Xanthine oxidase inhibition study of isolated secondary metabolites from Dolichandrone spathacea (Bignoniaceae): In vitro and in silico approach.

Xanthine oxidase (XO) has been widely recognized as a pivotal enzyme in developing hyperuricemia, primarily contributing to the excessive production of uric acid during purine metabolism in the liver. One of the standard treatment approaches involves reducing uric acid levels by inhibiting XO activity. In this study, the leaf extract of Dolichandrone spathacea, traditionally used in folk medicine, was found to inhibit XO activity in the ethyl acetate and butanol fractions at a concentration of 100 µg/mL, their values were 78.57 ± 3.85 % (IC50 = 55.93 ± 5.73 µg/ml) and 69.43 ± 8.68 % (IC50 = 70.17 ± 7.98 µg/ml), respectively. The potential XO inhibitory components were isolated by bioactivity assays and the HR-ESI-MS and NMR spectra system. The main constituents of leaf extracts of Dolichandrone spathacea, six compounds, namely trans-4-methoxycinnamic acid (3), trans-3,4-dimethoxycinnamic acid (4), p-coumaric acid (5), martynoside (6), 6-O-(p-methoxy-E-cinnamoyl)-ajugol (7), and scolymoside (17), were identified as potent XO inhibitors with IC50 values ranging from 19.34 ± 1.63 µM to 64.50 ± 0.94 µM. The enzyme kinetics indicated that compounds 3-5, 7, and 17 displayed competitive inhibition like allopurinol, while compound 6 displayed a mixed-type inhibition. Computational studies corroborated these experimental results, highlighting the interactions between potential metabolites and XO enzyme. The hydrogen bonds played crucial roles in the binding interaction, especially, scolymoside (17) forms a hydrogen bond with Mos3004, exhibited the lowest binding energy (-18.3286 kcal/mol) corresponding to the lowest IC50 (19.34 ± 1.63 µM). Furthermore, nine compounds were isolated for the first time from this plant. In conclusion, Dolichandrone spathacea and its constituents possess the potential to modulate the xanthine oxidase enzyme involved in metabolism.

Evaluating the effects of surfactant types on the properties and stability of oil-in-water Rhodiola rosea nanoemulsion.

Different types and ratios of surfactant, co-surfactant, and oil phase, have a greater impact on nanoemulsion preparation. The presence of surfactants in the nanoemulsion can reduce surface tension and characteristic stability. In this study, four groups of oil-in-water (O/W) nanoemulsions (NEs) with different ratios of surfactant and co-surfactant, and two oils were formulated as carriers of Rhodiola rosea. The variable optimization was investigated and then indicated as optimization group A (Opt A) with the formula of 10% of transcutol, 16.63% of tween 80, Opt B with 10% of tween 80, 29.87% of span 80, Opt C with 28.42% of transcutol, 30% of labrasol, and Opt D with 30% of transcutol, 30% of tween 80. Labrafac and soybean oil were used as the oil phase. The optimized formula using the response surface method (RSM) by design expert software showed the ideal conditions with a higher desirability score. Desirability score are 0.72% (Opt A), 0.81% (Opt B), 0.76% (Opt C) and 0.98% (Opt D), the desirability rating close to 1 indicates a high possibility that the projected values would closely match the experimental results for the optimum formula. All of the optimized formulation were also checked for the characteristics of nanoemulsion including particle size, polydispersity index (PDI), zeta potential, viscosity, encapsulation efficiency, transmission electron microscope (TEM), antioxidant activity, skin irritation test and stability studies. Our study provides a promising combination of surfactant-co-surfactant and oil phases to produce a stable nanoemulsion that can be used in pharmaceuticals and cosmetics in the future.

A heterologous expression platform in Aspergillus nidulans for the elucidation of cryptic secondary metabolism biosynthetic gene clusters: discovery of the Aspergillus fumigatus sartorypyrone biosynthetic pathway.

