Oxindole and Benzoxazinone Alkaloids from the Seeds of Persea americana (Avocado) and Their SIRT1 Stimulatory Activity.

Persea americana Mill. (Lauraceae), commonly known as avocado, is a well-known food because of its nutrition and health benefits. The seeds of avocado are major byproducts, and thus their phytochemicals and bioactivities have been of interest for study. The chemical components of avocado seeds were investigated by using UPLC-qTOF-MS/MS-based molecular networking, resulting in the isolation of seven new oxindole alkaloids (1-7) and two new benzoxazinone alkaloids (8 and 9). The chemical structures of the isolated compounds were identified by the analysis of NMR data in combination with computational approaches, including NMR and ECD calculations. Bioactivities of the isolated compounds toward silent information regulation 2 homologue-1 (SIRT1) in HEK293 cells were assessed. The results showed that compound 1 had the most potent effect on SIRT1 activation with an elevated NAD+/NADH ratio with potential for further investigation as an anti-aging agent.

A molecular networking-based isolation of gardneria alkaloids from Gardneria distincta and their anti-inflammatory activity.

Gardneria distincta P. T. Li is traditionally applied as a herbal medicine for treatment various ailments, and is mainly distributed in Southwestern China. Under the guided separation of MS/MS-based molecular networking, eight undescribed oxindole alkaloids, gardistines A-H, as well as 17 known alkaloids were discovered from the whole parts of Gardneria distincta. Structural elucidation of these undescribed alkaloids was performed by various spectroscopic methods. Gardistine A is a rare oxindole gardneria alkaloid bearing an ester carbonyl group attached to C-18, which is the second reported alkaloid of oxindole gardneria-type. All of the identified monoterpene indole alkaloids were investigated for their anti-inflammatory activity in LPS-induced RAW 264.7 cells. Gardistines A-B and akuammidine demonstrated significant inhibitory effects on the expressions of nitric oxide, tumor necrosis factor alpha, and interleukin-6 at 20 µM.

Profiles of Metabolic Genes in Uncaria rhynchophylla and Characterization of the Critical Enzyme Involved in the Biosynthesis of Bioactive Compounds-(iso)Rhynchophylline.

Rhynchophylline (RIN) and isorhynchophylline (IRN), two of the representative types of indole alkaloids, showed the unique spiroindole structures produced in Uncaria rhynchophylla. As the bioactive constituent of U. rhynchophylla, IRN has recently drawn extensive attention toward antihypertensive and neuroprotective activities. Despite their medicinal importance and unique chemical structure, the biosynthetic pathways of plant spiroindole alkaloids are still largely unknown. In this study, we used U. rhynchophylla, extensively used in traditional Chinese medicine (TCM), a widely cultivated plant of the Uncaria genus, to investigate the biosynthetic genes and characterize the functional enzymes in the spiroindole alkaloids. We aim to use the transcriptome platform to analyse the tissue-specific gene expression in spiroindole alkaloids-producing tissues, including root, bud, stem bark and leaf. The critical genes involved in the biosynthesis of precursors and scaffold formation of spiroindole alkaloids were discovered and characterized. The datasets from this work provide an essential resource for further investigating metabolic pathways in U. rhynchophylla and facilitate novel functional enzyme characterization and a good biopharming approach to spiroindole alkaloids.

A Sensitive LC-MS/MS Method for the Simultaneous Determination of Two Thia-Analogous Indirubin N-Glycosides and Indirubin-3'-Monoxime in Plasma and Cell Culture Medium.

