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AbstractContext The therapeutic potential of andrographolide is hindered by its poor oral bioavailability and unpredictable pharmacokinetics, primarily due to its limited water solubility.Objective This work aimed to enhance the solubility and pharmacokinetics of andrographolide, a bioactive compound in Andrographis paniculata (Burm. f.) Nees (Acanthaceae), using solubilizing agents and a bioenhancer.Materials and methods Four groups of beagles were compared: (1) A. paniculata powder alone (control), (2) A. paniculata powder with 50% weight/weight (w/w) β-cyclodextrin solubilizer, (3) A. paniculata powder with 1% w/w sodium dodecyl sulfate (SDS) solubilizer, and (4) A. paniculata powder co-administered with 1% w/w SDS solubilizer and 10% piperine bioenhancer. All groups received a consistent oral dose of 3 mg/kg of andrographolide, administered both as a single dose and multiple doses over seven consecutive days.Results Thirteen chemical compounds were identified in A. paniculata powder, including 7 diterpenoids, 5 flavonoids, and 1 phenolic compound. A. paniculata co-administration with either 50% w/w β-cyclodextrin or 1% w/w SDS, alone or in combination with 10% w/w piperine, significantly increased systemic andrographolide exposure by enhancing bioavailability (131.01% to 196.05%) following single and multiple oral co-administration. Glucuronidation is one possible biotransformation pathway for andrographolide, as evidenced by the excretion of glucuronide conjugates in urine and feces.Conclusion The combination of solubilizing agents and a bioenhancer improved the oral bioavailability and pharmacokinetics of andrographolide, indicating potential implications for A. paniculata formulations and clinical therapeutic benefits. Further investigation in clinical studies is warranted.

Abstract Cigarette smoke (CS) is one of the leading causes of pulmonary diseases and can induce lung secretome alteration. CS exposure-induced damages to human pulmonary epithelial cells and microvascular endothelial cells have been extensively demonstrated; however, the effects of the secretome of lung epithelial cells exposed to CS extracts (CSE) on lung microvascular endothelial cells are not fully understood. In this study, we aimed to determine the effects of the secretome of lung epithelial cells exposed to CSE on lung microvascular endothelial cells. Human lung epithelial cells, A549, were exposed to CSE, and the secretome was collected. Human lung microvascular endothelial cells, HULEC-5a, were used to evaluate the effect of the secretome of A549 exposed to CSE. Secretome profile, endothelial cell death, inflammation, and permeability markers were determined. CSE altered the secretome expression of A549 cells, and secretome derived from CSE-exposed A549 cells caused respiratory endothelial cell death, inflammation, and moderately enhanced endothelial permeability. This study demonstrates the potential role of cellular interaction between endothelial and epithelial cells during exposure to CSE and provides novel therapeutic targets or beneficial biomarkers using secretome analysis for CSE-related respiratory diseases.

Background: Cannabis sativa L. has long been used in Thai traditional medicine which has included a preparation containing powders of cannabis leaves and turmeric rhizomes, in cottonseed oil, that have been effectively used as a topical paste for haemorrhoids and skin diseases. This traditional formulation was successfully modified and developed as an ointment preparation for assisting patient compliance. In this study, our purpose was to 1) establish the quality control method, 2) study the stability of the formulation and 3) evaluate the anti-inflammatory activity of the formulated ointment and its ingredients. Methods: The ingredients in the ointment include cannabis leaf extract (CL), turmeric rhizome extract (TM) and cottonseed oil (CO). HPLC methods for the determination of cannabinoids and curcuminoids in the extracts and the ointment were validated. The ointment was subjected to stability evaluation. Each ingredient and the mixture of all ingredients (MI) were evaluated for their effects on LPS-induced pro-inflammatory cytokines in THP-1 monocytes and nitric oxide in RAW 264.7 macrophage cells. Results: The HPLC methods were accurate and precise. The ointment contained curcumin (2.10 %), bisdemethoxycurcumin (1.82 %), demethoxycurcumin (0.95 %) and delta9-tetrahydrocannabinol (0.08 %). The accelerated stability test at 50 °C for 1 month showed that the developed formula remained stable, with a shelf life of 9 months. The TM, CL and MI (10 µg/mL) expressed a high inhibitory effect on TNF-α whereas CL and MI strongly suppressed IL-1β. All samples (25 µg/mL) significantly inhibited nitric oxide production of which TM exhibited the strongest activity. Conclusion: The novel formulated ointment, derived from traditional practice for treating haemorrhoids and skin diseases, demonstrated satisfactory quality, stability, and in-vitro anti-inflammatory activity.

