled, extraction of microscopic images, collection site light not equal and images collected uneven illumination point which can make the acquisition of CHM microscopic images sub- ject to different unfavorable factors. Those often make microscopic images of CHM become blurred or introduce noise. In this paper, some restoration methods for the microscopic images of CHM using the total variation (TV) model are put forward. Firstly, the microscopic images of CHM are blurred utilizing the two dimensional different Gaussian convolution kernels and different Gauss white noise are added, and then the damaged images are restored by using the improved TV model. The TV restoration results are compared with the results using Wiener filtering, median filtering method etc. The theoretical analysis and experimental results show that the novel method is effectively able to restore image, and maintain the image detail information.

Preparation of fluorine functionalized magnetic nanoparticles for fast extraction and analysis of perfluorinated compounds from traditional Chinese medicine samples

Magnetic solid-phase extraction for the enrichment of PFCs from sample solution.

Evaluation of the mutagenicity of a traditional Chinese medicine Radix isatidis root by modified Ames tests, SOS/umu and SOS/rec tests

Radix isatidis root (RIR), the root of plant Isatis indigotica Fort., is a traditional Chinese medicine (TCM) that has been commonly used in China to control upper respiratory tract infections. However, the genetic toxicity of RIR has not been well studied and is the focus of this paper. RIR gave positive results in the Ames test at some doses; and gave positive results in modified Ames tests at a few doses. RIR gave negative results in SOS/umu-lacZ tests. In addition, RIR gave negative results in the mammalian bone marrow chromosomal aberration test and TK gene mutation test. Because the concentrations of RIR in the above tests were all greatly higher than the commonly clinical dose, it could be summarized that at the commonly used clinical dose, RIR showed a negative response in the Ames test, the umu/lacZ test, the TK gene mutation test and the mammalian bone marrow cell chromosomal aberration test. Moreover, different tests sometimes gave conflicting results which may be due to their different sensitivities. This suggests that it is important to choose the appropriate test method when assessing the mutagenicity of traditional Chinese medicine samples.

Quantitative analysis of catalpol in chinese patent medicine Lixin pill by near-infrared diffuse reflectance spectroscopy

Lixin pill is a typical Chinese patent medicine with anti-rheumatic heart disease activity that has been widely used in clinical practice. Therefore it is very important to detect the concentration of catalpol, as the main component of the active ingredient. Near-infrared reflectance(NIR) spectroscopy was used to study the content of catalpol in the unprocessed Chinese patent medicine of Lixin pills. NIR is applied to quantitatively analyze 77 samples, which were randomly divided into a calibration set containing 61 samples and a prediction set containing 16 samples. To get a satisfying result, partial least squares(PLS) regression was utilized to establish quantitative models. In PLS regression, the values of coefficient of determination(R2) and root mean square error of cross-validation (RMSECV) of PLS regression are 0.9419 and 0.0216, respectively. The process of establishing model, parameters of model, and prediction results were also discussed in detail(root mean square error of prediction is 0.0164). The overall results show that NIR spectroscopy can be efficiently utilized for the rapid and accurate analysis of routine chemical compositions in the Chinese patent medicine of Lixin pills. The prediction set suggests that this quantitative analysis model has excellent generalization ability and prediction precision. Accordingly, the result can provide technical support for the further analysis of catalpol in unprocessed Lixin pill. Moreover, this study supplied technical support for the further analysis of other Chinese patent medicine samples.

A new strategy based on cholesterol-functionalized iron oxide magnetic nanoparticles for determination of polycyclic aromatic hydrocarbons by high-performance liquid chromatography with cholesterol column

This study reported for the first time the use of cholesterol-functionalized magnetic nanoparticles (Fe3O4@SiO2@Chol) for the determination of polycyclic aromatic hydrocarbons (PAHs) in traditional Chinese medicine samples (TCMs) by high performance liquid chromatography (HPLC) coupled with fluorescence detection. The method was efficient, environmentally friendly, and fast. The solvent consumption of the proposed column is only half of the conventional column but with higher efficiency. Influencing factors, including sorbent amount, desorption solvent, sample volume and extraction time, were investigated in detail. Under the optimum conditions, good linearity (R2>0.991) was obtained over the range of 5–400ngg−1, with limits of detection (LOD) 0.75, 0.50, 1.0, 0.56, 0.60, 0.84 and 0.80ngg−1 for anthracene, fluoranthene, pyrene, chrysene, benzo[a]anthracene, benzo(b)fluoranthene and benzo(k)fluoranthene, respectively.