Indirubin was identified as an active component of Danggui Longhui Wan, an herbal mixture used in traditional Chinese medicine, and showed anticancer activity in clinical trials in patients with chronic leukemia. Investigations on the mechanisms of antitumor action of indirubins have mainly focused on the indirubin derivative indirubin-3'-monoxime (I3M). Meanwhile, antiproliferative and cytotoxic properties on cancer cells have also been demonstrated for several synthetic indirubin N-glycosides. In the present study, we demonstrate cytotoxic activity of the thia-analogous indirubin N-glycosides KD87 (3-[3'-oxo-benzo[b]thiophen-2'-(Z)-ylidene]-1-(ß-d-glucopyranosyl)-oxindole) and KD85 (3-[3'-oxo-benzo[b]thiophen-2'-(Z)-ylidene]-1-(ß-d-mannopyranosyl)-oxindole) against melanoma and squamous cell carcinoma cells as well as lung cancer and glioblastoma cells. The advanced state of preclinical studies on the effects of indirubins conducted to date underscores the need for pharmacokinetic data from cellular, animal, and human studies for which reliable quantification is required. Therefore, a sensitive liquid chromatography-tandem mass spectrometric (LC-MS/MS) method was developed and validated for the simultaneous measurement of KD87, KD85, and I3M in plasma and cell culture medium. Experimental conditions for sample preparation were optimized for human plasma protein precipitation and liquid-liquid extraction from plasma and cell culture medium. The methods were successfully validated in accordance with the U.S. Food and Drug Administration Bioanalytical Method Validation and evaluated for selectivity, sensitivity, matrix effect, recovery, carryover, calibration curve linearity, accuracy, precision, and stability. The applicability of the methods was demonstrated by the determination of KD87 in mouse plasma after prior intraperitoneal administration to mice.

Kratom (Mitragyna speciosa) Validation: Quantitative Analysis of Indole and Oxindole Alkaloids Reveals Chemotypes of Plants and Products.

Many consumers are turning to kratom (Mitragyna speciosa) to self-manage pain and opioid addiction. In the United States, an array of capsules, powders, and loose-leaf kratom products are readily available. Additionally, several online sites supply live kratom plants. A prerequisite to establishing quality control and quality assurance standards for the kratom industry, or understanding how alkaloid levels effect clinical outcomes, is the identification and quantitation of major and minor alkaloid constituents within available products and preparations. To this end, an ultra-high performance liquid chromatography-high resolution mass spectrometry method was developed for the analysis of 8 indole alkaloids (7-hydroxymitragynine, ajmalicine, paynantheine, mitragynine, speciogynine, isopaynantheine, speciociliatine, and mitraciliatine) and 6 oxindole alkaloids (isomitraphylline, isospeciofoleine, speciofoline, corynoxine A, corynoxeine, and rhynchophylline) in US-grown kratom plants and commercial products. These commercial products shared a qualitatively similar alkaloid profile, with 12 - 13 detected alkaloids and high levels of the indole alkaloid mitragynine (13.9 ± 1.1 - 270 ± 24 mg/g). The levels of the other major alkaloids (paynantheine, speciociliatine, speciogynine, mitraciliatine, and isopaynantheine) and the minor alkaloids varied in concentration from product to product. The alkaloid profile of US-grown M. speciosa "Rifat" showed high levels of the indole alkaloid speciogynine (7.94 ± 0.83 - 11.55 ± 0.18 mg/g) and quantifiable levels of isomitraphylline (0.943 ± 0.033 - 1.47 ± 0.18 mg/g). Notably, the alkaloid profile of a US-grown M. speciosa seedling was comparable to the commercial products with a high level of mitragynine (15.01 ± 0.20 mg/g). This work suggests that there are several M. speciosa chemotypes.

In vivo screening and toxicity studies of indolinone incorporated thiosemicarbazone, thiazole and piperidinosulfonyl moieties as anticonvulsant agents.

Based on the biological importance of the thiazole nucleus, we decided to prepare and evaluate the biological activity of some new isatin derivatives containing thiazole moiety. The 5-(piperidin-1-ylsulfonyl)indoline-2,3-dione (1) was prepared and used as a starting material in the synthesis of many isatin derivatives for anticonvulsant evaluation. All the newly synthesized thiazlidino/thiosemicarbazide-indolin-2-one derivatives screened in vivo for their anticonvulsant activity against pentylenetetrazole-induced convulsions in mice. The results were compared with phenobarbitone sodium as a standard anticonvulsant drug. Most of the tested compounds exhibited anticonvulsant activity with relative potency ranging from 0.02 to 0.2 in comparison to standard drug phenobarbitone. The most active compounds 3, 6a, 6c and 8, were exposed to further investigations in rats to evaluate the effect of most active derivatives on the haematological, liver, kidney functions as well as histopathological studies of the liver and kidney tissues. Finally, the most potent compounds 3, 6a, 6c and 8 observed good toxic properties for both liver and kidney function with mild variability changes on RBCs, WBCs, Platelets, Hb, AST, ALT, and creatinine level, as well as kidney and liver tissue and these good results obtained rather than used low dose from phenobarbitone.