Aspergillus fumigatus is a serious human pathogen causing life-threatening Aspergillosis in immunocompromised patients. Secondary metabolites (SMs) play an important role in pathogenesis, but the products of many SM biosynthetic gene clusters (BGCs) remain unknown. In this study, we have developed a heterologous expression platform in Aspergillus nidulans, using a newly created genetic dereplication strain, to express a previously unknown BGC from A. fumigatus and determine its products. The BGC produces sartorypyrones, and we have named it the spy BGC. Analysis of targeted gene deletions by HRESIMS, NMR, and microcrystal electron diffraction (MicroED) enabled us to identify 12 products from the spy BGC. Seven of the compounds have not been isolated previously. We also individually expressed the polyketide synthase (PKS) gene spyA and demonstrated that it produces the polyketide triacetic acid lactone (TAL), a potentially important biorenewable platform chemical. Our data have allowed us to propose a biosynthetic pathway for sartorypyrones and related natural products. This work highlights the potential of using the A. nidulans heterologous expression platform to uncover cryptic BGCs from A. fumigatus and other species, despite the complexity of their secondary metabolomes.

Anti-Aging Constituents from Pinus morrisonicola Leaves.

Pinus morrisonicola Hayata is a unique plant species found in Taiwan. Previous studies have identified its anti-hypertensive, anti-oxidative, and anti-inflammatory effects. In this study, a bioactivity-guided approach was employed to extract 20 compounds from the ethyl acetate fraction of the ethanol extract of Pinus morrisonicola Hayata's pine needles. The anti-aging effects of these compounds were investigated using HT-1080 cells. The structures of the purified compounds were confirmed through NMR and LC-MS analysis, revealing the presence of nine flavonoids, two lignans, one coumarin, one benzofuran, one phenylic acid, and six diterpenoids. Among them, PML18, PML19, and PML20 were identified as novel diterpene. Compounds 3, 4, and 5 exhibited remarkable inhibitory effects against MMP-2 and showed no significant cell toxicity at 25 µM. Although the purified compounds showed lower activity against Pro MMP-2 and Pro MMP-9 compared to the ethyl acetate fraction, we speculate that this is the result of synergistic effects.

Acrolein adducts and responding autoantibodies correlate with metabolic disturbance in Alzheimer's disease.

BACKGROUND: Alzheimer's disease (AD) is caused by many intertwining pathologies involving metabolic aberrations. Patients with metabolic syndrome (MetS) generally show hyperglycemia and dyslipidemia, which can lead to the formation of aldehydic adducts such as acrolein on peptides in the brain and blood. However, the pathogenesis from MetS to AD remains elusive. METHODS: An AD cell model expressing Swedish and Indiana amyloid precursor protein (APP-Swe/Ind) in neuro-2a cells and a 3xTg-AD mouse model were used. Human serum samples (142 control and 117 AD) and related clinical data were collected. Due to the involvement of MetS in AD, human samples were grouped into healthy control (HC), MetS-like, AD with normal metabolism (AD-N), and AD with metabolic disturbance (AD-M). APP, amyloid-beta (Aß), and acrolein adducts in the samples were analyzed using immunofluorescent microscopy, histochemistry, immunoprecipitation, immunoblotting, and/or ELISA. Synthetic Aß1-16 and Aß17-28 peptides were modified with acrolein in vitro and verified using LC-MS/MS. Native and acrolein-modified Aß peptides were used to measure the levels of specific autoantibodies IgG and IgM in the serum. The correlations and diagnostic power of potential biomarkers were evaluated. RESULTS: An increased level of acrolein adducts was detected in the AD model cells. Furthermore, acrolein adducts were observed on APP C-terminal fragments (APP-CTFs) containing Aß in 3xTg-AD mouse serum, brain lysates, and human serum. The level of acrolein adducts was correlated positively with fasting glucose and triglycerides and negatively with high-density lipoprotein-cholesterol, which correspond with MetS conditions. Among the four groups of human samples, the level of acrolein adducts was largely increased only in AD-M compared to all other groups. Notably, anti-acrolein-Aß autoantibodies, especially IgM, were largely reduced in AD-M compared to the MetS group, suggesting that the specific antibodies against acrolein adducts may be depleted during pathogenesis from MetS to AD. CONCLUSIONS: Metabolic disturbance may induce acrolein adduction, however, neutralized by responding autoantibodies. AD may be developed from MetS when these autoantibodies are depleted. Acrolein adducts and the responding autoantibodies may be potential biomarkers for not only diagnosis but also immunotherapy of AD, especially in complication with MetS.

Exploring the therapeutic efficacy of Chlorella pyrenoidosa peptides in ameliorating Alzheimer's disease.

