Experimental design approach for the quantitative analysis of multicomponents by single marker and HPLC fingerprinting of Thunbergia laurifolia aqueous extract.

INTRODUCTION: Thunbergia laurifolia is used in traditional Thai medicine to reduce fever and treat mouth ulcers. However, the quantitative analysis of chemical markers has not yet been officially defined. OBJECTIVE: The objective of this study is to develop a high-performance liquid chromatography (HPLC) method using a design of experiment (DoE) for the quantitative analysis of multicomponents by single marker (QAMS) and fingerprinting of the T. laurifolia aqueous extract. MATERIALS AND METHODS: Critical variables were screened using a two-level fractional factorial design, followed by the optimization of the selected variables using a central composite design. The validated method was applied for quality assessment based on QAMS and fingerprinting of the extract. RESULTS: Optimum conditions of DoE for the analysis of caffeic acid, vicenin-2, and rosmarinic acid were determined. The relative correction factors for caffeic acid and vicenin-2 were calculated using rosmarinic acid as an internal reference standard, and their contents in 30 samples were determined. The differences between the external standard method (ESM) and QAMS were compared. No significant difference was observed in the quantitative determination, proving the consistency QAMS and ESM. HPLC fingerprints of T. laurifolia were established with 8 of 12 characteristic peaks that were structurally characterized using HPLC-diode array detection-electrospray ionization/tandem mass spectrometry. The similarity of the fingerprints in all samples was ≥0.74, and the pattern recognition of the characteristic peaks was satisfied. CONCLUSION: The proposed method efficiently detected multiple components of the T. laurifolia extract. Thus, the method is beneficial in providing references for enhancing the quality control of other herbal medicines.

Determination of Phytochemical Contents in Extracts from Different Growth Stages of Oroxylum indicum Fruits Using HPLC-DAD and QAMS Methods.

Flavones are major compounds found in several parts of Oroxylum indicum (O. indicum). The quantification of multiple components by one marker (QAMS) method and the high-performance liquid chromatography (HPLC) method were developed for the quantitative analysis of extracts from the young fruits, green mature fruits, dry pod coats and seeds of O. indicum. Oroxin A, oroxin B and chrysin-7-O-glucuronide were identified in the O. indicum extracts. Oroxylin A and 5-hydroxymethylfurfural were isolated and structurally identified from the pod coat and young fruit extracts, respectively. From the HPLC analysis of the seven major flavones in the extracts, baicalin was the major compound in all extracts investigated (0.4-11% w/w of the extract). All flavone contents were low in the young fruit extract (<1% w/w of the extract). The green mature fruit and dry pod coat extracts showed similar constituent compositions. They contained small amounts of baicalin and oroxylin A, which were found only in these two extracts. Oroxylin A could be used as a marker to indicate the maturity of O. indicum fruits, while 5-hydroxymethylfurfural could be used as a marker for the young fruits. Baicalin was found to be a suitable single marker to calculate the contents of all flavones in the O. indicum extracts.

Design of Turmeric Rhizome Extract Nano-Formula for Delivery to Cancer Cells.

Novel turmeric rhizome extract nanoparticles (TE-NPs) were developed from fractions of dried turmeric (Curcuma longa Linn.) rhizome. Phytochemical studies, by using HPLC and TLC, of the fractions obtained from ethanol extraction and solvent-solvent extraction showed that turmeric rhizome ethanol extract (EV) and chloroform fraction (CF) were composed mainly of three curcuminoids and turmeric oil. Hexane fraction (HE) was composed mainly of turmeric oil while ethyl acetate fraction (EA) was composed mainly of three curcuminoids. The optimal TE-NPs formulation with particle size of 159.6 ± 1.7 nm and curcumin content of 357.48 ± 8.39 µM was successfully developed from 47-run D-optimal mixture-process variables experimental design. Three regression models of z-average, d50, and d90 could be developed with a reasonable accuracy of prediction (predicted r2 values were in the range of 0.9120-0.9992). An in vitro cytotoxicity study using MTT assay demonstrated that the optimal TE-NPs remarkably exhibited the higher cytotoxic effect on human hepatoma cells, HepG2, when compared with free curcumin. This study is the first to report nanoparticles prepared from turmeric rhizome extract and their cytotoxic activity to hepatic cancer cells compared with pure curcumin. These nanoparticles might serve as a potential delivery system for cancer therapy.

