[Research progress of pretreatment technology for mycotoxin detection in Chinese materia medica and complex matrices].

In the processes of planting, harvest, transport and storage, improper treatment of Chinese materia medica (CMM) and foodstuffs and agricultural products will result in fungal growth and mycotoxins contamination, which will not only directly affect the quality, safety and efficacy of these complex matrices, but also seriously threaten the consumers' health and lives. Therefore, the establishment of high-throughout analytical methods with high sensitivity for the determination of mycotoxins in CMM and foodstuffs and agricultural products at trace levels will provide reliable references for reducing the risk of mycotoxin exposure in humans. Due to the matrix complexity of CMM and foodstuffs and agricultural products, highly-effective pretreatment technologies are necessary for the establishment of such analytical techniques. In this review, the current extraction and purification methods commonly used for the detection of mycotoxins were summarized, the importance of pretreatment techniques for the precise quantification of mycotoxins in complex matrices such as Chinese herbal medicines was highlighted, as well as the development tendency about the pretreatment techniques for mycotoxins in complex matrices in the future was proposed.

[Contamination distribution and control measures of aflatoxins in Chinese herbal pieces].

As an important part of traditional medicine in China, traditional Chinese medicine(TCM) plays a significant role because of its unique medical efficiency, less adverse reactions and extensive resources. However, in recent years, the aflatoxins in medicinal herbs have been detected excessive both at home and abroad, seriously affecting the reputation and credibility of traditional Chinese medicine. In this paper, the current status of aflatoxins contamination in medicinal herbs was analyzed, and the internal and external factors of aflatoxins contamination in traditional Chinese medicine were also summarized. In view of the high toxicity of aflatoxins, it is proposed to strengthen the mildew prevention and control from the early planting to storage stage, and the reasonable detoxification mode should also be considered. This review aims to provide a reference in guaranteeing the clinical safe administration of medicinal herbs and reducing the risk of being poisoned by aflatoxins.

[Research progresses and formulation development of natural anti-mildew agents based on Chinese herbal medicines].

Owing to the intrinsic factors and some extrinsic environmental conditions, many foods, agricultural products and Chinese materia medicas (CMMs), if not handled properly in the processes of growth, harvesting, processing and storage, can be easily contaminated by all kinds of molds to produce mycotoxins of serious toxicity, which will not only affect the quality, safety and effectiveness of CMMs, but also result in potential threatens to human and animal's health and life. Therefore, in recent decades, it has become the focus on how to prevent and control the foods, agricultural products and CMMs from being moldy and producing toxicity for scientific preservation. Many Chinese herbal medicines (CHMs) especially those with high content of volatile oils with strong antifungal activities have been applied for the scientific preservation of foods, agricultural products and CMMs. Based on these situations, natural anti-mildew agents have been further developed and made into some useful dosage forms, such as tablets, aerosol, liposomes and inclusion, which will not only greatly expand the application scope of CHMs to make the use of anti-mildew agents more convenient, but also achieve the sustained or controlled release of the antifungal effect for scientific preservation of foods, agricultural products and CMMs.

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.

Simultaneous Quantification of Seven Bioactive Flavonoids in Citri Reticulatae Pericarpium by Ultra-Fast Liquid Chromatography Coupled with Tandem Mass Spectrometry.