Oxindole and its derivatives: A review on recent progress in biological activities.

Oxindole has been shown to be a pharmacologically advantageous scaffold having many biological properties that are relevant to medicinal chemistry. The simplicity and widespread occurrence of this scaffold in plant-based alkaloids have further reinforced oxindole's merit in the domain of novel drug discovery. First extracted from Uncaria tomentosa, commonly the known as cat claw's plant which was found abundantly in the Amazon rainforest, molecules with the oxindole moiety have been shown to be common in a wide variety of compounds extracted from plant sources. The role of oxindole as a chemical scaffold for fabricating and designing biological drugs agents can be ascribed to its ability to be modified by a number of chemical groups to generate novel biological functions. This review is aimed at providing a description of the general chemistry based on existing corresponding structure-activity relationships (SARs) and compile all recent developmentary studies on oxindole-derived compounds as a successful pharmaceutical agent. A substantial group of oxindole derivatives are chiefly being tested as anticancer agents, however, a several oxindole derivatives have been shown to possesses antimicrobial, α-glucosidase inhibitory, antiviral, antileishmanial, antitubercular, antioxidative, tyrosinase inhibitory, PAK4 inhibitory, antirheumatoid arthritis and intraocular pressure reducing activities, to name a few. In this review we show the potential value of developing newer oxindole derivatives with an improved range of pharmacological implications as well as identifying drugs possessing oxindole core, that are showing and serving increased efficacy in clinical practice.

[Antioxidant mechanism of gastrodin combined with isorhynchophylline in inhibiting MPP~+-induced apoptosis of PC12 cells].

Gastrodiae Rhizoma-Uncariae Ramulus cum Uncis is the most frequently used herbal pair in the treatment of Parkinson's disease(PD). Gastrodin and isorhynchophylline are important components of Gastrodiae Rhizoma-Uncariae Ramulus cum Uncis herb pair with anti-Parkinson mechanism. This study aimed to investigate the effect of gastrodin combined with isorhynchophylline on 1-methyl-4-phenylpyridinium(MPP~+)-induced apoptosis of PC12 cells and their antioxidant mechanism. The leakage of lactate dehydrogenase(LDH) from cells to media was analyzed by spectrophotometry. Apoptotic cells were labeled with Annexin V-fluorescein isothiocyanate(FITC) and propidium iodide(PI) and analyzed by flow cytometry. The cell cycle was analyzed using propidium iodide(PI) staining. Lipid peroxidation(LPO) level was analyzed by spectrophotometry. The mRNA expression of caspase-3 was examined by Real-time RT-PCR. The protein expressions of heme oxygenase 1(HO-1) and NADPH: quinoneoxidore-ductase 1(NQO-1) were determined by Western blot. Gastrodin combined with isorhynchophylline reduced the percentage of Annexin V-positive cells and cell cycle arrest in MPP~+-induced PC12 cells. Gastrodin combined with isorhynchophylline down-regulated the mRNA expression of caspase-3, up-regulated the protein expressions of HO-1 and NQO-1, and reduced LPO content in MPP~+-induced PC12 cells. PD98059, LY294002 or LiCl could partially reverse these changes pretreated with gastrodin combined with isorhynchophylline, suggesting that gastrodin combined with isorhynchophylline inhibited MPP~+-induced apoptosis of PC12 cells and oxidative stress through ERK1/2 and PI3 K/GSK-3ß signal pathways. Our experiments showed that gastrodin combined with isorhynchophylline could down-re-gulate the mRNA expression of caspase-3 and up-regulate the protein expressions of HO-1 and NQO-1, so as to reduce oxidative stress and inhibit apoptosis.

Pharmacokinetics of Eleven Kratom Alkaloids Following an Oral Dose of Either Traditional or Commercial Kratom Products in Rats.