Flavone-Rich Fractions and Extracts from Oroxylum indicum and Their Antibacterial Activities against Clinically Isolated Zoonotic Bacteria and Free Radical Scavenging Effects.

Oroxylum indicum extracts from the seeds collected from Lampang and Pattani provinces in Thailand, and young fruits and flowers exhibited in vitro display antioxidant and antibacterial activities against clinically isolated zoonotic bacteria including Staphylococcus intermedius, Streptococcus suis, Pseudomonas aeruginosa, ß-hemolytic Escherichia coli and Staphylococcus aureus. The orange crystals and yellow precipitates were obtained from the preparation processes of the seed extracts. The orange-red crystals from the seeds collected from Lampang province exhibited strong in vitro 2,2-diphenyl-1-picrylhydrazyl (DPPH) scavenging effects (EC50 value = 25.99 ± 3.30 µg/mL) and antibacterial effects on S. intermedius and ß-hemolytic E. coli while the yellow precipitate from the same source exhibited only antioxidant activity. Quantitative analysis of phytochemicals in O. indicum samples by spectrophotometric and HPLC techniques showed that they contained different amounts of total phenolic, total flavonoid and three major flavones; baicalin, baicalein and chrysin contents. Young fruit extract, which contained low amounts of flavone contents, still promoted antibacterial effects against the tested bacteria with IC50 values lower than 1 mg/mL and MIC values between 4 to 10 mg/mL in S. intermedius, S. aureus and S suis while higher IC50 and MIC values against P. aeruginosa and ß-hemolytic E. coli were found. From scanning electron microscopy, the extract of the young fruit of O. indicum promoted morphological changes in the bacterial cells by disrupting the bacterial cell walls, inducing leakage of the cellular content, and generating the abnormal accumulation of cells. The mechanism of action of the extract for this antibacterial effect may be the disruption of the cell membrane and abnormal cell aggregations. Regression analysis of the results suggests the correlation between total phenolic and total flavonoid contents and antioxidant and antibacterial effects. Baicalin was found to have a high correlation with an inhibitory effect against ß-hemolytic E. coli while three unidentified peaks, which could be flavones, showed high correlations with an inhibitory effect against S. intermedius, S. suis, P. aeruginosa and S. aureus.

Flavones Contents in Extracts from Oroxylum indicum Seeds and Plant Tissue Cultures.

Oroxylum indicum (L.) Benth. ex Kurz or Pheka, is a plant in the Bignoniaceae family with various traditional uses. The mature fruits promote anti-helminthic and stomachic effects, while the seeds have been used as a purgative and for the relief of tonsil pain. The young fruits are popularly consumed as vegetables, while the seeds are one of the components in traditional drink formulations. To develop new plant raw material sources, a plant tissue culture technique was used to generate plant tissue cultured samples from the seeds of O. indicum. Plant tissue cultured samples were collected from three different growth stages; 4 days, then at 3 and 9 weeks, and prepared as crude extracts by maceration with ethanol, along with the seed raw material sample. A high performance liquid chromatographic (HPLC) method was used for quantitative analysis of the contents of the three major flavones; baicalin, baicalein, and chrysin in the extracts from the seeds and plant tissue cultured samples of this plant. Baicalin was found in the highest amount among these three flavones in all extracts. The seed extract contained the highest baicalin content (24.24% w/w in the extract), followed by the shoot extract from tissue-cultured plant at week 3 (14.78% w/w of the extract). The amounts of chrysin in all O. indicum showed the same trend as the contents of baicalin, but the amounts were lower, while baicalein was accumulated at the lowest amount among three flavonoids and the amounts were quite stable in all O. indicum extracts. From the results, O. indicum seed and plant tissue cultured extracts have potential as sources of flavones, which could be further developed as health products in the future.