INTRODUCTION: Citri Reticulatae Pericarpium (CRP) is a commonly-used traditional Chinese medicine with flavonoids as the major bioactive components. Nevertheless, the contents of the flavonoids in CRP of different sources may significantly vary affecting their therapeutic effects. Thus, the setting up of a reliable and comprehensive quality assessment method for flavonoids in CRP is necessary. OBJECTIVE: To set up a rapid and sensitive ultra-fast liquid chromatography coupled with tandem mass spectrometry (UFLC-MS/MS) method for simultaneous quantification of seven bioactive flavonoids in CRP. METHODS: A UFLC-MS/MS method coupled to ultrasound-assisted extraction was developed for simultaneous separation and quantification of seven flavonoids including hesperidin, neohesperidin, naringin, narirutin, tangeretin, nobiletin and sinensetin in 16 batches of CRP samples from different sources in China. RESULTS: The established method showed good linearity for all analytes with correlation coefficient (R) over 0.9980, together with satisfactory accuracy, precision and reproducibility. Furthermore, the recoveries at the three spiked levels were higher than 89.71% with relative standard deviations (RSDs) lower than 5.19%. The results indicated that the contents of seven bioactive flavonoids in CRP varied significantly among different sources. Among the samples under study, hesperidin showed the highest contents in 16 samples ranged from 27.50 to 86.30 mg/g, the contents of hesperidin in CRP-15 and CRP-9 were 27.50 and 86.30 mg/g, respectively, while, the amount of narirutin was too low to be measured in some samples. CONCLUSION: This study revealed that the developed UFLC-MS/MS method was simple, sensitive and reliable for simultaneous quantification of multi-components in CRP with potential perspective for quality control of complex matrices. Copyright © 2016 John Wiley & Sons, Ltd.

[Application and prospect of "couplet medicine" techniques in preservation of Chinese medicinal materials].

Chinese medicinal materials (CMMs) are easily to be contaminated by all kinds of molds to produce various mycotoxins due to their internal factors and the external environmental conditions during the growth, harvesting, processing, and especially storage processes. This will not only affect the quality of CMMs, resulting in enormous financial loss, but also influence the safety and effectiveness of CMMs, posing potential threats to human health. With the increase in awareness of "traditional Chinese medicine health" idea, more and more attention has been paid on how to prevent and control these CMMs from being mouldy to guarantee their safety. Some physical and chemical techniques have been restricted for protecting CMMs due to their own disadvantages. As a green, safe and economic strategy for the preservation of CMMs, "couplet medicine" technique based on the principle of "protecting CMM with another CMM" has been developed: two kinds of CMMs are stored together and fight against each other to prevent mildew metamorphism, exhibiting no obvious changes in color, smell and quality. Nowadays, certain application results have been obtained for the "antagonistic storage" method based on the above mode and principle. In this paper, we would review and discuss the mechanism, practical application and the problems of "couplet medicine" technique, and provide scientific evidences for developing safe and effective tools to protect CMMs from being mouldy.

[Advancement of colloidal gold chromatographic technique in screening of ochratoxin A].

Ochratoxin A (OTA) is a toxic secondary metabolite mainly produced by Aspergillus and Penicillium species, existing in a variety of foodstuffs and Chinese medicines. OTA is difficult to be detected in practice because of the characteristics such as trace amounts, toxicity, existing in complex matrices. In the numerous detection technologies, colloidal gold chromatographic techniques are highly sensitive, specific, cost-effective and user-friendly, and are being used increasingly for OTA screening. Recently, with the development of aptamer technology and its application in chromatographic technique, a newly colloidal gold aptamer chromatographic technique has been developed. This review elaborates the structures and principles of both traditional and newly colloidal gold chromatographic techniques, focuses on newly colloidal gold aptamer chromatographic technique, summarizes and compares their use in rapid detection of OTA. Finally, in order to provide a reference for better research of related work, the development trends of this novel technique are prospected.

[Review of scientific preservation techniques for traditional Chinese medicine becoming mouldy during storage].

All kinds of traditional Chinese medicines (TCMs) differed from each other with different chemical components and properties. Owing to poor conditions and technologies, and the lack of scientific preservation, most TCMs will easily become mouldy during the processes of growth, harvest, handling, translation and especially storage, which will not only influence the quality, safety and efficacy of TCMs, but also cause serious waste and economic losses, and even do great harm to human health. The process of storage is closely related to the qualities of TCMs with many interference factors. Therefore, scientific preservation of TCM during storage is crucial to prevent them from being mouldy. This review analyzed the important harm of mouldy TCMs, summarized the internal and external factors of resulting in TCMs to become mouldy. Based on the above discussion, together with the characteristics and limitations of current TCM preservation techniques, we prospect novel, scientific and reasonable preservation techniques, expecting to provide references for scientific preservation of TCMs to avoid becoming mouldy and guarantee their qualities.