Kratom, Mitragyna speciosa Korth., is being widely consumed in the United States for pain management and the reduction of opioid withdrawal symptoms. The central nervous system (CNS) active alkaloids of kratom, including mitragynine, 7-hydroxymitragynine, and numerous additional compounds, are believed to derive their effects through opioid receptor activity. There is no literature describing the systemic exposure of many of these alkaloids after the consumption of kratom. Therefore, we have developed and validated a bioanalytical method for the simultaneous quantitation of 11 kratom alkaloids (mitragynine, 7-hydroxymitragynine, corynantheidine, speciogynine, speciociliatine, paynantheine, corynoxine, corynoxine-B, mitraphylline, ajmalicine, and isospeciofoline) in rat plasma. The validated method was used to analyze oral pharmacokinetic study samples of lyophilized kratom tea (LKT) and a marketed product, OPMS liquid shot, in rats. Among the 11 alkaloids, only mitragynine, 7-hydroxymitragynine, speciociliatine, and corynantheidine showed systemic exposure 8 h postdose, and the dose-normalized systemic exposure of these four alkaloids was higher (1.6-2.4-fold) following the administration of the commercial OPMS liquid. Paynantheine and speciogynine levels were quantifiable up to 1 h postdose, whereas none of the other alkaloids were detected. In summary, the method was successfully applied to quantify the exposure of individual kratom alkaloids after an oral dose of traditional or commercial products. This information will contribute to understanding the role of each alkaloid in the overall pharmacology of kratom and elucidating the pharmacokinetic differences between traditional and commercial kratom products.

Systematic elucidation of the pharmacological mechanisms of Rhynchophylline for treating epilepsy via network pharmacology.

ABSTACT: BACKGROUND: Epilepsy, one of the most common neurological disorders, affects over 70 million people worldwide. Rhynchophylline displays a wide variety of pharmacologic actives. However, the pharmacologic effects of rhynchophylline and its mechanisms against epilepsy have not been systematically elucidated. METHODS: The oral bioavailability and druglikeness of rhynchophylline were evaluated using the Traditional Chinese Medicine Systems Pharmacology Database. Rhynchophylline target genes to treat epilepsy were identified using PharmMapper, SwissTargetPrediction and DrugBank databases integration. Protein-protein interaction analysis was carried out by utilizing the GeneMANIA database. WebGestalt was employed to perform Gene ontology and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analyses. The drug-disease-target-Gene Ontology-pathway network was constructed using Cytoscape. RESULTS: The oral bioavailability and druglikeness of rhynchophylline were calculated to be 41.82% and 0.57, respectively. A total of 20 rhynchophylline target genes related to epilepsy were chosen. Among the 20 genes and their interacting genes, 54.00% shared protein domains and 16.61% displayed co-expression characteristics. Gene ontology, Kyoto Encyclopedia of Genes and Genomes and network analyses illustrate that these targets were significantly enriched in regulation of sensory perception, morphine addiction, neuroactive ligand-receptor interaction and other pathways or biological processes. CONCLUSION: In short, rhynchophylline targets multiple genes or proteins, biological processes and pathways. It shapes a multiple-layer network that exerts systematic pharmacologic activities on epilepsy.

Kinetic Resolution of Aziridines Enabled by N-Heterocyclic Carbene/Copper Cooperative Catalysis: Carbene Dose-Controlled Chemo-Switchability.

Catalytic kinetic resolution (KR) and dynamic kinetic asymmetric transformation (DyKAT) are alternative and complementary avenues to access chiral stereoisomers of both starting materials and reaction products. The development of highly efficient chiral catalytic systems for kinetically controlled processes has therefore been one of the linchpins in asymmetric synthesis. N-heterocyclic carbene (NHC)/copper cooperative catalysis has enabled highly efficient KR and DyKAT of racemic N-tosylaziridines by [3+3] annulation with isatin-derived enals, leading to highly enantioenriched N-tosylaziridine derivatives (up to >99 % ee) and a large library of spirooxindole derivatives with high structural diversity and stereoselectivity (up to >95:5 d.r., >99 % ee). Mechanistic studies suggest that the NHC can bind reversibly to the copper catalyst without compromising its catalytic activity and regulate the catalytic activity of the copper complex to switch the chemoselection between KR and DyKAT.