Abstract Cyperus rotundus rhizomes have been used in longevity remedies in Thailand for nourishing good health, which led us to investigate the effect on energy homeostasis, especially glucose utilization in myotubes and adipocytes, and on inhibition of lipogenesis in adipocytes. The results showed that an ethyl acetate extract of C. rotundus rhizomes (ECR) containing 1.61%w/w piceatannol, with a half-maximal concentration of 17.76 ± 0.03 μg/mL in 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging assay, caused upregulation and cell-membrane translocation of glucose transporters GLUT4 and 1 in L6 myotubes but downregulation and cytoplasmic localization of GLUT4 expression in 3T3-L1 adipocytes and was related to the p-Akt/Akt ratio in both cells, especially at 100 μg/mL. Moreover, ECR (25–100 μg/mL) significantly inhibited lipid accumulation via Adenosine Monophosphate-Activated Protein Kinase (AMPK), Acetyl CoA Carboxylase (ACC), and Glycogen Synthase Kinase (GSK) pathways. Its immunoblot showed increased expression of p-AMPKα/AMPKα and p-ACC/ACC but decreased expression of p-Akt/Akt and p-GSK3β/GSK3β in 3T3-L1 adipocytes. Moreover, the decreased expression of the adipogenic effectors, perilipin1 and lipoprotein lipase, in ECR-incubated adipocytes (50 and 100 μg/mL) indicated reduced de novo lipogenesis. Our study elucidated mechanisms of C. rotundus that help attenuate glucose tolerance in skeletal muscle and inhibit lipid droplet accumulation in adipose tissue.

The study aimed to investigate the antioxidant and cholinesterase inhibitory activities of the leaves and stems of Artocarpus sericicarpus and to analyse the phenolic compounds in the extracts. The modified Ellman’s method was used to determine the cholinesterase inhibitory activities against acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) enzymes. The antioxidant properties were evaluated using DPPH and ABTS methods. The total phenolic content (TPC) was measured by spectrometric assay, and compound identification was carried out by LC-MS/MS analysis. The results showed that the leaf and stem extracts of A. sericicarpus exerted significant inhibitory effects against AChE and BChE, as well as antioxidant activities. The stem ethanolic extract exhibited the highest potency against AChE and BChE with IC50 values of 5.81 and 11.46 µg/mL, respectively. The leaf and stem ethanolic extracts gave higher antioxidant activities and TPC compared to the water-based extracts. The LC-MS/MS analysis indicated the presence of phenolic compounds, such as flavones, flavonols, flavanones, prenylated chalcones, and xanthones in the extracts.

AbstractContext Attempts are ongoing to develop medications to fight against the COVID-19 pandemic. Our previous study revealed the in vitro anti-SARS-CoV-2 activity of fingerroot [Boesenbergia rotunda (L.) Mansf. (Zingiberaceae)] and its phytochemical, panduratin A.Objective To investigate the pharmacokinetic profiles of panduratin A as a pure compound and in a fingerroot extract formulation in beagle dogs.Materials and methods A total of 12 healthy dogs were randomly divided into three groups, a single dose of 1 mg/kg panduratin A by intravenous and multiple doses of 5 and 10 mg/kg panduratin A fingerroot extract formulation by oral administration for seven consecutive days. The plasma concentration of panduratin A was determined by LCMS.Results The peak concentrations of a single dose of 5 and 10 mg/kg panduratin A fingerroot extract formulation were 12,416 ± 2,326 and 26,319 ± 8,221 µg/L, respectively. Increasing the oral dose of fingerroot extract formulation, equivalent to panduratin A 5–10 mg/kg, showed dose proportionality, with an approximately 2-fold increase in Cmax and AUC. The absolute oral bioavailability of panduratin A in the fingerroot extract formulation was approximately 7–9%. The majority of panduratin A was biotransformed into several products via oxidation and glucuronidation, and predominantly excreted via the faecal route.Conclusion The oral formulation of fingerroot extract was safe in beagle dogs, and increasing dose showed dose proportionality in terms of the systemic exposure of panduratin A. This information will support the phytopharmaceutical product development of fingerroot extract against the COVID-19 pandemic.