[Contamination level of aflatoxin B1 in lotus seeds rapid screening by indirect competitive ELISA method].

A simple and cost-effective indirect competitive enzyme-linked immune sorbent assay (ic-ELISA) was developed to rapidly screen the content of aflatoxin B1 (AFB1) in lotus seeds, and the results were confirmed by ultra-fast liquid chromatography-tandem mass spectrometry( UFLC-MS/MS). Matrix-matched calibration expressed a good linearity ranging from 0. 171 to 7. 25 µg · L(-1) for AFB, with R2 > 0.978. The medium inhibitory concentration( IC50 ) for AFB1 was 1.29 µg · L(-1), the recovery for AFB1 was 74.73% to 126.9% with RSD < 5%, and the limit of detection (IC10) was 0.128 µg · L(-1). The developed ic-ELSIA method was applied to rapid analysis of AFB, in 20 lotus seeds samples and the results indicated that the contents of AFB, in samples 1-15 were in the range of 1. 19- 115. 3 µg · kg(-1) and in 40% of the samples exceeded the legal limit(5 µg · kg(-1)), while the contamination rate of AFB, in samples 16-20 was 40%. Pearson correlation coefficient(r) reached 0.997 for AFB1 content in the samples detected by ic-ELSIA and UFLC-MS/MS methods. The results proved that the developed ic-ELISA method is simple, sensitive and reliable, and can be used for rapid and high-throughput screening of AFB1 in lotus seeds

[Application of cytometric bead array in rapid high-throughput detection of small molecules].

Cytometric bead array (CBA) is a new analytical technique, which can achieve real-time and rapid detection of targeted components in a small amount of sample. With many advantages of high throughput screening, high specificity and sensitivity, low cost, easy operation and good repeatability, this CBA technique has been widely used for the detection of various components in foods, agricultural products and environmental samples. Recently, it has got significant development in rapid detection of small molecules. This review briefly introduced the theory of CBA technique, summarized the application in the analysis of small molecules, such as mycotoxins, pesticide residues, shellfish toxins, and then prospected the application of trace small molecules detection in the complex matrices of traditional Chinese medicine and the development trend of it.

[Research progress of Chinese herbal medicine raw materials in cosmetics].

Advocating green, nature, environmental protection, safety and the pursuit of efficacy are the trends of cosmetics in the world. In recent years, more and more Chinese herbal extracts with mild, high safety and small irritation are applied to cosmetics as the natural additives. This has become a new hot spot. The recent application advances of Chinese medicine raw materials in cosmetics are overviewed according to their main functions. This review will provide useful references for the future development and application of Chinese medicinal herbs cosmetics.

[Application prospect and expectation of fungistatic agents of plants in preservation of Chinese herbal medicines].

During the process of growth, harvesting, transportation, processing and storage, Chinese herbal medicines (CHMs) can be easily contaminated by fungi and their metabolites like mycotoxins, which not only express negative effects on the quality and safety of CHMs and their processed products, but also pose great threats to human health. Now, some chemical synthetic fungicides have been frequently used to control the growth of fungi and accumulation of mycotoxins in the preservation of CHMs. However, the concentration and type of chemical fungicides allowed for postharvest application are restricted due to the disadvantages of their high residual toxicity, long degradation period and pollution to the environment and so on. Therefore, it is critical to research and develop some highly effective, safe and non-toxic, natural, environment-friendly fungistatic agents from plants to prevent CHMs from being contaminated by fungi and mycotoxins. The paper reviews mycotoxins and their harmfulness, the effective compounds of fungistatic plants as well as the antifungal mechanism to provide scientific evidences for developing novel and effective fungistatic agents plants. Then, the application prospect of fungistatic agents from plants in the preservation of CHMs was discussed.