Multi-residual Pesticide Monitoring in Commercial Chinese Herbal Medicines by Gas Chromatography–Triple Quadrupole Tandem Mass Spectrometry

To investigate multi-residual pesticide monitoring data in commercial Chinese herbal medicines on major markets, an easy, rapid, and selective gas chromatography with mass spectrometry (GC/MS/MS) method was established for simultaneously determining multi-residual pesticides including organochlorine, pyrethroid, carbamate, and organophosphorus pesticides in Chinese herbal medicines. The analytical method was based on an efficient extraction procedure and further cleanup steps by solid-phase extraction columns, yielding recovery rates in the range of 70.0–120.0 % for the majority of pesticides, except for hexachlorobenze, diazinon, β-HCH, δ-HCH, and omethoate, with precision values expressed as relative standard deviation of 0.1–14.7 %. The limits of detection of the established GC/MS/MS method for all investigated pesticides ranged from 0.01 to 3.6 μg kg−1 and limits of quantification from 0.03 to 11.88 μg kg−1. With this validated method, multi-residual pesticides of 132 Chinese herbal medicine samples were analyzed. The monitoring results indicated that pesticide residue was found in 74 samples. In total, 51 pesticides were found with detection rate ranging from 0.76 to 18.94 %. An 82.3 % of positive pesticides were found in less than 6 % of samples. Hexachlorobenzene was found in 25 samples, quintozene in 15 samples, and acephate and simazine in 13 samples. Concentrations of pesticide residue from monitoring data obtained ranged from 0.5 to 203.5 μg kg−1. The simple and rapid method can be used as routine analysis method in multi-residual pesticide monitoring of Chinese herbal medicines.

Near infrared spectroscopy as a tool for the rapid analysis of the Honeysuckle extracts

There is a lack of methods to analyze large numbers of traditional Chinese medicine (TCM) samples quickly and efficiently. Here we present a near infrared (NIR) spectroscopy-based method for the rapid analysis of Honeysuckle extracts. NIR spectra of 124 Honeysuckle extract samples produced within a four-year period were collected in the diffuse reflection mode and pretreated with multiplicative scatter correction (MSC), smoothing and derivative. Using partial least squares regression (PLSR), including PLS1 and PLS2, we established and validated the calibration models of six organic acids. Our results showed that both accuracy and precision of the NIR spectroscopy-based method meet the requirements of a traditional Chinese medicine (TCM) factory. This new method has the potential to save time, efforts and cost associated with the analysis of large quantities of unknown samples and will be particularly applicable to TCM factories for cost reduction.

Chemometric resolution of coeluting peaks of eleven antihypertensives from multiple classes in high performance liquid chromatography: A comprehensive research in human serum, health product and Chinese patent medicine samples

A novel chemometric-assisted high performance liquid chromatography method coupled with diode array detector (HPLC-DAD) was presented for the simultaneous determination of eleven antihypertensives from multiple classes in most concerned matrix systems. With the aid of second-order calibration which enables specific information of analytes to be well extracted, the heavily overlapping profiles between analytes and the coeluting interferences can be successfully separated and thus accurately quantified. A great advantage of the novel strategy lies in the fact that the analysis could be carried out with the same isocratic mobile phase (methanol/KH2PO4: 58:42, v/v, pH 2.60) in a short time regardless of the changes of matrices, such as human serum, health product and Chinese patent medicine. Both qualitative and quantitative results indicate that the hybrid strategy that using HPLC-DAD coupled with second-order chemometric method would be a high performance approach for the purpose of simultaneously quantifying multiple classes of antihypertensives in complex systems. Additionally, the analytical strategy can potentially benefit drug monitoring in both therapeutic research and pharmaceutical quality control. Moreover, the accuracy and reliability of the proposed methodology has been evaluated using several statistical parameters such as root mean squared error of prediction (RMSEP), figures of merit (FOM) and reproducibility of inter-day analysis.