Alzheimer's disease (AD) is one of the neurodegenerative disorders, the hallmarks of which include deposits of extracellular beta-amyloid (Aß) as well as intracellular tau neurofibrillary tangles (NFTs) tangles. With disease progression, neuronal apoptosis combined with cerebral atrophy occurs, leading to cognitive impairment and long-term memory loss. Recently, Chlorella species have been identified as a functional food and are being explored for the prevention of various diseases widely studied to prevent or treat many neurodegenerative diseases. Hence, we for the first time investigated the neuroprotective effects of Chlorella pyrenoidosa short-chain peptides (CPPs) i.e. <1 kDa, 1-3 kDa, 3-10 kDa, and >10 kDa on the in vitro and in vivo neuronal injury models. Our in vitro results showed that CPP with a molecular weight of 1-3 kDa and 3-10 kDa could elevate the survival rate of Aß1-42 or l-Glutamic acid-injured N2A cells. These treatments also inhibited Aß and tau NFTs in N2A cells and prevented progressive neuronal cellular damage by suppressing inflammatory cytokines such as PGE2, iNOS, IL-6, TNF-α, COX-2, IL-1ß, TGF-ß1, and NF-κB. Further, our in vivo Aß1-42-induced AD mice model demonstrated that 1-3 kDa or 3-10 kDa CPP could improve spatial cognition and learning memory. We also observed a decreased cell loss ratio in CA1-CA3 hippocampal regions. Taken together, our findings imply that CPPs may exert their anti-AD impact through anti-inflammatory, and anti-amyloid activities via reducing APP and tau NFT.

Cytotoxicity Effect of Constituents of Pinus taiwanensis Hayata Twigs on B16-F10 Melanoma Cells.

Pinus taiwanensis Hayata (Pinaceae) is an endemic plant in Taiwan. According to the Chinese Materia Medica Grand Dictionary, the Pinus species is mainly used to relieve pain, and eliminate pus and toxicity. In this study, nineteen compounds were isolated from the ethyl acetate layer of the ethanolic extract of P. taiwanensis Hayata twigs using bioassay-guided fractionation, and their anti-melanoma effects were investigated through a B16-F10 mouse melanoma cell model. The structures of the purified compounds were identified by 2D-NMR, MS, and IR, including 1 triterpenoid, 9 diterpenoids, 2 lignans, 4 phenolics, 1 phenylpropanoid, 1 flavonoid, and 1 steroid. Among them, compound 3 was found to be a new diterpene. Some of the compounds (2, 5, 6, 17, 18) showed moderate cytotoxicity effects. On the other hand, the anti-melanoma effect was no better than that from the original ethyl acetate layer. We presumed it resulted from the synergistic effect, although further experimentation needs to be performed.

Formulation, characteristics and anti-bacterial effects of Euphorbia hirta L. mouthwash.

Objective: Mouthwash is a liquid solution used to improve oral health and breath freshness as well as reduce oral bacteria. This study aims to formulate a Euphorbia hirta L. ethanol extract for mouthwash, evaluate the physical properties, and determine its anti-bacterial effects against Streptococcus mutans. Methods: Each mouthwash formula was created by utilising a solubilisation technique. Three mouthwash formulas were created from different concentrations of Euphorbia hirta L. ethanol extract (0.5%, 1%, and 2%), and referred to as F1, F2, and F3. The organoleptic properties, pH levels, specific gravity, viscosity, flow properties, stability, irritation level, contact time, and anti-bacterial effects were evaluated for each concentration. Result: The resulting formulas featured a distinctive smell of oleum menthae piperitae and had a sweet and spicy taste. These characteristics remained unaffected during storage. The acidity levels ranged from 4.59 to 6.0, the weight masses ranged from 0.9693 to 1.0710 g/ml, and the viscosity ranged from 1.50 to 3.00 cP. F3 concentration was non-irritating with mucus production at 11.325%, had lower contact time with a neutralisation of 57.14% of Streptococcus mutans colonies in 30 seconds, and showed stable anti-bacterial control against Streptococcus mutans (p = 0.000) within the first week of use. Conclusion: The study results offer the first proof of Euphorbia hirta L. used for improving human health. The formulation features the ideal physical and stability characteristics of mouthwash, and possesses anti-bacterial properties that can potentially combat Streptococcus mutans. Clinical use of the formulated mouthwash must be explored in future research.