Chemical Standardization and Anti-Proliferative Activity of Ardisia elliptica Fruit against the HCT116 Human Colon Cancer Cell Line.

The present study is intended to carry out the chemical standardization and evaluation of the anti-proliferative activity of A. elliptica fruit extract. A. elliptica fruit powder was extracted with ethanol. The obtained extract was assessed for total phenolic content using the Folin-Ciocalteu method. Moreover, a simple, accurate, and precise reversed phase high-performance liquid chromatographic method was developed and validated to determine the embelin content of A. elliptica fruit extract. Then, the extract and embelin were investigated for their anti-proliferative effect against HCT-116 cells. Finally, the mechanisms of inhibition of the extract and embelin on the mRNA expression of pro-apoptotic genes Bad, Bax, and Caspase-8 and anti-apoptotic genes c-IAP1, Mcl-1, and XIAP were determined by real-time qRT-PCR. The phenolic content and embelin content of the extract were 5.20 ± 0.01 g of gallic acid equivalent per 100 g of dried fruit (g% GAE) and 5.57 ± 0.56 mg/g of extract, respectively. The extract and embelin showed strong anti-proliferative effects on HCT-116 cells with 50% inhibition concentration (IC50) values of 19.16 ± 1.09 µg/mL and 25.93 ± 1.75 µg/mL, respectively. The A. elliptica extract exhibited a significant increase in the mRNA level of Bad, Bax, and Caspase-8 and a significant decrease in c-IAP1, Mcl-1, and XIAP. Embelin showed a significant decrease in Mcl-1 and XIAP.

Inhibitory Effects on Clinical Isolated Bacteria and Simultaneous HPLC Quantitative Analysis of Flavone Contents in Extracts from Oroxylum indicum.

Oroxylum indicum is a medicinal plant in Thailand, which has been used as a tonic and for the treatment of various diseases. Extracts from various parts of O. indicum were reported as promoting in vitro antioxidant and antibacterial effects. Phytochemical analysis suggested that this plant contained some flavones. O. indicum fruit and seed water and ethanol extracts and their major flavonoids including baicalein, baicalin, and chrysin were tested for in vitro antibacterial activities on four clinical isolated bacteria, namely, Staphylococcus intermedius, Streptococcus suis, Pseudomonas aeruginosa, and ß-Escherichia coli, using a broth micro-dilution assay. The amounts of these three major flavonoids were also quantitatively analyzed using the high-performance liquid chromatographic (HPLC) method. O. indicum fruit ethanol extract from Nakhon Pathom province (OFNE) promoted the strongest antimicrobial activity against four clinical pathogenic bacteria, including S. intermedius (IC50 = 1.30 mg/mL), S. suis (13.59% inhibition at 7.81 mg/mL), P. aeruginosa (IC50 = 39.20 mg/mL), and ß-E. coli (IC50 = 66.85 mg/mL). Baicalin showed high in vitro antibacterial effect to all tested bacteria. From the optimized and validated HPLC method, baicalin, baicalein, and chrysin contents in O. indicum extracts were 0.19 ± 0.00 - 9.45 ± 0.13, 0.14 ± 0.00 - 1.27 ± 0.02, and 0.02 ± 0.00 - 0.96 ± 0.02 g/100 g extract, respectively. Baicalin was found to be the major compound in O. indicum seed extract followed by baicalein, whereas chrysin was found in lower amounts than the amounts of the other two flavonoids in all O. indicum extracts.

Screening for Antibacterial and Antioxidant Activities and Phytochemical Analysis of Oroxylum indicum Fruit Extracts.