Abstract Andrographis paniculata (Burm. F.) Nees is a medicinal plant previously reported with broad-spectrum antivirals but the mode of inhibition remains elusive. The objective of this study was to identify the most active fraction from A. paniculata ethanol extract (APE, APE-2A, APE-2B and APE-2C) and dry powder extract (APSP) against influenza A (H3N2), representing RNA viruses, and herpes simplex virus-1 (HSV-1), representing DNA viruses. The results showed that the fractions APSP, APE, APE-2B, and APE-2C directly neutralized the HSV-1 and influenza A (H3N2) when incubated at room temperature for 60 min before infecting the cells. The results also showed that the additional APE-2A fraction also directly neutralized the influenza A (H3N2), but not the HSV-1. The APE, APE-2B and APE-2C inhibited the HSV-1 by more than 0.5 log when the fractions were introduced after infection. Similarly, the APSP and APE inhibited the influenza A (H3N2) more than 0.5 log after infection. Only 50 μg/mL APE-2C inhibited the viruses greater than 0.5 log. In addition, A. paniculata extracts were also evaluated for their interfering capacities against nitric oxide (NO) production in LPS-activated RAW 264.7 macrophages. As well, APE-2C potently inhibited NO production at the IC50 of 6.08 μg/mL. HPLC and LC–MS analysis indicated that the most actively antiviral fractions did not contain any andrographolide derivatives, whereas the andrographolide-rich fractions showed moderate activity.

Abstract Industrial wastewater treatment generates sludge with high concentrations of metals and coagulants, which can cause environmental problems. This study developed a sequential sludge washing and metal recovery process for industrial sludge containing > 4500 mg/kg Cu and > 5000 mg/kg Cr. The washing agent was formulated by mixing glycolipid, lipopeptide, and phospholipid biosurfactants from Weissella cibaria PN3 and Brevibacterium casei NK8 with a chelating agent, ethylenediaminetetraacetic acid (EDTA). These biosurfactants contained various functional groups for capturing metals. The optimized formulation by the central composite design had low surface tension and contained relatively small micelles. Comparable Cu and Cr removal efficiencies of 37.8% and 38.4%, respectively, were obtained after washing the sludge by shaking with a sonication process at a 1:4 solid-to-liquid ratio. The zeta potential analysis indicated the bonding of metal ions on the surface of biosurfactant micelles. When 100 g/L iron oxide nanoparticles were applied to the washing agent without pH adjustment, 83% Cu and 100% Cr were recovered. In addition, X-ray diffraction and X-ray absorption spectroscopy of the nanoparticles showed the oxidation of nanoparticles, the reduction of Cr(V) to the less toxic Cr(III), and the absorption of Cu. The recovered metals could be further recycled, which will be beneficial for the circular economy.

[This corrects the article DOI: 10.1371/journal.pone.0302662.].

Kaab Dum, a prominent indigenous rice variety cultivated in the Pak Phanang Basin of Nakhon Si Thammarat, Thailand, is the focus of our study. We investigate the therapeutic potential of indigenous Kaab Dum rice extract in the context of chronic wounds. Our research encompasses an examination of the nutritional compositions and chemical profiles of Kaab Dum rice extract. Additionally, we assess how the extract affects chronic wounds in TGF-β-induced HaCaT cells. Our evaluation methods include the detection of cellular oxidative stress, the examination of endoplasmic reticulum (ER) stress, wound healing assays, analysis of cell cycle arrest and the study of cellular senescence through senescence-associated β-galactosidase (SA-β-gal) staining. Our research findings demonstrate that TGF-β induces oxidative stress in HaCaT cells, which subsequently triggers ER stress, confirmed by the expression of the PERK protein. This ER stress results in cell cycle arrest in HaCaT cells, characterized by an increase in p21 protein, a cyclin-dependent kinase inhibitor (CDKI). Ultimately, this leads to cellular senescence, as confirmed by SA-β-gal staining. Importantly, our study reveals the effectiveness of Kaab Dum rice extract in promoting wound healing in the chronic wound model. The extract reduces ER stress and senescent cells. These beneficial effects are potentially linked to the antioxidant and anti-inflammatory properties of the rice extract. The findings of our study have the potential to make significant contributions to the development of enhanced products for both the prevention and treatment of chronic wounds.

Gynmena inodorum (GI) is a green leafy vegetable used in the Northern Thai cuisine which has antioxidant activities and may be applicable for preventing oxidative stress and aging-related disease. However, understanding the relationship between GI phytonutrients and their antioxidant properties has been unclear. The aims of this study were to identify the GI leaf phytochemicals and to study their antioxidant activities. A chromatogram of LC-ESI-MS/QTOF-MS showed that the GI leaves were potentially composed of phenolics, quinic acids, flavonoids, and triterpenoid saponins. This study was able to authenticate quercetin, kaempferol, and triterpenoid GIA1 in the samples. The GI materials with high contents of phenolics, flavonoids, quercetin, and kaempferol showed significant relation to antioxidation and protection in endothelial cell death suppressed by reactive nitrogen species. Meanwhile, triterpenoids had a low antioxidant impact. Ultimately, GI leaves with high phenolic compounds are a promising raw material to develop as an antioxidant functional food.