New Trichothecenes Isolated from the Marine Algicolous Fungus Trichoderma brevicompactum.

Eight trichothecenes, including four new compounds 1-4 and four known entities 5-8, together with one known cyclonerane (9) were isolated from the solid-state fermentation of Trichoderma brevicompactum NTU439 isolated from the marine alga Mastophora rosea. The structures of 1-9 were determined by 1D/2D NMR (nuclear magnetic resonance), MS (mass spectrometry), and IR (infrared spectroscopy) spectroscopic data. All of the compounds were evaluated for cytotoxic activity against HCT-116, PC-3, and SK-Hep-1 cancer cells by the SRB assay, and compound 8 showed promising cytotoxic activity against all three cancer cell lines with the IC50 values of 3.3 ± 0.3, 5.3 ± 0.3, and 1.8 ± 0.8 µM, respectively. Compounds 1-2, 4-6, and 7-8 potently inhibited LPS-induced NO production, and compounds 5 and 8 showed markedly inhibited gelatinolysis of MMP-9 in S1 protein-stimulated THP-1 monocytes.

Serum levels of 4-hydroxynonenal adducts and responding autoantibodies correlate with the pathogenesis from hyperglycemia to Alzheimer's disease.

OBJECTIVE: Hyperglycemia leads to lipid peroxidation, producing 4-hydroxynonenal (HNE) adducts which correlate with the production of amyloid-beta (Aß), one of the hallmarks of Alzheimer's disease (AD). This study is to investigate the interactions of Aß, HNE adducts and responding autoantibodies during the pathogenesis from hyperglycemia to AD. METHODS: A total of 239 Taiwanese serum samples from a healthy control group and patients with hyperglycemia, and AD with and without hyperglycemia were analyzed. Aß was immunoprecipitated from randomly pooled serum in each group and immunoblotted. Synthetic Aß1-16 and Aß17-28 peptides were modified with HNE in vitro and verified with LC-MS/MS. The levels of Aß, HNE adducts, and autoantibody isotypes IgG and IgM against either native or HNE-modified Aß were determined with ELISA. The diagnostic power of potential biomarkers was evaluated. RESULTS: Increased fasting glucose and decreased high-density-lipoprotein cholesterol in AD groups indicated abnormal metabolism in the pathogenesis progression from hyperglycemia to AD. Indeed, serum Aß, HNE adducts and most of the autoantibodies recognizing either native or HNE-modified Aß were increased in the diseased groups. However, HNE adducts had better diagnostic performances than Aß for both hyperglycemia and AD. Additionally, HNE-Aß peptide levels were increased, and the responding autoantibodies (most notably IgM) were decreased in hyperglycemic AD group compared to the hyperglycemia only group, suggesting an immunity disturbance in the pathogenesis progression from hyperglycemia to AD. CONCLUSION: Hyperglycemia increases the level of HNE adducts which may be neutralized by responding autoantibodies. Depletion of these autoantibodies promotes AD-like pathogenesis. Thus, levels of a patient's HNE adducts and associated responding autoantibodies are potential biomarkers for AD with diabetes.

Bioactive compounds from an endophytic fungi Nigrospora aurantiaca.

BACKGROUND: Many groups of fungi live as an endophyte in plants. Both published and undiscovered bioactive compounds can be found in endophytic fungi. Various biological activities of bioactive compounds from endophytic fungi had been reported, including anti-inflammatory and anticancerous effects. The chemical investigation of biologically active compounds from endophytic fungi Melaleuca leucadendra Linn. have not yet been stated. RESULTS: One new compound, namely nigaurdiol (1), along with five known compounds, xyloketal K (2), bostrycin (3), deoxybostrycin (4), xylanthraquinone (5), and ergosterol (6), were isolated from the Melaleuca leucadendra Linn. associated fungal strain Nigrospora aurantiaca #TMU062. Their chemical structures were elucidated by spectroscopic data and compared with literature. All isolated compounds were evaluated for inhibitory effect of NO production in LPS-activated microglial BV-2 cells. CONCLUSIONS: Compound 6 exhibited considerable inhibitory effect on NO production with IC50 values of 7.2 ± 1.4 µM and the survival rate of the cells was 90.8 ± 6.7% at the concentration of 10 µM.

Quantification and Optimization of Ethanolic Extract Containing the Bioactive Flavonoids from Millettia pulchra Radix.