Oroxylum indicum, which is called Pheka in Thai, is a traditional Thai plant in the Bignoniaceae family with various ethnomedical uses such as as an astringent, an anti-inflammatory agent, an anti-bronchitic agent, an anti-helminthic agent and an anti-microbial agent. The young fruits of this plant have also been consumed as vegetables. However, there has been no report concerning its antibacterial activities, especially activities related to clinically isolated pathogenic bacteria and the in vitro antioxidant effects of this plant. Therefore, the extracts from O. indicum fruits and seeds collected from different provinces in Thailand were prepared by decoction and maceration with ethanol and determined for their in vitro antibacterial effects on two clinically isolated bacteria, Streptococcus suis and Staphylococcus intermedius, using disc diffusion assay. Ethanol extracts from O. indicum fruits collected from Nakorn Pathom province at the concentration of 1000 mg/mL exhibited intermediate antibacterial activity against S. intermedius with an inhibition zone of 15.11 mm. Moreover, it promoted moderate inhibitory effects on S. suis with an inhibition zone of 14.39 mm. The extracts prepared by maceration with ethanol promoted higher antibacterial activities than those prepared with water. The ethanol extract from the seeds of this plant, purchased in Bangkok, showed stronger in vitro antioxidant activities than the other extracts, with an EC50 value of 26.33 µg/mL. Phytochemical analysis suggested that the seed ethanol extract contained the highest total phenolic and flavonoid contents (10.66 g% gallic acid equivalent and 7.16 g% quercetin equivalent, respectively) by a significant amount. Thin layer chromatographic analysis of the extracts showed the chromatographic band that could correspond to a flavonoid baicalein. From the results, extracts from O. indicum fruits have an in vitro antioxidant effect, with antibacterial potential, on clinically pathologic bacteria and they contain an antioxidant flavonoid which could be developed for medicinal and pharmaceutical purposes in the future.

Thunbergia laurifolia extract ameliorates cognitive and emotional deficits in olfactorectomized mice.

CONTEXT: Thunbergia laurifolia Lindl. (Acanthaceae) is a Thai medicinal plant used for the detoxification of poison which is likely to be beneficial for the treatment of cognitive deficits including Alzheimer's disease. OBJECTIVE: To elucidate the effects of Thunbergia laurifolia leaf extract (TLL) on cognitive dysfunction and depression-like behavior in olfactory bulbectomized mice (OBX). MATERIALS AND METHODS: OBX mice were treated daily with TLL at the dose of 250 and 500 mg/kg, tacrine, and imipramine, on the day after 10 d of OBX operation. The effects of TLL on cognitive and depression-like behavior of the animals were analyzed. After completing behavioral experiments, the expression levels of cholinergic marker genes encoding ChAT and muscarinic M1 receptor were quantitatively analyzed. RESULTS: TLL and tacrine reduced OBX-induced cognitive deficits in the object recognition test (ORT) with the time spent for the novel object two times longer than that of the familiar object. Moreover, TLL at the dose of 500 mg/kg and imipramine ameliorated depression-like behavior in the tail suspension test (TST) by reducing the duration of immobility from 25.18% to 3.16% and from 25.18% to 6.48%, respectively. TLL at the dose of 250 and 500 mg/kg reversed the OBX-induced down-regulation of ChAT mRNA expression in the hippocampus from 0.12 to 0.17 and 0.24, respectively, while the down-regulation of mRNA expression of muscarinic M1 receptor was also reversed by TLL from 0.23 to 0.38 and 0.48, respectively. CONCLUSIONS: TLL ameliorates non-spatial short-term memory deficits in OBX mice, and has the potential to exhibit an antidepressant-like action.

Green extraction and partial purification of roselle (Hibiscus sabdariffa L.) extracts with high amounts of phytochemicals and in vitro antioxidant and antibacterial effects.