Context: Marine sources such as sponges have shown a significant impact on the drug development from nature. Metabolites isolated from sponges show diversity in terms of structural features and pharmacological properties. Several sponges have been reported to have potency as cholinesterase inhibitors as one of the target therapies for Alzheimer’s disease. Aims: To investigate the potency of marine sponge Aaptos suberitoides as cholinesterase inhibitors and to explore the chemistry of the sponge. Methods: The cholinesterase inhibitory assay was carried out against two enzymes, acetylcholinesterase (AChE) and butyrylcholinesterase (BChE), based on the modified Ellman’s method. The chemistry of the active fractions was studied by LC-MS/MS method, followed by molecular networking using GNPS. Results: The results suggested that the extract and fractions inhibited both AChE and BChE enzymes. All samples demonstrated more potent inhibition against AChE compared to BChE enzymes. The n-hexane fraction gave the strongest inhibition against both AChE and BChE, with IC50 values of 4.76 µg/mL and 6.79 µg/mL, respectively. Based on the LC-MS/MS analysis, alkaloids were detected in the n-hexane and ethyl acetate fractions. Four alkaloids were identified in the ethyl acetate fraction, namely demethylaaptamine, aaptamine, isoaaptamine, and 8,9,9-trimethoxy-9H-benzo[de][1,6]naphthyridine at RT 1.52, 1.67, 2.92, and 3.22 mins, respectively. Aaptamine was also identified in the n-hexane fraction together with demethyloxyaaptamine. Conclusions: The extract, n-hexane, and ethyl acetate fractions of A. suberitoides have shown promising cholinesterase inhibitory properties against both AChE and BChE enzymes. The alkaloids present in the active fractions may be responsible for the bioactivity.

Abstract Eulophia macrobulbon (E.C.Parish & Rchb.f.) Hook.f. contains a natural PDE5A1 inhibitor, phenanthrene, 1-(4'-hydroxybenzyl)-4,8- dimethoxyphenanthrene-2,7-diol (HDP), a potential agent for the treatment of erectile dysfunction. The aim of this study was to improve the extraction efficiency of HDP from E. macrobulbon by using a more environmentally friendly extraction method, subcritical liquid dimethyl ether extraction (sDME), instead of classical solvent extraction (CSE) and ultrasound-assisted extraction (UAE). The efficiency and quality of the extracts obtained were evaluated using the following criteria: %process yield; solvent amount; extraction time; temperature; %HDP content by LC–MS, bioactivity as inhibition of phosphodiesterase-5A1 (PDE5A1) by radio-enzymatic assay; and chemical profiles by LC-QTOF-MS. sDME provided the highest content of HDP in the extract at 4.47%, much higher than the use of ethanol (0.4–0.5%), ethyl acetate (1.2–1.7%), or dichloromethane (0.7–1.4%). The process yield for sDME (1.5–2.7%) was similar to or lower than the other solvents (0.9–17%), but as long as the process yield is not prohibitively low, the concentration is a more important measure for clinical use. The optimal conditions for sDME extraction were: Extraction time, 40 min; 200% water as co-solvent; sample-to-solvent ratio of 1:8; temperature, 35 °C. Phenanthrene aglycone and glycoside derivatives were the major constituents of the sDME extracts and lesser amounts of phenolic compounds and sugars. The inhibition of PDE5A1 by sDME (IC50 0.67 ± 0.22 µg/ml) was tenfold more potent than ethanolic extract and other extraction methods, suggesting a high probability of clinical efficacy. Thus, sDME was a more efficient, faster, solvent-saving and environmentally friendly extraction method and more selective for phenanthrene when extracted from E. macrobulbon.

