Evaluation of Anti-Inflammatory Effects of Celery Leaf and Stem Extracts in LPS-Induced RAW 264.7 Cells Using Nitric Oxide Assay and LC-MS Based Metabolomics.

The present work demonstrated and compared the anti-inflammatory effects of celery leaf (CLE) and stem (CSE) extracts. LC-MS-based metabolomics were an effective approach to achieve the biomarker identification and pathway elucidation associated with the reduction in inflammatory responses. The celery extracts suppressed LPS-induced NO production in RAW 264.7 cells, and CLE was five times more effective than CSE. Distinct differences were revealed between the control and celery-treated samples among the 24 characteristic metabolites that were identified. In celery-treated LPS cells, reversals of intracellular (citrulline, proline, creatine) and extracellular (citrulline, lysine) metabolites revealed that the therapeutic outcomes were closely linked to arginine metabolism. Reversals of metabolites when treated with CLE (aspartate, proline) indicated targeted effects on the TCA and urea cycles, while, in the case of CSE (histidine, glucose), the glycolysis and the pentose phosphate pathways were implicated. Subsequently, apigenin and bergapten in CLE were identified as potential biomarkers mediating the anti-inflammatory response.

Untargeted LC-QTOF-MS/MS based metabolomics approach for revealing bioactive components in probiotic fermented coffee brews.

Amidst trends in non-dairy probiotic foods and functional coffees, we recently developed a fermented coffee brew containing high live counts of the probiotics Lacticaseibacillus rhamnosus GG and Saccharomyces boulardii CNCM-I745. However, probiotic fermentation did not alter levels of principal coffee bioactive components based on targeted analyses. Here, to provide therapeutic justification compared to other non-fermented coffee brews, we aimed to discover postbiotics in coffee brews fermented with L. rhamnosus GG and/or S. boulardii CNCM-I745. By using an untargeted LC-QTOF-MS/MS based metabolomics approach coupled with validated multivariate analyses, 37 differential metabolites between fermentation treatments were putatively annotated. These include the production of postbiotics such as 2-isopropylmalate by S. boulardii CNCM-I745, and aromatic amino acid catabolites (indole-3-lactate, p-hydroxyphenyllactate, 3-phenyllactate), and hydroxydodecanoic acid by L. rhamnosus GG. Overall, LC-QTOF based untargeted metabolomics can be an effective approach to uncover postbiotics, which may substantiate additional potential functionalities of probiotic fermented foods compared to their non-fermented counterparts.

Simultaneous Determination of Quercitrin, Afzelin, Amentoflavone, Hinokiflavone in Rat Plasma by UFLC-MS-MS and Its Application to the Pharmacokinetics of Platycladus orientalis Leaves Extract.

Leaves of Platycladus orientalis have been used as blood cooling and homeostatic therapy for thousands of years in traditional Chinese medicine. Emerging evidences of modern pharmacology have proved flavonoids as the key elements responsible for the efficacies. However, there has been no report on pharmacokinetic study of the flavonoids from Platycladus orientalis leaves extract. In this study, a sensitive and rapid ultra-flow liquid chromatography-tandem mass spectrometry method was established and validated for the simultaneous determination of amentoflavone, afzelin, hinokiflavone and quercitrin in rat plasma. The four flavonoids and luteolin (internal standard, IS) were recovered from rat plasma by methanol-ethyl acetate (v:v, 50:50). Chromatographic separation was performed on a C18 column with gradient elution. Our results showed that the recoveries from spiked control samples were more than 85% for all analytes and IS. The relative standard deviations of intra-day and inter-day precision were within 15% while the REs ranged from -6.6% to 8.0%. The validated method in this study was successfully applied to pharmacokinetic study in healthy rats after oral administration of P. orientalis leaves extract.

Chemical Fingerprint and Quantitative Analysis for the Quality Evaluation of Platycladi cacumen by Ultra-performance Liquid Chromatography Coupled with Hierarchical Cluster Analysis.