Millettia pulchra is traditionally used for treating diseases, including joint pain, fever, anemia, and allergies. It is also a potential resource of natural flavonoid derivatives, which represents major constituents of this plant. This study aimed to isolate the major compounds from M. pulchra radix, develop and validate the HPLC-PDA method to determine their contents, and optimize its extraction. Four major flavonoid derivatives (karanjin, lanceolatin B, 2",2"-dimethylpyrano-[5″,6″:7,8]-flavone, and pongamol) were isolated using silica gel column chromatography, crystallization techniques in large amounts with high purities (>95%). A simple, accurate high-performance liquid chromatography-photodiode array (HPLC-PDA) detection method has been developed and validated with significantly statistical impacts according to International Conference on Harmonization (ICH) guidelines. The Response Surface Methodology (RSM), Artificial Neural Network (ANN) models were employed to predictive performance and optimization of the extraction process. The optimized conditions for the extraction of major flavonoids were: extraction time (twice), solvent/material ratio (9.5), and ethanol concentration (72.5%). Our research suggests an effective method, which will be helpful for quality control in the pharmaceutical development of this species.

Constituents of Aquilaria sinensis Leaves Upregulate the Expression of Matrix Metalloproteases 2 and 9.

In this novel study, we isolated 28 compounds from the leaves of Aquilaria sinensis (Lour.) Gilg based on a bioassay-guided procedure and also discovered the possible matrix metalloprotease 2 (MMP-2) and 9 (MMP-9) modulatory effect of pheophorbide A (PA). To evaluate the regulatory activity on MMP-2 and MMP-9, the HT-1080 human fibrosarcoma cells were treated with various concentrations of extracted materials and isolated compounds. PA was extracted by methanol from the leaves of A. sinensis and separated from the fraction of the partitioned ethyl acetate layer. PA is believed to be an active component for MMP expression since it exhibited significant stimulation on MMP-2 and proMMP-9 activity. When treating with 50 µM of PA, the expression of MMP-2 and MMP-9 were increased 1.9-fold and 2.3-fold, respectively. PA also exhibited no cytotoxicity against HT-1080 cells when the cell viability was monitored. Furthermore, no significant MMP activity was observed when five PA analogues were evaluated. This study is the first to demonstrate that C-17 of PA is the deciding factor in determining the bioactivity of the compound. The MMP-2 and proMMP-9 modulatory activity of PA indicate its potential applications for reducing scar formation and comparative medical purposes.

Fast dereplication of xanthine oxidase-inhibiting compounds in alfalfa using comparative metabolomics.

Xanthine oxidase (XO) inhibition is a major strategy for preventing hyperuricemia and associated comorbidities, such as gout. Alfalfa extract has been demonstrated to possess XO-inhibiting activity; however, the elaborate conventional fraction-by-fraction analyses hindered the identification of the active components. In this study, we established a streamlined approach to rapidly screen, identify, and characterize XO-interacting compounds in alfalfa, by incorporating protein-subtraction, mass profiling, and molecular docking analysis. Crude extract was incubated with or without XO protein before UPLC-ESI-Q-TOF-MS/MS composition profiling. By dereplicating the component profile of XO-subtracted extract from that of untreated extract, the targets were rapidly narrowed down to twelve XO-interacting compounds, regarded as potential xanthine oxidase inhibitors (XOIs). Molecular docking analysis revealed that nine of these compounds, namely salicylic acid, tricin 7-O-glucuronopyranoside, chrysoeriol-7-glucoside, ferulic acid, apigenin 7-O-ß-glucuronopyranoside, apigenin, tricin, chrysoeriol, and liquiritigenin, exhibited high affinity with XO, and depicted the possible mechanisms of inhibition. In vitro bioassay further verified the XO inhibitory activities of selected compounds, among which apigenin, chrysoeriol and liquiritigenin were more potent XO inhibitors (XOIs), with IC50 of 0.25, 0.5 and 1 µM, respectively, compared to allopurinol (IC50 = 1.41 µM), the well-known XO-inhibiting drug. Together, the results demonstrated that alfalfa is a promising natural source for potent XOIs which might be applied for nutraceuticals development and that the approach used is applicable for efficient screening, identification, and mechanistic analyses of enzyme-inhibiting compounds from plant-based resources.