Abstract A newly standardised extract of Centella asiatica (Centell-S) with better water solubility than the previous standardised extract of C. asiatica (ECa 233) was developed, and pharmacokinetic profiles of bioactive triterpenoids were investigated in beagle dogs. The test substances were administered via intravenous or oral administration with single and multiple doses for 7 days. The concentrations of major bioactive triterpenoids, including madecassoside, asiaticoside, madecassic acid, and asiatic acid, in biological samples were measured by liquid chromatography–tandem mass spectrometry. The dogs in this study showed good tolerability to all test substances, based on the physical appearance and blood chemistry 24 h after dosing. The major bioactive triterpenoids found in systemic blood circulation were madecassoside, asiaticoside, and asiatic acid; the concentration of these components ranged from 1 to 10,000 µg/L after intravenous administration of 1.0 mg/kg Centell-S. Oral administration of 10 and 20 mg/kg Centell-S generated approximately twofold higher plasma levels of both madecassoside and asiaticoside compared with equivalent doses of ECa 233. In addition, there was an accumulation of triterpenoid glycosides after multiple oral administrations of Centell-S for 7 days, while triterpenic acids showed little tendency for accumulation. Beagles had good tolerability to both standardised extracts of C. asiatica, and showed a similar pattern of bioactive triterpenoids to humans. Centell-S increased oral bioavailability of major triterpenoid glycosides and can be further developed into a phytopharmaceutical product.

Background. Cassia spectabilis is a flowering plant containing various metabolites that provide potential for pharmacological activities. The current study aimed to investigate the ethanolic and water extracts of C. spectabilis as cholinesterase inhibitor as one of the target treatments for Alzheimer’s disease. The chemical composition of the extracts was also studied to determine which components are responsible for the bioactivity. Methods. The cholinesterase inhibitory activity assay was carried out by the modified Ellman’s method against acetylcholinesterase (AChE) and butyrylcholinesterase (BChE). LC-MS/MS analysis was carried out to investigate the chemical profiles of the extracts, followed by a molecular networking study by GNPS. Results. Both extracts showed inhibition against AChE and BChE in a dose-dependent manner, with the higher potency exhibited by the ethanolic extract with IC50 values of 7.88 and 3.78 μg/mL. The chemical analysis and molecular networking study of the flower extracts revealed similarity between the ethanolic and water extracts. Piperidine alkaloids were identified in both extracts, while the sphingolipid compounds were found in the ethanolic extract. Conclusion. The water and ethanolic extracts of C. spectabilis flowers displayed potency for Alzheimer’s disease treatment. The presence of piperidine alkaloids in the extract may be responsible for the cholinesterase inhibitory activity. The higher potency of the ethanolic extract compared to the water extract is possibly due to the higher amount of piperidine alkaloids in the ethanolic extract. Further study is needed to quantify the concentration of alkaloids in the extracts.

Abstract Since 2006, Ficus dubia has been reported as a new Ficus species in Thailand. As per our recent report, the red-brown aqueous extract of F. dubia sap (FDS) has been determined to strongly exhibit in vitro anti-radicals. However, the phytochemicals in the FDS extract related to health-promoting antioxidation have not been explored. Thus, in this study, we aimed to investigate the chemical components of the F. dubia sap extract by liquid chromatography-electrospray ionization quadrupole time-of-flight mass spectrometry (LC–ESI–MS/QTOF-MS) and its potential use in cosmetics in terms of cellular antioxidation on keratinocytes (HaCaT), phototoxicity, and irritation on 3D skin cell models following standard tests suggested by the Organization for Economic Cooperation and Development (OECD). It was found that the sap extract was composed of quinic acid and caffeoyl derivatives (e.g., syringoylquinic acid, 3-O-caffeoylquinic acid, 4-O-caffeoylquinic acid, and dimeric forms of caffeoylquinic acids). The extract has significantly exhibited antioxidant activity against H2O2-induced oxidative stress in HaCaT cells. The cellular antioxidative effect of the FDS extract was remarkably dependent on the presence of 3- and 4-O-caffeoylquinic acid in the extract. Furthermore, the FDS extract showed negative results on skin phototoxicity and irritation. Overall, the results reveal that the FDS extract could be developed as a new antioxidant candidate for a skin healthcare product.

Centotheca lappacea (L.) Desv. or “hee yum” grass has been used in Thailand for postpartum care. This study focused on analyses of elements, antioxidants and volatile compounds from C. lappacea. Wavelength dispersive X-ray fluorescence showed the presence of silicon in this plant. Liquid chromatography–electrospray ionization-quadrupole-time of flight-mass spectrometer coupled with DPPH assay was used for separation, detection and identification of antioxidants in C. lappacea in one run. Twelve antioxidants including C-and O-glycosides of luteolin and apigenin were identified. Moreover, a headspace gas chromatography-mass spectrometer was used for analyses of volatile compounds by direct burning of the plant material, which is similar to how smoke is made during a traditional treatment. Nineteen and twenty-five compounds were found from the roots and the aerial parts of C. lappacea, respectively. In conclusion, antioxidants and volatile constituents after C. lappacea has been burnt are reported here for the first time.