Protection by rhynchophylline against MPTP/MPP+-induced neurotoxicity via regulating PI3K/Akt pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Isolated from Uncaria rhynchophylla (U. rhynchophylla), rhynchophylline (Rhy) has been applied for treating diseases related to central nervous system such as Parkinson's disease. Nevertheless, the molecular mechanism of the neuroprotective effect has not been well interpreted. AIM OF THE STUDY: To investigate the effects of Rhy on MPTP/MPP + -induced neurotoxicity in C57BL/6 mice or PC12 cells and study the mechanisms involved. MATERIALS AND METHODS: The neuroprotective effect of Rhy on MPTP-induced neurotoxicity was evaluated by spontaneous motor activity test, as well as a test of rota-rod on a rat model of Parkinson's disease. The numbers of TH-positive neurons in the substantia nigra pars compacta (SNpc) was assessed by immunohistological. CCK-8, lactate dehydrogenase (LDH), reactive oxygen species (ROS), the concentration of intracellular calcium ([Ca2+]i) and flow cytometry analysis were performed to evaluate the pharmacological property of Rhy on 1-methyl-4-phenylpyridinium (MPP+) induced neurotoxicity in PC12 cells. Besides, LY294002, a PI3K inhibitor was employed to determine the underlying molecular signaling pathway revealing the effect of Rhy by western-blot analysis. RESULTS: The results showed that Rhy exhibited a protective effect against the MPTP-induced decrease in tyrosine hydroxylase (TH)-positive fibers in the substantia nigra at 30 mg/kg, demonstrated by the immunohistological and behavioral outcomes. Furthermore, it has been indicated that cell viability was improved and the MPP+-induced apoptosis was inhibited after the treatment of Rhy at 20 µM, which were severally analyzed by the CCK-8 and the Annexin V/propidium iodide staining method. In addition, Rhy treatment attenuated MPP+-induced up-regulation of LDH, ([Ca2+]i), and the levels of ROS. Besides, it can be revealed from the Western blot assay that LY294002, as a selective Phosphatidylinositol 3-Kinase (PI3K) inhibitor, effectively inhibited the Akt phosphorylation caused by Rhy, which suggested that Rhy showed its protective property through the activated the PI3K/Akt signaling pathway. Moreover, the Rhy-induced decreases of Bax and caspase-3 as the proapoptotic markers and the increase of Bcl-2 as the antiapoptotic marker, were blocked by LY294002 in the MPP+-treated PC12 cells. CONCLUSIONS: Rhy exerts a neuroprotective effect is partly mediated by activating the PI3K/Akt signaling pathway.

Recent research progress of Uncaria spp. based on alkaloids: phytochemistry, pharmacology and structural chemistry.

Medicinal plants are well-known in affording clinically useful agents, with rich medicinal values by combining with disease targets through various mechanisms. Plant secondary metabolites as lead compounds lay the foundation for the discovery and development of new drugs in disease treatment. Genus Uncaria from Rubiaceae family is a significant plant source of active alkaloids, with anti-hypertensive, sedative, anti-Alzheimer's disease, anti-drug addiction and anti-inflammatory effects. This review summarizes and discuss the research progress of Uncaria based on alkaloids in the past 15 years, mainly in the past 5 years, including biosynthesis, phytochemistry, pharmacology and structural chemistry. Among, focusing on representative compounds rhynchophylline and isorhynchophylline, the pharmacological activities surrounding the central nervous system and cardiovascular system are described in detail. On the basis of case studies, this article provides a brief overview of the synthesis and analogues of representative compounds types. In summary, this review provides an early basis for further searching for new targets and activities, discussing the mechanisms of pharmacological activity and studying the structure-activity relationships of active molecules.

An oxindole efflux inhibitor potentiates azoles and impairs virulence in the fungal pathogen Candida auris.

Candida auris is an emerging fungal pathogen that exhibits resistance to multiple drugs, including the most commonly prescribed antifungal, fluconazole. Here, we use a combinatorial screening approach to identify a bis-benzodioxolylindolinone (azoffluxin) that synergizes with fluconazole against C. auris. Azoffluxin enhances fluconazole activity through the inhibition of efflux pump Cdr1, thus increasing intracellular fluconazole levels. This activity is conserved across most C. auris clades, with the exception of clade III. Azoffluxin also inhibits efflux in highly azole-resistant strains of Candida albicans, another human fungal pathogen, increasing their susceptibility to fluconazole. Furthermore, azoffluxin enhances fluconazole activity in mice infected with C. auris, reducing fungal burden. Our findings suggest that pharmacologically targeting Cdr1 in combination with azoles may be an effective strategy to control infection caused by azole-resistant isolates of C. auris.