The aim of this study is to develop roselle (Hibiscus sabdariffa L.) extracts with high amounts of phytochemicals and in vitro antioxidant and antibacterial effects using green extraction technique. Roselle extract prepared by ultrasonic extraction with 80% ethanol and 1% hydrochloric acid (RSUEH) promoted the highest yield and high total phenolic (5.21 ± 0.05 g% gallic acid equivalent of dried extract), total flavonoid (1.29 ± 0.06 g% quercetin equivalent of dried extract), and total anthocyanin contents (0.21 ± 0.00 and 0.44 ± 0.00 g% cyanidin 3-glucoside equivalent of dried extract determined by pH-differential method with standard calculation and pH-differential method with standard curve of cyanidin 3-glucoside, respectively). From the validated high-performance liquid chromatography analysis, delphinidin-3-sambubioside, delphinidin-3-glucoside, and cyanidin-3-glucoside were found to be the major anthocyanins in roselle extract. This extract exhibited moderate 2,2-diphenyl-1-picrylhydrazyl (DPPH) scavenging effect (EC50 289.61 ± 0.16 µg/mL) and strong inhibitory effects against Escherichia coli, Staphylococcus aureus, Staphylococcus intermedius, Proteus mirabilis, and Pseudomonas aeruginosa with the inhibitory concentration (IC50) values in the range of 0.15 ± 0.05 to 4.64 ± 0.07 mg/mL and the minimum inhibitory concentration (MIC) values ranged from 1.66 ± 0.06 to 7.11 ± 1.05 mg/mL. From partial purification, fraction 28 (F28) contained higher amounts of all analyzed phytochemicals than roselle extracts and promoted stronger DPPH scavenging effects (EC50 54.88 ± 2.23 µg/mL). F28 showed antibacterial activity with IC50 values against all tested bacteria in the range of 0.19 ± 0.07 to 4.69 ± 0.09 mg/mL, and the MIC values ranged from 2.46 ± 0.08 to 6.24 ± 0.09 mg/mL.

Inhibitory effects against zoonotic bacteria by Oroxylum indicum ointment and effects to dog wound.

A topical antimicrobial ointment was developed from the fruit extract of Oroxylum indicum and was evaluated for its antibacterial and wound-healing effects and acute toxicity in animal models. O. indicum fruit and seed extracts exhibited antibacterial activities against clinically isolated bacteria and showed in vitro antioxidant activities. To develop a topical antimicrobial ointment from the fruit extract of O. indicum and evaluate for its antibacterial and wound-healing effects. O. indicum fruit extract ointment was prepared and qualitatively controlled. Acute toxicity of the extract was evaluated in the animal model. Antibacterial effects and healing effects of the ointment to the dog wound were investigated. The results are expressed as mean ± standard deviation. The ointment exhibited in vitro antibacterial effects. A single daily application of the ointment to a dog's wound exhibited a wound-healing effect with complete epithelialization within 7 days while the wound was completely healed with the removal of the scabs, the size was decreased to 14% of the original size within 12 days. The ointment was found no acute toxicity in the animal model. O. indicum ointment promoted in vitro antibacterial activity and wound-healing effect in dogs with no acute toxicity.

Delivery of Avocado Seed Extract Using Novel Charge-Switchable Mesoporous Silica Nanoparticles with Galactose Surface Modified to Target Sorafenib-Resistant Hepatocellular Carcinoma.

Background: Sorafenib-resistant (SR) hepatocellular carcinoma (HCC) is a current serious problem in liver cancer treatment. Numerous phytochemicals derived from plants exhibit anticancer activity but have never been tested against drug-resistant cells. Methods: Avocado seed extract (APE) isolated by maceration was analysed for its phytochemical composition and anticancer activity. Novel design charge-switchable pH-responsive nanocarriers of aminated mesoporous silica nanoparticles with conjugated galactose (GMSN) were synthesised for delivering APE and their physicochemical properties were characterized. The drug loading efficiency (%LE) and entrapment efficiency (%EE) were evaluated. Anticancer activity of APE loaded GMSN was measured against HCC (HepG2, Huh-7) and SR-HCC (SR-HepG2). Results: Anticancer activity of APE against non-resistant HepG2 (IC50 50.9 ± 0.83 µg mL-1), Huh-7 (IC50 42.41 ± 1.88 µg mL-1), and SR-HepG2 (IC50 62.58 ± 2.29 µg mL-1) cells was confirmed. The APE loaded GMSN had a diameter of 131.41 ± 14.41 nm with 41.08 ± 2.09%LE and 44.96 ± 2.26%EE. Galactose functionalization (55%) did not perturb the original mesoporous structure. The GMSN imparted positive surface charges, 10.3 ± 0.61mV at acidic medium pH 5.5 along with rapid release of APE 45% in 2 h. The GMSN boosted cellular uptake by HepG2 and SR-HepG2 cells, whereas the amine functionalized facilitated their endosomal escape. Their anticancer activity was demonstrated in non-resistant HCC and SR-HCC cells with IC50 values at 30.73 ± 3.14 (HepG2), 21.86 ± 0.83 (Huh-7), 35.64 ± 1.34 (SR-HepG2) µg mL-1, respectively, in comparison to the control and non-encapsulated APE. Conclusion: APE loaded GMSN is highly effective against both non-resistant HCC and SR-HCC and warrants further in vivo investigation.