Overcoming the limitations of directed C-H functionalizations of heterocycles.

In directed C-H activation reactions, any nitrogen or sulphur atoms present in heterocyclic substrates will coordinate strongly with metal catalysts. This coordination, which can lead to catalyst poisoning or C-H functionalization at an undesired position, limits the application of C-H activation reactions in heterocycle-based drug discovery, in which regard they have attracted much interest from pharmaceutical companies. Here we report a robust and synthetically useful method that overcomes the complications associated with performing C-H functionalization reactions on heterocycles. Our approach employs a simple N-methoxy amide group, which serves as both a directing group and an anionic ligand that promotes the in situ generation of the reactive PdX2 (X = ArCONOMe) species from a Pd(0) source using air as the sole oxidant. In this way, the PdX2 species is localized near the target C-H bond, avoiding interference from any nitrogen or sulphur atoms present in the heterocyclic substrates. This reaction overrides the conventional positional selectivity patterns observed with substrates containing strongly coordinating heteroatoms, including nitrogen, sulphur and phosphorus. Thus, this operationally simple aerobic reaction demonstrates that it is possible to bypass a fundamental limitation that has long plagued applications of directed C-H activation in medicinal chemistry.

Development of a sensitive and reliable high performance liquid chromatography method with fluorescence detection for high-throughput analysis of multi-class mycotoxins in Coix seed.

As an edible and medicinal plant, Coix seed is readily contaminated by more than one group of mycotoxins resulting in potential risk to human health. A reliable and sensitive method has been developed to determine seven mycotoxins (aflatoxins B1, B2, G1, G2, zearalenone, α-zearalenol, and ß-zearalenol) simultaneously in 10 batches of Coix seed marketed in China. The method is based on a rapid ultrasound-assisted solid-liquid extraction (USLE) using methanol/water (80/20) followed by immunoaffinity column (IAC) clean-up, on-line photochemical derivatization (PCD), and high performance liquid chromatography coupled with fluorescence detection (HPLC-FLD). Careful optimization of extraction, clean-up, separation and detection conditions was accomplished to increase sample throughput and to attain rapid separation and sensitive detection. Method validation was performed by analyzing samples spiked at three different concentrations for the seven mycotoxins. Recoveries were from 73.5% to 107.3%, with relative standard deviations (RSDs) lower than 7.7%. The intra- and inter-day precisions, expressed as RSDs, were lower than 4% for all studied analytes. Limits of detection and quantification ranged from 0.01 to 50.2 µg kg(-1), and from 0.04 to 125.5 µg kg(-1), respectively, which were below the tolerance levels for mycotoxins set by the European Union. Samples that tested positive were further analyzed by HPLC tandem electrospray ionization mass spectrometry for confirmatory purposes. This is the first application of USLE-IAC-HPLC-PCD-FLD for detecting the occurrence of multi-class mycotoxins in Coix seed.

[Rapid detection of extrinsic harmful contaminants using quantum dots-coated probes and their application prospects in traditional Chinese medicine].

The contamination of extrinsic harmful contaminants including mycotoxins, heavy metals and pesticides, etc, brings serious risks to traditional Chinese medicines (TCMs), further to human health. Due to their unique photoluminescence, chemiluminescence, electrochemical and electrochemiluminescence properties, semiconductor quantum dots (QDs) nanoparticles are widely used to immobilize bioprobes and biosensors, etc. In this review, the luminescence characteristics and specific ligands of QDs probles which are used to determine contaminants were summed up. Then, the applications of QDs-coated novel probes in the determination of mycotoxins, heavy metals and pesticides were discussed in detail. In addition, the contamination levels and characteristics of extrinsic harmful residues in TCMs were investigated. Further, the maximum levels of those contaminants in TCMs were compared with those set by various countries. Finally, the future development trends and problems of QDs-coated probes in the determination of those extrinsic residues in TCMs were prospected.