Rice bran is an essential source of natural bioactive compounds with widely applications. This study aimed to determine the effects of extraction methods on the bioactivities of the extracts. Jasmine and Riceberry rice bran were extracted using 50% ethanolic maceration, cold pressed and subcritical dimethyl ether (DME) extraction. The extracts were analyzed for antioxidant, pro-collagen type I synthesis and 5α-reductase inhibition activities. The results showed that Riceberry rice bran extract exhibited superior antioxidant activity than Jasmine rice bran extract. The highest pro-collagen type I synthesis of Jasmine and Riceberry rice resulted from subcritical DME extraction (50.91%) and maceration (46.99%) respectively. It is a newly found that the subcritical DME extract of Jasmine rice bran displayed the strongest 5α-reductase inhibition activity (73.14 %inhibition) resulting from the biological markers linoleic acid (22.39±0.61 %w/w) and oleic acid (39.48±1.07 %w/w). Therefore, subcritical DME extraction is suitable for preparing rice bran extract for 5α-reductase inhibitor application.

Abstract Background Cucurbitacin B is the major bioactive constituent in Trichosanthes cucumerina L. fruits, which the pharmacological properties have been studied for decades particularly an anti-tumor activity. The pharmacokinetic profile of this compound is still limited and investigation is needed for further phytopharmaceutical product development. This study aimed to investigate the pharmacokinetic profile of cucurbitacin B after administering the compound at different doses and routes to rats. Methods Male Wistar rats (n = 6) were treated by cucurbitacin B extracted from Trichosanthes cucumerina L. The cucurbitacin B was administered at 0.1 mg/kg intravenously or by oral gavage at 2–4 mg/kg. Blood samples and internal organs were collected serially within 24 h after administration. Urine and feces were collected from time 0 to 48 h. The level of cucurbitacin B in biological samples was determined by liquid chromatography-tandem mass spectrometry. Results The absolute oral bioavailability of cucurbitacin B was approximately 10%. The maximum concentration in plasma after normalization by dose ranged from 4.85–7.81 μg/L and the time to reach maximum value was approximately within 30 min after oral dosing. The level of cucurbitacin B in plasma increased proportionally to the given dose. After intravenous administration, cucurbitacin B had a large volume of distribution of about 51.65 L/kg and exhibited a high tissue to plasma concentration ratio, approximately 60 to 280-fold in several organs. Negligible amount of unchanged cucurbitacin B could be detected in urine and feces and accounted less than 1% of administered dose. Conclusion Cucurbitacin B had low oral bioavailability, but could be distributed extensively into internal organs with a high volume of distribution and tissue to plasma ratio. Only negligible amounts of unchanged cucurbitacin B were excreted via urine and feces suggesting that the compound might be biotransformed before undergoing an excretion. Further studies of the metabolic pathway and tissue uptake mechanism are required to strategize the future development of cucurbitacin B into clinical studies.

The holistic approach of herbal medicines is becoming acceptable in many parts of the world. In order to determine, not only chemical but also biological activity profiles of traditional medicines, on-line liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS) coupled to antioxidant assay was introduced. Here, we demonstrate the performance of this on-line monitor for quantitatively and qualitatively analyses of antioxidants in Sahasthara, a Thai traditional medicine. HPLC real-time coupling to ESI-MS and DPPH assay was applied to separate and identify the constituent compounds responsible for antioxidant activity. The antioxidant fingerprint was obtained together with the LC-MS chromatogram. The full scan MS and the multiple reaction monitoring (MRM) were used for structural confirmation and quantitation. Peak identification was performed by comparing retention times and mass spectra with reference compounds and published data. Based on their mass spectra, 13 antioxidant compounds were identified as gallic acid, chebulic acid, digalloyl glucoside, caffeoylferuloylquinic lactone, galloylshikimic acid, trigalloyl glucoside, corilagin, chebulanin, chebulagic acid, dicaffeoylquinic lactone, dicaffeoyl quinic acid, ellagic acid rhamnoside and ellagic acid. Gallic acid and ellagic acid were further quantitated and comprised 126.8±6.0 mg/g and 253.7±17.6 mg/g of Sahasthara, respectively. This on-line monitor is shown to be an effective tool for fast and simultaneous analyses of antioxidant compounds in complex formulations characteristic of many traditional medicines.

