Oral Glucose Tolerance Test (OGTT) Evidence for the Postprandial Anti-Hyperglycemic Property of Salacca zalacca (Gaertn.) Voss Seed Extract.

Salak seed extract (Salacca zalacca) is known for its high antioxidant content and low caffeine levels, making it a promising candidate for the development of value-added health products. However, there is a lack of scientific evidence for its anti-hyperglycemic effects. To address this, we investigated the in vitro and in vivo anti-hyperglycemic and antioxidant effects of salak seed extract. The HPLC chromatogram of salak seed extract shows a prominent peak that corresponds to chlorogenic acid. In vitro studies revealed that salak seeds inhibited α-glucosidase activity and glucose uptake in Caco-2 cells in a concentration-dependent manner, while also exhibiting antioxidant properties. The extract exhibits a non-competitive inhibition on α-glucosidase activity, with an IC50 and Ki of 16.28 ± 7.22 and 24.81 µg/mL, respectively. In vivo studies utilizing streptozotocin-nicotinamide-induced diabetic mice showed that the extract significantly reduced fasting blood glucose (FBG) levels in the oral glucose tolerance test. Continuous administration of the salak seed extract resulted in lower FBG levels by 13.8% as compared with untreated diabetic mice, although this change was not statistically significant. The estimated LD50 value of salak seed extract exceeds 2000 mg/kg, and no toxicity symptoms have been detected. Our research supports that salak seed extract has the potential to serve as a functional food or supplement that may be beneficial in reducing postprandial hyperglycemia among people with type 2 diabetes. This effect was explained by the salak's inhibitory mechanisms of glucose absorption due to inhibition of both α-glucosidase activity and intestinal glucose uptake, coupled with its antioxidant effects.

In vitro and in vivo evidence of hypouricemic and anti-inflammatory activities of Maclura cochinchinensis (Lour.) Corner heartwood extract.

Maclura cochinchinensis (Lour.) Corner (MC) heartwood extracts have been used for the treatment of gout, hyperuricemia, and inflammation in Thai traditional medicine. Despite their traditional use, their mechanisms of action remain unknown. The aim of this study was to determine the mechanisms of MC heartwood extract activity using both in vitro and in vivo models . The extraction methods were optimized to yield the highest contents of biochemical compounds and antioxidant activities. The effects of MC heartwood extract on xanthine oxidase and its enzyme kinetics were determined in vitro and the antihyperuricemic effect was evaluated in potassium oxonate (PO)-induced hyperuricemic mice. The anti-inflammatory effect of MC heartwood extract was also tested against lipopolysaccharide-induced proinflammatory mRNA upregulation in RAW 264.7 mouse macrophage cells. Soxhlet extraction of MC heartwood with 70% ethanol produced stronger antioxidant activity, and higher total phenolic and flavonoid contents than conventional methods did (maceration or decoction). By using HPLC, we found that MC contains morin as a major constituent, which may account for its pharmacological activities. Moreover, administration of MC heartwood extract (500 mg/kg) markedly decreased uric acid levels in PO-induced hyperuricemic mice (p < 0.05). MC heartwood extract inhibited the hepatic activity of xanthine oxidase ex vivo by approximately 53%. In addition, MC heartwood extract markedly downregulated mRNA expression of inflammatory mediators (TNF-α, TGF-ß, iNOS, and COX-2) and this inhibition was comparable with that of dexamethasone. Therefore, MC heartwood extract is a promising candidate as a natural treatment for inflammation and the hyperuricemia that causes gout.

Development and validation of a high-performance thin layer chromatography method for the simultaneous quantitation of α- and γ-mangostins in Thai stingless bee propolis

ABSTRACT Stingless bees (Apoidea) are widely distributed and commercially cultivated in artificial hives in fruit gardens. Their propolis are commonly used in traditional medicine to treat various diseases (e.g., abscesses, inflammations, and toothaches) and as a constituent of numerous health products. Thus, this study aimed to (i) develop and validate a high-performance thin layer chromatography method for the quantitation of major active constituents (α- and γ-mangostins) in propolis produced by five stingless bee species (Tetragonula fuscobalteata Cameron, T. laeviceps Smith, T. pagdeni Schwarz, Lepidotrigona terminata Smith, and L. ventralis Smith) cultivated in Thai mangosteen orchards and (ii) determine an optimal extraction solvent. Separation was performed on a silica gel 60 F254 plate using toluene/ethyl acetate/formic acid (8:2:0.1, v/v/v) as a mobile phase, and the developed method was validated to assure its linearity, precision, accuracy, and limits of detection/quantitation. Propolis extract from T. fuscobalteata exhibited the highest mangostin content, and acetone was shown to be more a more effective extraction solvent than dichloromethane, ethanol, or methanol. Thus, the simplicity and reliability of the developed method make it well suited for the routine analysis (e.g., for quality control) of commercial products containing stingless bee propolis.