Platycladi cacumen (dried twigs and leaves of Platycladus orientalis (L.) Franco) is a frequently utilized Chinese medicinal herb. To evaluate the quality of the phytomedcine, an ultra-performance liquid chromatographic method with diode array detection was established for chemical fingerprinting and quantitative analysis. In this study, 27 batches of P. cacumen from different regions were collected for analysis. A chemical fingerprint with 20 common peaks was obtained using Similarity Evaluation System for Chromatographic Fingerprint of Traditional Chinese Medicine (Version 2004A). Among these 20 components, seven flavonoids (myricitrin, isoquercitrin, quercitrin, afzelin, cupressuflavone, amentoflavone and hinokiflavone) were identified and determined simultaneously. In the method validation, the seven analytes showed good regressions (R ≥ 0.9995) within linear ranges and good recoveries from 96.4% to 103.3%. Furthermore, with the contents of these seven flavonoids, hierarchical clustering analysis was applied to distinguish the 27 batches into five groups. The chemometric results showed that these groups were almost consistent with geographical positions and climatic conditions of the production regions. Integrating fingerprint analysis, simultaneous determination and hierarchical clustering analysis, the established method is rapid, sensitive, accurate and readily applicable, and also provides a significant foundation for quality control of P. cacumen efficiently.

A Review on the Phytochemistry, Pharmacology, Pharmacokinetics and Toxicology of Geniposide, a Natural Product.

Iridoid glycosides are natural products occurring widely in many herbal plants. Geniposide (C17H24O10) is a well-known one, present in nearly 40 species belonging to various families, especially the Rubiaceae. Along with this herbal component, dozens of its natural derivatives have also been isolated and characterized by researchers. Furthermore, a large body of pharmacological evidence has proved the various biological activities of geniposide, such as anti-inflammatory, anti-oxidative, anti-diabetic, neuroprotective, hepatoprotective, cholagogic effects and so on. However, there have been some research articles on its toxicity in recent years. Therefore, this review paper aims to provide the researchers with a comprehensive profile of geniposide on its phytochemistry, pharmacology, pharmacokinetics and toxicology in order to highlight some present issues and future perspectives as well as to help us develop and utilize this iridoid glycoside more efficiently and safely.

A Review on the Phytochemistry, Pharmacology, and Pharmacokinetics of Amentoflavone, a Naturally-Occurring Biflavonoid.

Amentoflavone (C30H18O10) is a well-known biflavonoid occurring in many natural plants. This polyphenolic compound has been discovered to have some important bioactivities, including anti-inflammation, anti-oxidation, anti-diabetes, and anti-senescence effects on many important reactions in the cardiovascular and central nervous system, etc. Over 120 plants have been found to contain this bioactive component, such as Selaginellaceae, Cupressaceae, Euphorbiaceae, Podocarpaceae, and Calophyllaceae plant families. This review paper aims to profile amentoflavone on its plant sources, natural derivatives, pharmacology, and pharmacokinetics, and to highlight some existing issues and perspectives in the future.

(1)H nuclear magnetic resonance-based metabolomics study of earthworm Perionyx excavatus in vermifiltration process.

In this study, (1)H nuclear magnetic resonance (NMR)-based metabolomics approach was used to characterize the metabolic response of the earthworm Perionyx excavatus in continuous vermifiltration for two months under hydraulic loading rates of 1m(3)m(-2)d(-1) (VF1) and 1.5m(3)m(-2)d(-1) (VF1.5). Both VF1 and VF1.5 showed higher removal of chemical oxygen demand and total nitrogen than the biofilter without earthworms. Principal component analysis of the NMR spectra of earthworm metabolites showed significant separations between those not subjected to wastewater filtration (control) and VF1 or VF1.5. Temporal variations of earthworm biomass, and the identified metabolites that are significantly different between control, VF1 and VF1.5 revealed that worms underwent increasing metabolic activity within 20days in VF1 and 14days in VF1.5, then decreasing metabolic activity. The use of NMR-based metabolomics in monitoring earthworm metabolism was demonstrated to be a novel approach in studying engineered vermifiltration systems.

A green and effective approach for characterisation and quality control of chrysanthemum by pressurized hot water extraction in combination with HPLC with UV absorbance detection.

Chrysanthemum is a ubiquitous plant with many species and wide uses, and it is usually consumed as functional food. The main aim of this paper is to demonstrate that chromatographic fingerprints obtained from the HPLC/UV analysis of the pressurized hot water extraction (PHWE) extracts together with the aid of principal component analysis (PCA), allowed for the clustering of various chrysanthemums of different species and provenance. In addition, a parallel study of pressurized fluid extraction (PFE) with methanol was carried out for comparison. From the results, a clearer separation and clustering was obtained with the environmentally-benign water extracts compared with methanol extracts. This study shows that PHWE in combination with HPLC/UV and PCA can be used successfully as a green and effective approach for characterisation and quality control of ubiquitous functional food such as chrysanthemum.