[Status of pesticide registration and residue analysis for traditional Chinese medicine in China].

The present paper outlined pesticide registration status for traditional Chinese medicines (TCMs) and summarized the characteristics of pesticide contamination in different regions of some widely used TCMs by retrieving last 10 years' literatures. At present, the problems of pesticide residues for TCM include less pesticide registrations, widespread high-residue organochlorine pesticides contamination, pesticide abuse, irregular GAP bases and imperfect pesticide limit standards, etc. According to the current situation, we should adopt some control measures to strengthen the quality control of TCMs so as to ensure the safety of TCMs and related products.

Analysis of zearalenone and α-zearalenol in 100 foods and medicinal plants determined by HPLC-FLD and positive confirmation by LC-MS-MS.

BACKGROUND: Mycotoxins, which may contaminate many foods and medicinal plants, are poisonous to humans. A high-performance liquid chromatography with fluorescence detection (HPLC-FLD) method was successfully developed for analysing the contamination levels of zearalenone (ZON) and its metabolite α-zearalenol (α-ZOL) in 100 widely consumed foods and medicinal plants in China. Samples were extracted with methanol-water (80:20, v/v), and cleaned up by using an immunoaffinity column. RESULTS: The limits of detection of this developed method for ZON and α-ZOL were 4 µg kg(-1) and 2.5 µg kg(-1) , respectively. Recoveries for the samples spiked with three levels (30, 60 and 300 µg kg(-1) for ZON and α-ZOL) ranged from 85.8% to 96.1% with relative standard deviation (RSD) of 2.6-7.1% for ZON, and from 89.9% to 98.7% with RSD of 1.9-9.2% for α-ZOL. Twelve (12%) of these tested samples were contaminated with ZON at levels ranging from 5.3 to 295.8 µg kg(-1). The most contaminated samples were Semen coicis, four of them in a concentration level exceeding 60 µg kg(-1) 'maximum level' (range 68.9-119.6 µg kg(-1)). Positive samples were further confirmed by liquid chromatography-tandem mass spectrometry. CONCLUSION: The results suggest that it is necessary to control ZON contamination in medicinal plants, especially Semen coicis. This is a successful study on the analysis of ZON and α-ZOL in medicinal plants in China by HPLC-FLD. Immunoaffinity clean-up and HPLC-FLD proved to have broad applicability in the field of simultaneously detecting ZON and α-ZOL in foods and medicinal plants and other complicated matrices.

Multi-component analysis of bile acids in natural Calculus bovis and its substitutes by ultrasound-assisted solid-liquid extraction and UPLC-ELSD.

An ultrasound-assisted solid-liquid extraction (USLE) coupled to ultra-performance liquid chromatography-evaporative light scattering detection (UPLC-ELSD) method has been developed for the simultaneous extraction and determination of six bile acids (BAs) in natural Calculus bovis and its substitutes, collected from different origins. The USLE conditions, UPLC chromatographic and ELSD conditions for BAs were optimized. Under optimum conditions, the six target analytes were efficiently extracted and baseline separated within 10 min. The limits of detection (LODs) and quantification (LOQs) for six BAs were less than 7 ng and 22 ng, respectively. Average recoveries were within the range of 98.8-100.7% with relative standard deviations (RSDs) <2% for the six analytes. This method, due to its convenience, high selectivity, fast analysis efficiency and good reproducibility can be employed for analyzing the content differences of six BAs in 40 batches of natural C. bovis and its existing substitutes. The differences of the content of each BA in natural C. bovis and its substitutes were significant, and the total contents of six BAs in 13 batches of natural C. bovis were in the range of 7.96-160.17 mg g(-1), in 20 natural C. bovis of 0-245.89 mg g(-1), in 2 artificial cultivated C. bovis of 178.48-194.22 mg g(-1), in 3 cultured C. bovis of 41.01-107.3 mg g(-1), and in 2 counterfeit C. bovis of 144.9-340.25 mg g(-1). The significant differences of multi-component contents reflected the various inherent qualities of these samples, so, the use of these substitutes as the replacers of natural source in clinic should be paid more attention. Some substitutes could not be used as the replacers.