Simultaneous HPTLC quantification of three caffeoylquinic acids in Pluchea indica leaves and their commercial products in Thailand

ABSTRACT In Thai traditional medicine, Pluchea indica (L.) Less., Asteraceae, leaf has been widely used for the treatment of diabetes mellitus, tumors, hypertension, cystitis, and wounds. P. indica herbal tea is commercially available in Thailand as a health-promoting drink. The study was conducted to develop and validate a high-performance thin-layer chromatography (HPTLC) method for the quantitative analysis of chlorogenic acid, 3,4-O-dicaffeoylquinic acid, and 3,5-O-dicaffeoylquinic acid in P. indica leaf extract and their commercial products in Thailand. The method was validated according to ICH guidelines. The proposed HPTLC method showed acceptable validation parameters. The content of chlorogenic acid, 3,4-O-dicaffeoylquinic acid, and 3,5-O-dicaffeoylquinic acid in P. indica leaves from seven different provinces in Thailand was in the range of not detectable −1.94 ± 0.02%w/w, 0.71 ± 0.01-1.89 ± 0.05%w/w, and 1.00 ± 0.01-2.18 ± 0.03%w/w, respectively, while in the commercial products, it was in the range of 0.59 ± 0.03-2.17 ± 0.05%w/w, 0.53 ± 0.04-3.77 ± 0.03%w/w, and 0.88 ± 0.05-4.72 ± 0.10%w/w, respectively. The results indicated that plantation of P. indica in coastal saline land would be beneficial as it would increase the concentration of its active compounds and improve its medicinal quality. The developed high-performance thin-layer chromatography could be used as a rapid, reliable, less demanding, and cost-effective analytical method.

Optimization of extraction method and HPLC analysis of six caffeoylquinic acids in Pluchea indica leaves from different provenances in Thailand

ABSTRACT Pluchea indica (L.) Less., Asteraceae, is a medicinal plant which contains a high amount of phenolic compounds such as caffeoylquinic acid derivatives. The leaves have been traditionally used as a nerve tonic and extensively as herbal tea. This study aimed to develop and validate an HPLC method to quantitatively analyze six caffeoylquinic acid derivatives, viz. 3-O-caffeoylquinic acid, 4-O-caffeoylquinic acid, 5-O caffeoylquinic acid, 3,4-O-dicaffeoylquinic acid, 3,5-O-dicaffeoylquinic acid, and 4,5-O-dicaffeoylquinic acid in P. indica leaf extract. HPLC was carried out in a Hypersil BDS C18-column eluted with 0.5% acetic acid in water and methanol using gradient elution with a flow rate of 1 ml/min and detection at 326 nm. The method validation was performed to assure its linearity, precision, accuracy and limits of detection and quantitation. Several extraction techniques including maceration, decoction, digestion, Soxhlet extraction, and ultrasound extraction, were used to extract active constituents. The ultrasound extraction with 50% ethanol yielded the highest concentration of these caffeoylquinic acid derivatives in the P. indica leaf extract. Our developed HPLC method is simple and reliable for a routine analysis of the six caffeoylquinic acids in P. indica leaves and could potentially be applied to be used in commercial herbal products.

In vitro alpha glucosidase inhibition and free-radical scavenging activity of propolis from Thai stingless bees in mangosteen orchard

ABSTRACTThe chemical component and biological activity of propolis depend on flora area of bee collection and bee species. In the study, the propolis from three stingless bee species, Lepidotrigona ventralis Smith, Lepidotrigona terminata Smith, and Tetragonula pagdeni Schwarz, was collected in the same region of mangosteen garden from Thailand. Total phenolic content, alpha glucosidase inhibitory effect, and free-radical scavenging activity using FRAP, ABTS, DPPH assays were determined. The most potent activity of propolis extract was investigated for bioactive compounds and their quantity. The ethanol extract of T. pagdeni propolis had the highest total phenolic content 12.83 ± 0.72 g of gallic acid equivalents in 100 g of the extract, and the strongest alpha glucosidase inhibitory effect with the IC50 of 70.79 ± 6.44 µg/ml. The free-radical scavenging activity evaluated by FRAP, ABTS, DPPH assays showed the FRAP value of 279.70 ± 20.55 µmol FeSO4 equivalent/g extract and the IC50 of 59.52 ± 10.76 and 122.71 ± 11.76 µg/ml, respectively. Gamma- and alpha-mangostin from T. pagdeni propolis extract were isolated and determined for the biological activity. Gamma-mangostin exhibited the strongest activity for both alpha glucosidase inhibitory effect and free-radical scavenging activity. Using HPLC quantitative analysis method, the content of gamma- and alpha-mangostin in the extract was found to be 0.94 ± 0.01 and 2.77 ± 0.08% (w/w), respectively. These findings suggested that T. pagdeni propolis may be used as a more suitable raw material for nutraceutical and pharmaceutical products and these mangostin derivatives as markers.