Antioxidant activity of Acanthopanax trifoliatus.

OBJECTIVE: The aim of this study was to investigate the antioxidant activity of extracts of various parts of Acanthopanax trifoliatus obtained by different extraction methods. The total phenolic and flavonoid contents of the extracts were also determined. MATERIALS AND METHODS: The leaves, stems, stem bark, roots and root bark of A. trifoliatus were extracted separately using decoction, maceration and refluxing methods. The extracts were analysed for free-radical-scavenging activity using the 1,1-diphenyl-2-picryl-hydrazyl scavenging assay and the thiobarbituric-acid-reactive substance method for the inhibition of lipid peroxidation of a rat brain homogenate. Total phenolic and total flavonoid contents of the extracts were measured by UV spectrophotometry. RESULTS: The leaf decoction extracts possessed a significantly stronger antioxidant activity as revealed by both methods. Total phenolic and flavonoid contents of the extracts ranged from 2.16 to 21.79 g% chlorogenic acid equivalent and from 0.37 to 9.61 g% rutin equivalent, respectively. Analysis of the leaf decoction extract, which exhibited the most potent antioxidant activity, by thin-layer chromatography revealed flavonoid and polyphenolic compounds corresponding to rutin and chlorogenic acid. CONCLUSION: The leaf aqueous extracts showed a high level of antioxidative activity and contained high levels of both phenolic and flavonoid compounds. The magnitude of antioxidant activity corresponded with the level of phenolic and flavonoid compounds.

Clinical efficacy of 0.5% topical mangosteen extract in nanoparticle loaded gel in treatment of mild-to-moderate acne vulgaris: A 12-week, split-face, double-blinded, randomized, controlled trial.

BACKGROUND: Acne vulgaris is the most common inflammatory sebaceous gland disorder in young adults. The resistant strains of Propionibacterium acnes (P. acnes) are of increasing concern in the treatment of acne. OBJECTIVES: To evaluate the efficacy of 0.5% topical mangosteen extract in nanoparticle loaded gel (containing alpha-mangostin) compared with 1% clindamycin gel for treatment of mild-to-moderate acne vulgaris. METHODS: Patients aged 18-40 years were enrolled in this double-blinded, split-face, randomized, control study. The 2.5% benzoyl peroxide cream was applied to both sides of the faces once daily for 5 minutes and washed off. Each patient was randomly treated with the mangosteen fruit rind extract on one side and 1% clindamycin on another side of the face twice daily for 12 weeks. Treatment efficacies and side effects were evaluated on every follow-up. RESULTS: Twenty-eight patients, 24 female (85.7%), mean ± SD age of 25.14 ± 5.8, with Global Acne Grading system (GAGs) score of 15.43 ± 5.96 were included. Mangosteen fruit rind extract significantly showed significant 66.86% and 67.05% reduction of comedone and inflammatory lesions (P < 0.001) after 12-week treatment. The improvement on both treated sides significantly showed since 2 weeks after treatment, without statistical difference between two groups. Nonetheless, the mangosteen fruit rind extract revealed significantly better improvement of clinical severity, with no severe side effects. CONCLUSIONS: The mangosteen fruit rind extract formation could be a phytopharmaceutical medication for effective treatment of mild and moderate acne vulgaris treatment comparable to 1% clindamycin gel, with no severe side effects.

Simultaneous HPTLC Determination of Rhein and Aloe-emodin in Senna alata Leaves from Thailand and their Commercial Products.