Triterpene glycosides and phenylpropane derivatives from Staurogyne concinnula possessing anti-angiogenic activity.

After anti-angiogenic activity screening, the potential n-butanol layer partitioned from the ethanol extract of Staurogyne concinnula was conducted. Further purification by Diaion HP20 column and preparative HPLC chromatography, four undescribed triterpenoid saponin derivatives, along with the known baptisiasaponin I, and four known phenylpropanoid glycosides were isolated and characterized from n-butanol layer. The structures of isolated compounds were elucidated by ESI-MS, 1D, and 2D MNR data. Biological evaluation revealed that baptisiasaponin I possessed significant anti-angiogenic effects (IC50 4.0 ± 0.2 µM). Further mechanism of action of baptisiasaponin I by inhibition of integrin/FAK/paxillin signaling pathway and its downstream effectors as MMP2 and MMP9 are also presented.

Determination of phenylalanine enantiomers in the plasma and urine of mammals and á´ -amino acid oxidase deficient rodents using two-dimensional high-performance liquid chromatography.

A two-dimensional (2D) HPLC system focusing on the determination of phenylalanine (Phe) enantiomers in mammalian physiological fluids has been developed. ᴅ-Phe is indicated to have potential values as a disease biomarker and therapeutic molecule in several neuronal and metabolic disorders, thus the regulation of ᴅ-Phe in mammals is a matter of interest. However, the precise determination of amino acid enantiomers is difficult in complex biological samples, and the development of an analytical method with practically acceptable sensitivity, selectivity and throughput is expected. In the present study, a 2D-HPLC system equipped with a reversed-phase column in the 1st dimension and an enantioselective column in the 2nd dimension has been designed, following the fluorescence derivatization of the target amino acid enantiomers with 4-fluoro-7-nitro-2,1,3-benzoxadiazole (NBD-F). The analytical method was validated using both plasma and urine samples, and successfully applied to human, rat and mouse fluids. Trace levels of ᴅ-Phe were determined in the plasma, and the %ᴅ values were around 0.1% for all species. In the urine, relatively large amounts of ᴅ-Phe were observed, and the %ᴅ values for humans, rats and mice were 3.99, 1.76 and 5.25%, respectively. The relationships between the enzymatic activity of ᴅ-amino acid oxidase (DAO) and the amounts of intrinsic ᴅ-Phe have also been clarified, and high ᴅ-Phe amounts were observed (around 0.3% in the plasma and around 50% in the urine) in the DAO deficient rats and mice.

Protective Evaluation of Compounds Extracted from Root of Rhodiola rosea L. against Methylglyoxal-Induced Toxicity in a Neuronal Cell Line.

Rhodiola rosea L. (R. rosea) is one of the most beneficial medicinal plants and it is studied as an adaptogen. This study aims to evaluate the neuroprotective activity of compounds extracted from the root of R. rosea against methylglyoxal (MG)-induced apoptosis in neuro-2A (N2A) cells. The root of R. rosea was extracted with ethanol and partitioned with water, ethyl acetate, and n-butanol fractions to evaluate acetylcholinesterase (AChE) inhibitory activity and neuroprotective activity. The ethyl acetate fraction exhibited the highest values of AChE inhibitory activity (49.2% ± 3%) and cell viability (50.7% ± 4.8%) for neuroprotection. The structure identification of the most potential fraction (ethyl acetate fraction) revealed 15 compounds, consisting of three tannins, five flavonoids, and seven phenolics by infrared spectroscopy, nuclear magnetic resonance, and mass spectroscopy. All compounds were evaluated for their neuroprotective activity. Salidroside had the most potential neuroprotective activity. Gallic acid and methyl gallate had potential cytotoxicity in N2A cells. This study showed that R. rosea might have potential neuroprotective activities.

Reduction of AHI1 in the serum of Taiwanese with probable Alzheimer's disease.