Deciphering the Mechanism of the Anti-Hypertensive Effect of Isorhynchophylline by Targeting Neurotransmitters Metabolism of Hypothalamus in Spontaneously Hypertensive Rats.

Essential hypertension is a major risk factor for cardiovascular disease that can lead eventually to structural and functional alterations in the brain. Accumulating evidence has suggested that the increased activities in renin-angiotensin system and sympathetic nerve participated in the pathogenesis of hypertension that is related to the imbalance between neurotransmitters. The potential role in essential hypertension arising from alterations of neurotransmitters in the central nervous system remains understudied. Isorhynchophylline is a major oxindole alkaloid extracted from Uncaria rhynchophylla, which has been widely used for treating hypertension and neurodegenerative diseases. Whether isorhynchophylline acts on neurotransmitters to lower blood pressure has been hypothesized but rarely demonstrated unequivocally. Here, we studied the metabolic neurotransmitter profiles in the hypothalamus using a targeted metabolomic approach in spontaneously hypertensive rats after isorhynchophylline intervention. Our study demonstrated that isorhynchophylline exhibited a strong anti-hypertensive effect in spontaneously hypertensive rats by improving the neurotransmitter imbalance in the hypothalamus and inhibiting the overactivation of the renin-angiotensin system and sympathetic nerve system. Overall, this study played an essential role in enhancing our understanding of the mechanism of isorhynchophylline in essential hypertension and in providing theoretical evidence for future research and clinical application.

A Phytochemical and Ethnopharmacological Recapitulation on Hamelia patens.

The world health organization reports that 80% of the population living within the developing countries depends basically on traditional medicine for his or her primary health care. Quite half the entire world's population still depends entirely on plants for medicines, and plants provides the active ingredients to the most traditional medical products. Hamelia patens that belong to the family Rubiaceae, is mainly found in tropical and sub-tropical areas. It is used in folk medicine against a wide range of diseases such as athlete's foot, skin problems, insect sting, psychiatric disorder, rheumatism, headache, asthma, dysentery, menses, ovarian and uterine disorders. Hamelia consists of an important bioactive constituent such as oxindole alkaloids, flavonoids, and phenolic content. Due to the presence of chlorogenic acid and epicatechin constituent in the methanolic extract of Hamelia patens, there is a noticeable anti-hyperglycemic activity as well as it possesses antioxidant activity. Acute and sub-acute toxicity study has been performed on Hamelia patens which shows that it is safe and can be used for humans. Against fungi and bacteria, the ethanol leaf extract of Hamelia has a maximum inhibitory effect. The plant has various therapeutic effects like anti-inflammatory, analgesic, anti-diarrheal, anthelmintic, antidepressant, hepatoprotective, antiurolithiatic, diuretic, wound healing, and many others. In this article, we have discussed chemical constituent, pharmacological activity and traditional use of Hamelia patens.

Synthesis, Evaluation and Proposed Binding Pose of Substituted Spiro-Oxindole Dihydroquinazolinones as IRAP Inhibitors.

Insulin-regulated aminopeptidase (IRAP) is a new potential macromolecular target for drugs aimed for treatment of cognitive disorders. Inhibition of IRAP by angiotensin IV (Ang IV) improves the memory and learning in rats. The majority of the known IRAP inhibitors are peptidic in character and suffer from poor pharmacokinetic properties. Herein, we present a series of small non-peptide IRAP inhibitors derived from a spiro-oxindole dihydroquinazolinone screening hit (pIC50 5.8). The compounds were synthesized either by a simple microwave (MW)-promoted three-component reaction, or by a two-step one-pot procedure. For decoration of the oxindole ring system, rapid MW-assisted Suzuki-Miyaura cross-couplings (1 min) were performed. A small improvement of potency (pIC50 6.6 for the most potent compound) and an increased solubility could be achieved. As deduced from computational modelling and MD simulations it is proposed that the S-configuration of the spiro-oxindole dihydroquinazolinones accounts for the inhibition of IRAP.

Rhynchophylline Loaded-mPEG-PLGA Nanoparticles Coated with Tween-80 for Preliminary Study in Alzheimer's Disease.