A high performance thin layer chromatographic (HPTLC) method was developed and validated for simultaneous determination of rhein and aloe-emodin, major anthraquinone constituents, in S. alata leaves. The separation was performed on a silica gel 60 F254 HPTLC plate using ethyl acetate: methanol: water (100:17:10, v/v/v) as the development system. Validation of the analytical method for rhein and aloe-emodin promoted acceptable parameters. Good linearity in the range of 40-480 ng/band was obtained while intra-day and inter-day precisions were shown to be precise with relative standard deviations of less than 5%. The average percentage recoveries of rhein and aloe-emodin were 98.8% and 98.9%, suggesting acceptable accuracy. The content of rhein and aloe-emodin in S. alata leaves collected from 5 different provinces in Thailand analyzed by the validated HPTLC method were in the ranges of 0.098 ± 0.017 - 0.30±0.02%, w/w, and 0.081 ± 0.0006 - 0.34 ? 0.0009 %, w/w, respectively. Five commercial products of S. alata tea available in the market were purchased and analyzed for rhein and aloe-emodin contents. The contents of rhein and aloe-emodin in the tea samples were in the ranges of 0.085 ± 0.004 - 0:23 ± 0.04%, w/w, and 0.096 ±0.006 - 0.30 ± 0.01 %, w/w, respectively. The HPTLC method is rapid, reliable, sensitive and economical for routine analysis of rhein and aloe-emodin contents in-S. alata leaf raw materials and its commercial products.

Comparative Phytochemical Profiling and In Vitro Antioxidant Activity of Extracts from Raw Materials, Tissue-Cultured Plants, and Callus of Oroxylum indicum (L.) Vent.

Extracts from raw materials from different plant parts, tissue-cultured plants, and callus cultures of Oroxylum indicum were analyzed for in vitro antioxidant activities determined by DPPH radical scavenging assay and evaluated for phytochemical profiles by TLC and LC-MS methods. The results were analyzed by principal component analysis (PCA) to evaluate the similarity. Stalk, pedicel, flower, seed, and whole fruit and callus extracts promoted strong antioxidant activity with high total phenolic and total flavonoid contents. The main phytochemicals found in extracts were baicalin, baicalein, and chrysin. Baicalein and baicalin promoted strong antioxidant effects and existed in most extracts while chrysin, which promoted very low antioxidant activity, was a major flavonoid in the leaves and tissue-cultured plants. From PCA analysis by total phenolic and total flavonoid contents, four main clusters including callus and tissue-cultured plant groups from different growth stages, flower group, and whole fruit and leaf group could be organized. When the results were analyzed by PCA using antioxidant activity with total phenolic or total flavonoid contents, all O. indicum samples could be grouped together except the extracts from the root of tissue-cultured plants which separated from the rest due to their low phytochemical contents and weak antioxidant activities.

HPLC quantitative analysis of protocatechuic acid contents in 11 Phellinus mushroom species collected in Thailand

Abstract Phellinus mushrooms have been traditionally used for various medicinal purposes. Protocatechuic acid, which was previously reported to be a component in some Phellinus mushrooms, has some pharmacological effects. This study aimed to validate a HPLC method for the quantitative analysis of the protocatechuic acid contents in the extracts from different Phellinus mushroom species collected in Thailand. HPLC was carried out using a C18 column and the gradient mobile phases of 0.1% formic acid in water and 0.1% formic acid in acetonitrile. Method validation was performed to assure the linearity, accuracy, precision, limit of detection and limit of quantitation of the analytical method. The linearity range of protocatechuic acid was 1 - 10 µg/ml. The average recovery was 104.16%. The method was shown to be precise with the RSD of repeatability and intermediate precision at less than 3%. The protocatechuic contents in 11 Phellinus mushrooms were in the range of less than 0.0099 - 0.4121 %w/w of the extract. The developed HPLC method was reliable and suitable for the quantitative analysis of protocatechuic acid content in Phellinus mushrooms.

Antioxidant activity of Siamese neem tree (VP1209).