Background: This study explored the bioactivities and nutrient compositions of coffee (Coffea Arabica L.) pulp which was prepared in three different ways [Coffee Pulp Extracts (CPE) 1–3]. Methods: The coffee pulp was prepared in three different ways by distinct selecting and freezing processes. The nutritional values, polyphenol contents, antioxidant activity, and antibacterial properties of the coffee pulp as well as the characterization of the active ingredients by liquid chromatography-electrospray ionization-quadrupole-time-of-flight mass spectrometry (LC-ESI-Q-TOF-MS) were evaluated. Results: The chemical profiles of three aqueous extracts were compared and characterized using LC-ESI-QTOF-MS. They showed slightly different nutrient compositions. The total phenolic content was highest in CPE1, and decreased in the following order: CPE1 > CPE2 > CPE3. Among the CPEs tested, CPE1 showed the most potent antioxidant activity with IC50 18 μg/mL and 82 μg/mL by 2,2’-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) and 1,1-diphenyl-2-picryl-hydrazyl assay, respectively. Chlorogenic acid and caffeine were the most prominent in CPE1 and it contained more compounds than the others. Moreover, CPE1 demonstrated antibacterial activity against both gram-positive (Staphylococcus aureus and Staphylococcus epidermidis) and gram-negative bacteria (Pseudomonas aeruginosa and Escherichia coli). Conclusion: These findings indicated that CPE1 has powerful nutrients with antioxidant and antibacterial properties—the potency of which is impacted by the preparation process.

Brahmi essence, developed from Bacopa monnieri (L.) Wettst. standardized extract and mulberry juice, was proven to improve the memory speed of healthy participants aged 55–80 years old, following a 12-week dietary program. However, the metabolites have not yet been reported. Our objective was to characterize the altered metabolites in the plasma, urine, and feces of healthy volunteers after consumption of Brahmi essence for 12 weeks, using the LC-MS metabolomics approach. The altered metabolites were selected from OPLS-DA S-plots; 15 metabolites in the plasma, 7 in the urine, and 17 in the feces samples were tentatively identified by comparison with an online database and literature. The metabolites in the plasma samples were in the classes of amino acids, acylcarnitine, and phospholipids. Benzeneactamide-4-O-sulphate and 3-hydroxyhippuric acid were found in urine samples. The metabolites in the class of amino acids, together with jujubogenin and pseudojujubogenin, were identified in the fecal samples. The aminoacyl-tRNA, aromatic amino acids, and branched-chain amino acid biosynthetic pathways were mainly related to the identified metabolites in all three samples. It could be implied that those metabolites and their pathways might be linked with the effect of Brahmi essence on memory speed.

Citrus hystrix DC. (CH) is found in many countries in Southeast Asia. This plant has been reported for anti-microbial, anti-cancer and anti-inflammatory bioactivities. However, the anti-inflammatory and anti-inflammasome properties of the leaves remain poorly understood. This study aimed to investigate the effect of CH leaves on NLRP3 and NF-κB signaling pathways. CH leaves were sequentially extracted using hexane, ethyl acetate and 95% ethanol to give three crude extracts. An active compound, lupeol was fractionated from the ethanolic extract using chromatographic techniques, and its structure was identified and confirmed by spectroscopic methods. Anti-inflammatory activities were observed on both lipopolysaccharide-stimulated and NLRP3 adenosine triphosphate-induced macrophages. The release of pro-inflammatory cytokines (IL-1β, IL-6 and TNF-α) was analyzed by Enzyme-Linked Immunosorbent Assay (ELISA). Real-time qRT-polymerase chain reaction (PCR) was used to measure inflammatory-associated gene expression. NF-κB protein expressions were investigated using the immunoblotting technique. The active fraction of ethanolic CH leaves and lupeol significantly reduced the release of pro-inflammatory cytokines and suppressed the expression of both inflammasome genes and NF-κB proteins. The ethanolic extract of CH leaves and lupeol showed potent anti-inflammatory activities by targeting NF-κB and NLRP3 signaling pathways.

Moringa oleifera Lam. (MO) is a medicinal plant distributed across the Middle East, Asia, and Africa. MO has been used in the traditional treatment of various diseases including cancer. This study aimed to perform bioassay-guided fractionation and identification of bioactive compounds from MO leaf against MDA-MB-231 breast cancer cells. MO leaf was sequentially extracted with hexane, ethyl acetate (EtOAc), and ethanol. The most effective extract was subjected to fractionation. MO extract and its derived fractions were continuously screened for anti-cancer activities. The strongest fraction was selected for re-fractionation and identification of bioactive compounds using LC-ESI-QTOF-MS/MS analysis. The best anticancer activities were related to the fraction no. 7-derived crude EtOAc extract. This fraction significantly reduced cell viability and clonogenic growth and increased cells apoptosis. Moreover, sub-fraction no. 7.7-derived fraction no. 7 was selected for the identification of bioactive compounds. There were 10 candidate compounds tentatively identified by LC-ESI-QTOF-MS. Three of identified compounds (7-octenoic acid, oleamide, and 1-phenyl-2-pentanol) showed anticancer activities by inducing cell cycle arrest and triggering apoptosis through suppressed Bcl-2 expression which subsequently promotes activation of caspase 3, indicators for the apoptosis pathway. This study identified 10 candidate compounds that may have potential in the field of anticancer substances.