Antioxidant Activity and Induction of mRNA Expressions of Antioxidant Enzymes in HEK-293 Cells of Moringa oleifera Leaf Extract.

The leaves of Moringa oleifera, collected in different provinces in Thailand, were determined for the contents of total phenolics, total flavonoids, major components, and antioxidant activity. The extract and its major active components were investigated for the inhibition of H2O2-induced reactive oxygen species production and the effects on antioxidant enzymes mRNA expression. The extract, crypto-chlorogenic acid, isoquercetin and astragalin, significantly reduced the reactive oxygen species production inducing by H2O2 in HEK-293 cells. Treatment with isoquercetin significantly increased the mRNA expression levels of antioxidant enzymes such as superoxide dismutase, catalase and heme oxygenase 1. These results confirm that M. oleifera leaves are good sources of natural antioxidant with isoquercetin as an active compound.

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.

Simultaneous Determination of Crypto-Chlorogenic Acid, Isoquercetin, and Astragalin Contents in Moringa oleifera Leaf Extracts by TLC-Densitometric Method.

Moringa oleifera Lamarck (Moringaceae) is used as a multipurpose medicinal plant for the treatment of various diseases. Isoquercetin, astragalin, and crypto-chlorogenic acid have been previously found to be major active components in the leaves of this plant. In this study, a thin-layer-chromatography (TLC-)densitometric method was developed and validated for simultaneous quantification of these major components in the 70% ethanolic extracts of M. oleifera leaves collected from 12 locations. The average amounts of crypto-chlorogenic acid, isoquercetin, and astragalin were found to be 0.0473, 0.0427, and 0.0534% dry weight, respectively. The method was validated for linearity, precision, accuracy, limit of detection, limit of quantitation, and robustness. The linearity was obtained in the range of 100-500 ng/spot with a correlation coefficient (r) over 0.9961. Intraday and interday precisions demonstrated relative standard deviations of less than 5%. The accuracy of the method was confirmed by determining the recovery. The average recoveries of each component from the extracts were in the range of 98.28 to 99.65%. Additionally, the leaves from Chiang Mai province contained the highest amounts of all active components. The proposed TLC-densitometric method was simple, accurate, precise, and cost-effective for routine quality controlling of M. oleifera leaf extracts.

In vitro inhibitory effects of Moringa oleifera leaf extract and its major components on chemiluminescence and chemotactic activity of phagocytes.

The ethanol extract of Moringa oleifera Lam. leaves and its major constituents, crypto-chlorogenic acid, quercetin 3-O-glucoside and kaempferol 3-O-glucoside, were investigated on the respiratory burst of human whole blood and isolated human polymorphonuclear leukocytes (PMNs) using a luminol-based chemiluminescence assay. The chemotactic migration of PMNs was also investigated using the Boyden chamber technique. The ethanol extract demonstrated inhibitory activities on the oxidative burst and the chemotactic migration of PMNs. Quercetin 3-O-glucoside, crypto-chlorogenic acid, and kaempferol 3-O-glucoside, isolated from the extract, expressed relatively strong inhibitory activity on the oxidative burst of PMNs with IC50 values of 4.1, 6.7 and 7.0 microM, respectively, comparable with that of aspirin. They also demonstrated strong inhibition of chemotatic migration of PMNs with IC50 values of 9.5, 15.9 and 18.2 microM, respectively. The results suggest that M. oleifera leaves could modulate the immune response of human phagocytes, linking to its ethnopharmacological use as an anti-inflammatory agent. The immunomodulating activity of the plant was mainly due to its major components.

Sequencing analysis of the medicinal plant Stemona tuberosa and five related species existing in Thailand based on trnH-psbA chloroplast DNA.

In Thailand, Stemona tuberosa Lour. , S. phyllantha Gagnep. , S. collinsae Craib , S. burkillii Prain , S. aphylla Craib and S. sp. are found. The identification based on morphological characters alone is difficult and can lead to confusion regarding chemical constituents and biological activities. The tuberous roots of S. tuberosa have long been used for treatment of respiratory diseases and as anthelmintics. However, accurate identification of S. tuberosa is needed to ensure efficacy. Sequence comparison indicated that these Stemona spp. could be identified from the sequence of the trnH- psbA locus. As a result of different sequence lengths, the PCR products generated from newly designed primers could be used to preliminarily group the two species, S. tuberosa and S. phyllantha, apart from others. However, these products could be further sequenced to discriminate among Stemona spp.