PURPOSE: Alzheimer's disease (AD) is a growing concern in the modern society. The current drugs approved by FDA are not very promising. Rhynchophylline (RIN) is a major active tetracyclic oxindole alkaloid stem from traditional Chinese medicine uncaria species, which has potential activities beneficial for the treatment of AD. However, the application of rhynchophylline for AD treatment is restricted by the low water solubility, low concentration in brain tissue and low bioavailability. And there is no study of brain-targeting therapy with RIN. In this work, we prepared rhynchophylline loaded methoxy poly (ethylene glycol)-poly (dl-lactide-co-glycolic acid) (mPEG-PLGA) nanoparticles (NPS-RIN), which coupled with Tween 80 (T80) further for brain targeting delivery (T80-NPS-RIN). METHODS: Preparation and characterization of T80-NPS-RIN were followed by the detection of transportation across the blood-brain barrier (BBB) model in vitro, biodistribution and neuroprotective effects of nanoparticles. RESULTS: The results indicated T80-NPS-RIN could usefully assist RIN to pass through the BBB to the brain. T80-NPS-RIN treatment regulated the activity of neurons in vitro. CONCLUSION: The presented data confirmed that rhynchophylline encapsulated mPEG-PLGA nanoparticles coated with Tween 80 could across through the BBB and exhibited efficient neuroprotective effects. The T80-NPS-RIN nanoparticles have a chance to be an alternative drug to the therapy of AD.

Identification of metabolites and potential biomarkers of kratom in urine.

Mitragyna speciosa (kratom) is a drug that is increasingly used recreationally and "therapeutically", in the absence of medical supervision. The drug has been associated with a growing number of fatalities, and although its medicinal properties as an atypical opioid require further study, there are legitimate concerns regarding its unregulated use. Mitragynine is the most widely reported alkaloid within the plant, although more than forty other alkaloids have been identified. 7-Hydroxymitragynine is reported to have greater abuse liability due to its increased potency relative to mitragynine. In this report, biomarkers for mitragynine were investigated using liquid chromatography-quadrupole/time of flight mass spectrometry (LC-Q/TOF-MS). Speciociliatine and speciogynine were identified as alternative biomarkers, often exceeding the concentration of mitragynine in unhydrolyzed urine. 9-O-Demethylmitragynine and 7-hydroxymitragynine were identified in unhydrolyzed urine in 75% and 63% of the cases. Deconjugation of phase II metabolites using chemical hydrolysis was not suitable due to degradation of the Mitragyna alkaloids. Enzymatic hydrolysis was evaluated using three traditional glucuronidases, four sulfatases and four recombinant enzymes. Although enzymatic hydrolysis increased the concentration of 16-carboxymitragynine, it had nominal benefit for other metabolites. Deconjugation of urine was not necessary due to the abundance of parent drug (mitragynine), its diastereoisomers (speciociliatine and speciogynine) or metabolites (9-O-demethylmitragynine and 7-hydroxymitragynine).

Temperature and pH-Dependent Stability of Mitragyna Alkaloids.

Mitragynine (MG) is the principal psychoactive alkaloid in kratom. The drug produces a variety of dose-dependent effects that appeal to recreational drug users and individuals seeking therapeutic benefits in the absence of medical supervision. In light of documented intoxications, hospitalizations and fatalities, MG and other alkaloids from Mitragyna speciosa are of growing importance to the forensic toxicology community. However, the chemical stability of these compounds has not been thoroughly described. In this report, the stability of MG, 7-hydroxymitragynine (MG-OH), speciociliatine (SC), speciogynine (SG) and paynantheine (PY) are investigated. Short-term stability of the Mitragyna alkaloids was determined over a range of pH (2-10) and temperature (4-80°C) over 8 hours. Liquid chromatography--quadrupole/time-of-flight mass spectrometry was used to estimate half-lives and identify degradation products where possible. The stability of MG and other alkaloids was highly dependent on pH and temperature. All of the Mitragyna alkaloids studied were acid labile. Under alkaline conditions, MG undergoes chemical hydrolysis of the methyl ester to produce 16-carboxymitragynine. MG-OH was the most unstable alkaloid studied, with significant drug loss at 8 hours experienced at temperatures of 40°C and above. No significant drug losses were observed for MG in aqueous solution (pH 2-10) at 4, 20 or 40°C. Diastereoisomers of MG (SC and SG) demonstrated even greater stability. These findings are discussed within the context of the identification of Mitragyna alkaloids in toxicological specimens.