Leaves, fruits, flowers and stem bark extracts from the Siamese neem tree (Azadirachta indica A. Juss var. siamensis Valeton, Meliaceae) were assessed for antioxidant activity in vitro using the 1,1-diphenyl-2-picryl hydrazyl (DPPH) scavenging assay, total antioxidant activity and inhibition of lipid peroxidation in Chago K1 cancer cell culture by the thiobarbituric acid reactive substances (TBARS) method. The results showed that leaf aqueous extract, flower and stem bark ethanol extracts exhibited higher free radical scavenging effect on the DPPH assay with 50% scavenging activity at 26.5, 27.9 and 30.6 microg/ml, respectively. The total antioxidant activity of these extracts was found to be 0.959, 0.988 and 1.064 mM of standard trolox, respectively. At 100 microg/ml, the flower ethanol and leaf aqueous extracts significantly decreased malondialdehyde (MDA) levels (46.0 and 50.6%, respectively) by the TBARS method. The results suggest that extracts from leaf, flower and stem bark of the Siamese neem tree have strong antioxidant potential. This report supports the ethnomedical use of young leaves and flowers of this plant as a vegetable bitter tonic to promote good health.

Antioxidant Activity and Antibacterial Effects on Clinical Isolated Streptococcus suis and Staphylococcus intermedius of Extracts from Several Parts of Cladogynos orientalis and Their Phytochemical Screenings.

The in vitro antioxidant and antibacterial assays against clinically isolated Streptococcus suis and Staphylococcus intermedius of the extracts prepared by decoction and ethanolic reflux of different parts of Chettaphangki (Cladogynos orientalis Zipp. ex Span), including the leaves, roots, and stems, using 1,1-diphenyl-2-picrylhydrazyl (DPPH) scavenging assay and disc diffusion method were conducted. Quantitative analysis of total phenolic and total flavonoid contents in the extracts using spectrophotometric methods was also performed. Finally, phytochemical screening by thin layer chromatography (TLC) and high performance liquid chromatography (HPLC) was conducted. Leaf ethanolic reflux extract (100 g) contained the highest total phenolic and total flavonoid contents of 7.21 ± 0.28 µg gallic acid equivalent (GAE) and 11.51 ± 2.02 µg rutin equivalent (RE), respectively. Chettaphangki extracts promoted low antioxidant activity with EC50 values in the range of 0.27-0.48 mg/mL. Extracts and fractions from the roots and stems of this plant promoted low to intermediate antibacterial activity against S. intermedius with the inhibition zones between 7 and 14 mm. The chromatographic data suggested that the leaf extracts of C. orientalis contained rutin while the root and stem extracts contained scopoletin and chettaphanin I. Rutin promoted strong antioxidant activity while chettaphanin I showed low antibacterial activity against Staphylococcus intermedius.

Simultaneous HPLC quantitative analysis of active compounds in leaves of Moringa oleifera Lam.

Moringa oleifera Lam. has been used as a traditional medicine for the treatment of numerous diseases. A simultaneous high-performance liquid chromatography (HPLC) analysis was developed and validated for the determination of the contents of crypto-chlorogenic acid, isoquercetin and astragalin, the primary antioxidative compounds, in M. oleifera leaves. HPLC analysis was successfully conducted by using a Hypersil BDS C18 column, eluted with a gradient of methanol-1% acetic acid with a flow rate of 1 mL/min, and detected at 334 nm. Parameters for the validation included linearity, precision, accuracy and limits of detection and quantitation. The developed HPLC method was precise, with relative standard deviation < 2%. The recovery values of crypto-chlorogenic acid, isoquercetin and astragalin in M. oleifera leaf extracts were 98.50, 98.47 and 98.59%, respectively. The average contents of these compounds in the dried ethanolic extracts of the leaves of M. oleifera collected from different regions of Thailand were 0.081, 0.120 and 0.153% (w/w), respectively. The developed HPLC method was appropriate and practical for the simultaneous analysis of crypto-chlorogenic acid, isoquercetin and astragalin in the leaf extract of M. oleifera. This work is valuable as guidance for the standardization of the leaf extracts and pharmaceutical products of M. oleifera.