Squamous cell carcinoma is the most common type of head and neck cancer worldwide. Radiation and chemotherapy are general treatments for patients; however, these remedies can have adverse side effects and tumours develop drug resistance. Effective treatments still require improvement for cancer patients. Here, we investigated the anti-cancer effect of Moringa oleifera (MO) Lam. leaf extracts and their fractions, 3-hydroxy-β-ionone on SCC15 cell line. SCC15 were treated with and without MO leaf extracts and their fractions. MTT assay was used to determine cell viability on SCC15. Cell cycle and apoptosis were evaluated by the Muse™ Cell Analyser. Colony formation and wound closure analysis of SCC15 were performed in 6-well plates. Apoptosis markers were evaluated by immunoblotting. We found that Moringa extracts and 3-HBI significantly inhibited proliferation of SCC15. Moreover, they induced apoptosis and cell cycle arrest at G2/M phase in SCC15 compared to the untreated control. MO extracts and 3-HBI also inhibited colony formation and cell migration of SCC15. Furthermore, we observed the upregulation of cleaved caspase-3 and Bax with downregulation of anti-apoptotic Bcl-2, indicating the induction of cancer cell apoptosis. Our results revealed that MO extracts and 3-HBI provided anti-cancer properties by inhibiting progression and inducing apoptosis of SCC15.

Moringa oleifera (MO) is an important plant for traditional medicine. The present study aimed to identify the MO active phytochemical compounds for their ability against inflamed macrophages. An ethyl acetate extract fraction of MO was fractionation by flash column chromatography. Human macrophages were stimulated by Lipopolysaccharide and then treated with fractions of MO to examine their anti-inflammatory activity and cellular mechanism. The active fractions were analyzed by liquid chromatography with electrospray ionization quadrupole time-of-flight mass spectrometer (LC-ESI-QTOF-MS). MO treated cells showed a decreased production of pro-inflammatory mediator in response to lipopolysaccharide. This was evident at both mRNA and protein levels. The study revealed that MO suppressed mRNA expression of IL-1, IL-6, TNF-α, PTGS2, NF-κB (P50), and RelA. Furthermore, the extract effectively inhibited the expression of inflammatory mediators, including IL-6, TNF-α, and cyclooxygenase-2. Interestingly, the effect of MO inhibited phosphorylation of IκB-α and the ability to reduce expression of the nuclear factor (NF)-κB p65, suppressing its nuclear translocation. Moreover, LC-ESI-QTOF-MS analysis of the MO active fraction revealed seven compounds, namely 3,4-Methyleneazelaic acid, (2S)-2-phenylmethoxybutane-1,4-diol, (2R)-2-phenylmethoxybutane-1, 4-diol, γ-Diosphenol, 2,2,4,4-Tetramethyl-6-(1-oxobutyl)-1,3,5-cyclohexanetrione, 3-Hydroxy-β-ionone, and Tuberonic acid. Our findings highlight the ability of MO compounds to inhibit inflammation through regulation of the NF-κB pathway.

A major goal in the discovery of bioactive natural products is to rapidly identify active compound(s) and dereplicate known molecules from complex biological extracts. The conventional bioassay-guided fractionation process can be time consuming and often requires multi-step procedures. Herein, we apply a metabolomic strategy merging multivariate data analysis and multi-informative molecular maps to rapidly prioritize bioactive molecules directly from crude plant extracts. The strategy was applied to 59 extracts of three Bacopa species (B. monnieri, B. caroliniana and B. floribunda), which were profiled by UHPLC-HRMS2 and screened for anti-lipid peroxidation activity. Using this approach, six lipid peroxidation inhibitors 1−6 of three Bacopa spp. were discovered, three of them being new compounds: monnieraside IV (4), monnieraside V (5) and monnieraside VI (6). The results demonstrate that this combined approach could efficiently guide the discovery of new bioactive natural products. Furthermore, the approach allowed to evidence that main semi-quantitative changes in composition linked to the anti-lipid peroxidation activity were also correlated to seasonal effects notably for B. monnieri.