Effect of berberine on Escherichia coli, Bacillus subtilis, and their mixtures as determined by isothermal microcalorimetry.

The strong toxicity of pathogenic bacteria has resulted in high levels of morbidity and mortality in the general population. Developing effective antibacterial agents with high efficacy and long activity is in great demand. In this study, the microcalorimetric technique based on heat output of bacterial metabolism was applied to evaluate the effect of berberine on Escherichia coli, Bacillus subtilis, individually and in a mixture of both using a multi-channel microcalorimeter. The differences in shape of the power-time fingerprints and thermokinetic parameters of microorganism growth were compared. The results revealed that low concentration (20 µg/mL) of berberine began to inhibit the growth of E. coli and mixed microorganisms, while promoting the growth of B. subtilis; high concentration of berberine (over 100 µg/mL) inhibited B. subtilis. The endurance of E. coli to berberine was obviously lower than B. subtilis, and E. coli could decrease the endurance of B. subtilis to berberine. The sequence of half-inhibitory concentration (IC(50)) of berberine was: B. subtilis (952.37 µg/mL) > mixed microorganisms (682.47 µg/mL) > E. coli (581.69 µg/mL). Berberine might be a good selection of antibacterial agent used in the future. The microcalorimetric method should be strongly suggested in screening novel antibacterial agents for fighting against pathogenic bacteria.

Evaluation of the liver protection and toxicity of Da-Huang-Zhe-Chong pill in rats.

CONTEXT: Da-Huang-Zhe-Chong pill (DHZCP), a classical traditional Chinese formula, consists of 12 crude drugs which have been widely used with significant therapeutic effects. Some drugs in this formula have toxicities that might result in some adverse effects of DHZCP. OBJECTIVE: The liver protection and toxicity of DHZCP were first evaluated against chronic carbon tetrachloride (CCl(4))-induced liver injury in rats. MATERIALS AND METHODS: The rats were treated by intraperitoneal injection of 10% CCl(4) for 12 weeks. At the end of week 4, DHZCP at doses of 44 g/kg (high-dose group) and 22 g/kg (low-dose group) was intragastrically administered to CCl(4)-treated rats, once a day for four weeks. At the end of weeks 8 and 12, the general status of the rats, histopathology of liver, serum alanine aminotransaminase (ALT), aspartate aminotransaminase (AST), alkaline phosphatase (ALP) and total bilirubin (TBIL) levels were observed or determined and recorded. By correspondence analysis (CA) on biochemical markers and liver histopathological score (HS), the "dose-time-response" relationship of DHZCP on the hepatic injury rats was evaluated. RESULTS: The results showed that DHZCP exhibited a significant protective effect on liver injury by reversing the biochemical parameters and histopathological changes. However, this hepatoprotective effect may be weakened, or even be transferred to toxicity with the increase of the administration dose (44 g·kg(-1)·d(-1)) and time (more than 2 months) of this formula. These results were consistent with the histopathological observation and the serum levels. DISCUSSION AND CONCLUSION: Administration of proper dose and time of DHZCP could well play its hepatoprotective effect and even treat hepatitis, but the safety on liver should be considered when large-dose (44 g·kg(-1)·d(-1)) DHZCP is used for long time (more than 2 months). We suggest that the administration dose and time of DHZCP in clinical use should not be increased and prolonged, and simultaneously liver function should be regularly monitored.