Characterization of oils and fats by 1H NMR and GC/MS fingerprinting: classification, prediction and detection of adulteration.

The correct identification of oils and fats is important to consumers from both commercial and health perspectives. Proton nuclear magnetic resonance ((1)H NMR) spectroscopy, gas chromatography-mass spectrometry (GC/MS) fingerprinting and chemometrics were employed successfully for the quality control of oils and fats. Principal component analysis (PCA) of both techniques showed group clustering of 14 types of oils and fats. Partial least squares discriminant analysis (PLS-DA) and orthogonal projections to latent structures discriminant analysis (OPLS-DA) using GC/MS data had excellent classification sensitivity and specificity compared to models using NMR data. Depending on the availability of the instruments, data from either technique can effectively be applied for the establishment of an oils and fats database to identify unknown samples. Partial least squares (PLS) models were successfully established for the detection of as low as 5% of lard and beef tallow spiked into canola oil, thus illustrating possible applications in Islamic and Jewish countries.

Dengue virus detection using impedance measured across nanoporous alumina membrane.

The prevalence of dengue around the world makes it critical to develop a simple diagnostic device that can be easily handled by end users and provides fast results. In this paper, we described the use of a small and thin piece of alumina membrane, 60 µm thick and 13 mm in diameter as the sensing platform for the detection of dengue infection. The electrochemical setup is simplified by using the membrane as both the working and the counter electrode. This is achieved by coating both sides of the membrane with a submicron layer of platinum. Electrochemical impedance spectroscopy was utilized for the characterization of the immunosensor as well as the acquisition of data. The change in the pore resistance of the membrane displayed a good correlation with the concentration of the dengue 2 and dengue 3 viruses in plaque forming unit (PFU mL⁻¹), giving detection limit of 0.230 PFU mL⁻¹ and 0.710 PFU mL⁻¹ respectively. This thin piece of membrane sensor, coupled with the simple electrochemical setup, fast detection time of 40 min and high sensitivity, showed potential to be developed into a disposable point-of-care diagnostic tool for clinical uses.

Green synthesis of gold nanoparticles using palm oil mill effluent (POME): a low-cost and eco-friendly viable approach.

The present study reports the synthesis of gold nanoparticles (AuNps) from gold precursor using palm oil mill effluent (POME) without adding external surfactant, capping agent or template. The biosynthesized AuNps were characterized by using UV-vis spectroscopy, transmission electron microscopy (TEM), X-ray diffraction (XRD), and Fourier transform infrared spectroscopy (FTIR). According to the image analysis performed on a representative TEM micrograph by counting 258 particles, the obtained AuNps are predominantly spherical with an average size of 18.75 ± 5.96 nm. In addition, some triangular and hexagonal nanoparticles were also observed. The influence of various reaction parameters such as reaction pH, concentration of gold precursor and interaction time to the morphology and size of biosynthesized AuNps was investigated. This study shows the feasibility of using agro waste material for the biosynthesis of AuNps which is potentially more scalable and economic due to its lower cost.

Determination of pyrrolizidine alkaloids in comfrey by liquid chromatography-electrospray ionization mass spectrometry.

Symphytum officinale L. (comfrey) is a medicinal plant commonly used in decoctions and aliments. Besides therapeutic bioactive compounds present in the herb, it is found to contain hepatotoxic pyrrolizidine alkaloids (PAs), such as lycopsamine and others. In the present study, PAs such as lycopsamine, echimidine and lasiocarpine were determined using electrospray liquid chromatography-mass spectrometry (LC-MS) with the method precision (relative standard deviation, RSD) <10%. Detection of lycopsamine, symviridine and their N-oxides could be confirmed with a newly developed method based on HPLC ion-trap and orbitrap MS with electrospray ionization interface. With LC-MS, quantitative analysis of lycopsamine in the botanical extract was carried out. The effect of extraction solvent was optimized by sonication and methanol: H(2)O (50:50) was selected. Then a rapid method based on pressurized hot water extraction (PHWE) was employed for the extraction of lycopsamine from comfrey followed by the comparison with heating under reflux with the RSD ranging from 2.49% to 19.32%. Our results showed a higher extraction efficiency for heating under reflux compared with PHWE. It was proposed that the lower extraction efficiency for PHWE was attributable to dissolved nitrogen from air which caused the reduction in the solubility of lycopsamine in the compressed hot solvent. In this study, quantitative analysis of PAs in comfrey was demonstrated. In addition, it was found that the use of subcritical water for extractions depended on the physical properties of the dissolved solutes and their tendency to degrade under the chosen extraction conditions.