OBJECTIVE: The development of blood-based biomarkers for early diagnosis and treatment of Alzheimer's disease (AD) is desirable. In AD model mouse brain and neuronal cells, Abelson helper integration site-1 (AHI1) protein is reduced. AHI1 facilitates intracellular amyloid precursor protein (APP) translocation to inhibit amyloidogenic pathology of AD, and thus may be an AD biomarker. METHODS: This study was conducted among 32 AD patients and 54 healthy control (HC) subjects. AHI1-related protein levels from initially collected serum samples in each group were screened using Western blotting. The protein concentrations of AHI1 and amyloid-ß (Aß), peptide(s) derived from APP, from all serum samples were analyzed using ELISA. RESULTS: In AD serum, AHI1 and a large truncated C-terminal APP fragment were significantly reduced. The average concentrations of serum AHI1 and Aß in AD were significantly lower than those in HC. Notably, AHI1 concentration in HC serum was decreased in an age-dependent manner, while it was consistently low in AD serum and had no correlation with Aß or mini-mental state examination score. The receiver operating characteristic analysis on all subjects demonstrated an area under curve (AUC) value of 0.7 for AHI1 on AD diagnosis, while the AUC increased to 0.82 on the subjects younger than 77 years old, suggesting a good diagnostic performance of serum AHI1 for AD especially at relatively young age. CONCLUSION: An early event of AHI1 reduction in the body of AD patients was observed. Serum AHI1 may be valuable for early diagnosis of AD.

Efficient identification of fungal antimicrobial principles by tandem MS and NMR database.

The continuous re-isolation of the known and non-applicable compounds that is time-consuming and wasting resources is still a critical problem in the discovery of bioactive entities from natural resources. To efficiently address the problem, high performance liquid chromatography-diode array detector-microfractionation (HPLC-DAD-microfractionation) guided by disk agar diffusion assay was developed, and the active compounds were further identified using the tandem mass spectrometry (MS/MS)-based molecular networking. Of 150 fungal strains screened, the methanolic extracts of Phoma herbarum PPM7487, Cryptosporiopsis ericae PPM7405, and Albifimbria verrucaria PPM945 exhibited potent antimicrobial activity against Candida albicans SC5314 and Cryptococcus neoformans H99 in the preliminary agar diffusion assay. The concept of OSMAC (one strain many compounds) was employed in the fungal cultures in order to enrich the diversity of the 2nd metabolites in this study. HPLC coupled with off-line bioactivity-directed profiling of the extracts enabled a precise localization of the compounds responsible for the conspicuous antimicrobial activity. The purified active compounds were identified based mainly on MS/MS database, and further supported by 13C nuclear magnetic resonance (NMR) spectral data compared to the literatures. In addition to nineteen known compounds, a new trichothecene derivative 1, namely trichoverrin D, was isolated and identified through this protocol. The antifungal activities of all the pure isolates were evaluated, and the structure activity relationships were also inferred. This report has demonstrated the combination of HPLC microfractination and MS/MS coupled by NMR spectral dereplication for speeding up the antimicrobial natural products discovery process.

Characterizing Tyrosinase Modulators from the Roots of Angelica keiskei Using Tyrosinase Inhibition Assay and UPLC-MS/MS as the Combinatorial Novel Approach.

In this study, an in vitro tyrosinase inhibition assay in combination with ultra performance liquid chromatography-orbitrap mass spectrometry (UPLC-orbitrap-MS) was developed for the rapid screening and identification of tyrosinase modulators from roots of Angelica keiskei. Of the 15 candidates considered, nine chalcones, xanthoangelols (1), B (2), D (3), E (4), G (5), H (6), 4-hydroxyderricin (7), xanthokeismin B (8) and (2E)-1-[4-hydroxy-2-(2-hydroxy-2-propanyl)-2,3-dihydro-1-benzofuran-7-yl]-3-(4-hydroxyphenyl)-2-propen-1-one (9), five coumarins, umbelliferone (10), selinidin (11), isopimpinellin (12), phellopterin (13) and xanthyletin (14), and one other compound, ashitabaol A (15), were distinguished between the test samples and the controls with statistical significance, and the structure of each compound was determined by comparing with in-house standards and the literature. Among these, six compounds, xanthoangelol (1), xanthoangelol D (3), xanthoangelol H (6), 4-hydroxyderricin (7), laserpitin (16) and isolaserpitin (17), were isolated from roots of A. keiskei. Of the compounds isolated, compounds 1, 7 and 16 were subjected to tyrosinase inhibitory assay, and the IC50 values were 15.87 ± 1.21, 60.14 ± 2.29 and >100 µM, respectively. The present study indicated that the combination of in vitro tyrosinase inhibition assay coupled with UPLC-MS/MS could be widely applied to the rapid screening of active substances from various natural resources.