Determination of senkirkine and senecionine in Tussilago farfara using microwave-assisted extraction and pressurized hot water extraction with liquid chromatography tandem mass spectrometry.

Tussilago farfara (Kuan Donghua) is an important Chinese herbal medicine which has been shown to contain many bioactive compounds and widely used to relieve cough and resolve phlegm. However, besides therapeutic bioactive compounds, this herb has been found to contain toxic pyrrolizidine alkaloids (PAs), mainly senkirkine and traces of senecionine. In this report, conditions for microwave-assisted extraction (MAE) and pressurized hot water extraction (PHWE) were optimized for the extraction of the PAs. The results were compared against heating under reflux. It was found that the binary mixture of MeOH:H(2)O (1:1) acidified using HCl to pH 2-3 was the optimal solvent for the extraction of the PAs in the plant materials. Liquid chromatography (LC) with ultra-violet (UV) detection and electrospray ionization mass spectrometry (ESI-MS) in the positive mode was used for the determination and quantitation of senkirkine and senecionine in the botanical extract. The proposed extraction methods with LC/MS allow for the rapid detection of the major and the minor alkaloids in T. farfara in the presence of co-eluting peaks. With LC/MS, the quantitative analysis of PAs in the extract was done using internal standard calibration and the precision was found to vary from 0.6% to 5.4% on different days. The limits of detection (LODs) and limits of quantitation (LOQs) for MAE and PHWE were found to vary from 0.26 microg/g to 1.04 micro/g and 1.32 micro/g to 5.29 microg/g, respectively. The method precision of MAE and PHWE were found to vary from 3.7% to 10.4% on different days. The results showed that major and minor alkaloids extracted using MAE and PHWE were comparable to that by heating under reflux. Our data also showed that significant ion suppression was not observed in the analysis of senkirkine and senecionine in the botanical extracts with co-eluting peaks.

Metabonomics investigation of human urine after ingestion of green tea with gas chromatography/mass spectrometry, liquid chromatography/mass spectrometry and (1)H NMR spectroscopy.

A method using gas chromatography/mass spectrometry (GC/MS), liquid chromatography/mass spectrometry (LC/MS) and (1)H NMR with pattern recognition tools such as principle components analysis (PCA) was used to study the human urinary metabolic profiles after the intake of green tea. From the normalized peak areas obtained from GC/MS and LC/MS and peak heights from (1)H NMR, statistical analyses were used in the identification of potential biomarkers. Metabolic profiling by GC/MS provided a different set of quantitative signatures of metabolites that can be used to characterize the molecular changes in human urine samples. A comparison of normalized metabonomics data for selected metabolites in human urine samples in the presence of potential overlapping peaks after tea ingestion from LC/MS and (1)H NMR showed the reliability of the current approach and method of normalization. The close agreements of LC/MS with (1)H NMR data showed that the effects of ion suppression in LC/MS for early eluting metabolites were not significant. Concurrently, the specificity of detecting the stated metabolites by (1)H NMR and LC/MS was demonstrated. Our data showed that a number of metabolites involved in glucose metabolism, citric acid cycle and amino acid metabolism were affected immediately after the intake of green tea. The proposed approach provided a more comprehensive picture of the metabolic changes after intake of green tea in human urine. The multiple analytical approach together with pattern recognition tools is a useful platform to study metabolic profiles after ingestion of botanicals and medicinal plants.

Capillary electrophoresis with capacitively coupled contactless conductivity detection for low molecular weight organic acids in different samples.

CE with capacitively coupled contactless conductivity detection (CE-C(4)D) was explored and validated for the identification and quantification of organic acids in various types of samples. The analyses were performed under optimized conditions, using a buffer system composed of 20 mM MES-histidine (His), pH 6.0, 0.1 mM CTAB, 0.025% HP-beta-CD, and 10% methanol. The investigation included a study of the effects of buffer pH, concentration of CTAB, type and concentration of organic additives, on the migration behavior, resolution and selectivity of the organic acids. The intra- and interday RSDs (n = 6) obtained for migration time and peak area were typically in the range of 0.12-2% and 0.5-4%, respectively. Linearity, detection limits, and repeatability were evaluated. In order to evaluate the application potential of the developed method, real samples from different sources were analyzed. The results demonstrate that CE-C(4)D is a versatile tool for analyzing organic acids in beverages, Chinese herbal medicines (CHM) and plants as it allows for their detection, identification, and quantification.

Dynamic pH junction-sweeping capillary electrophoresis for online preconcentration of toxic pyrrolizidine alkaloids in Chinese herbal medicine.

There is a need to develop simple yet effective preconcentration methods to enhance concentration sensitivity for CE analysis of trace level analytes in real samples, particularly when commonly available but less sensitive detection methods, e.g., UV detection, are used. In this report, a hyphenated online preconcentration strategy combining dynamic pH junction with sweeping (i.e., dynamic pH junction-sweeping) was employed for the analysis of four toxic pyrrolizidine alkaloids (PAs) of senkirkine, senecionine, retrorsine, and seneciphylline in Chinese herbal medicine (Kuan donghua). Direct electrokinetically focusing of a large sample volume injection (up to 20% of capillary length) on the capillary was performed using the dynamic pH junction-sweeping method. A sample matrix consisting of 10 mM phosphate with 20% methanol at pH 4.0 and a BGE containing 20 mM borate, 30 mM SDS, and 20% methanol at pH 9.1 were utilized to realize dynamic pH junction-sweeping for PAs. This online preconcentration strategy resulted in sensitivity enhancement factors ranging from 23.8- to 90.0-fold for the four toxic PAs, giving an LOD as low as 30 ppb for the PAs. Critical factors such as sample matrix type, pH, and salt concentration were also examined to achieve higher sensitivity enhancement, shorter analysis time, and better resolution. The results indicate that the proposed dynamic pH junction-sweeping technique is a powerful alternative approach for identification and determination of trace levels of these toxic PAs and other hydrophobic, protonatable compounds in real samples.

Separation and determination of toxic pyrrolizidine alkaloids in traditional Chinese herbal medicines by micellar electrokinetic chromatography with organic modifier.

A simple and rapid micellar electrokinetic chromatography (MEKC) method was developed for the separation and determination of four toxic pyrrolizidine alkaloids (PAs) (senkirkine, senecionine, retrorsine, and seneciphylline) in two traditional Chinese herbal medicines (Qian liguang and Kuan donghua). Separation was performed in the running buffer consisting of 20 mM borate, 30 mM SDS, and 20% methanol at pH 9.1. With the optimized separation conditions, four PAs were separated in 17 min by a single run. The calibration curves showed good linearity with correlation efficiencies (R(2)) between 0.9940 and 0.9988. RSDs in migration time and peak area were 0.31, 0.40, 0.39, 0.48% and 3.28, 3.48, 4.16, 3.42% for senkirkine, senecionine, retrorsine, and seneciphylline, respectively. Limits of detection (S/N = 3) varied from 1.19 to 2.70 microg/mL. The proposed method was applied to determine the PAs extracted from Chinese herbal medicines (Qian liguang and Kuan donghua). PA of senkirkine in Kuan donghua was detected and the amount was found to be 79.1 microg/g. The results obtained indicate that the proposed MEKC method could potentially become an effective alternative tool for qualification control and quantitative analysis of herbal medicines in pharmaceutical industry.

Analysis of Chinese medicine preparations by capillary electrophoresis-mass spectrometry.

In Chinese medicines, herbs are usually prepared before use by patients. Since the preparation procedures convert the original component into one or more products, study of the procedures is usually complex and involves several compounds. On-line coupling of capillary electrophoresis (CE) to mass spectrometry (MS) allows both the efficient separation of CE and the specific and sensitive detection of MS to be achieved. In this study, CE-MS was applied to the determination of alkaloids in Maqianzi (the seed of Strychnos pierrian) and Wutou (aconite root, Radix aconiti praeparata) during the preparation procedure. With optimal CE-MS conditions, alkaloids in both prepared and unprepared Maqianzi were determined successfully in the total ion current (TIC) mode. However, single ion monitoring (SIM) had to be applied for the separation of aconitum alkaloids and their hydrolysis products. Quantification data indicated that MS detection under SIM mode is more sensitive than UV detection. Based on the CE-MS method developed, the hydrolysis of aconitum alkaloids in water and methanol was also studied.