A fatal case of desvenlafaxine and paroxetine poisoning.

Desvenlafaxine (O-desmethylvenlafaxine) and paroxetine are antidepressants that inhibit serotonin reuptake. Despite their relatively safe profiles, several serious side effects, including serotonin syndrome, bleeding, mania, and high blood pressure, are observed. We report the confirmation of the death of a 41-year-old female, with an overdose of desvenlafaxine and paroxetine suspected as the main cause of death. To quantify the level of desvenlafaxine and paroxetine in whole blood and urine, solid phase extraction combined with liquid chromatography-tandem mass spectrometry was developed and validated. Calibration curves were linear with coefficients of determination (r2) >0.999 for desvenlafaxine and paroxetine. The limits of detection and the limits of quantification for both desvenlafaxine and paroxetine were 0.001 µg/mL and 0.02 µg/mL, respectively. Desvenlafaxine and paroxetine were detected in the postmortem samples, along with various psychiatric drugs, and the blood alcohol content level was below 0.010%. The concentrations of desvenlafaxine and paroxetine in the heart blood were 11.0 µg/mL and 2.1 µg/mL, respectively, indicating lethal concentrations. In the urine, the concentrations of desvenlafaxine and paroxetine were 87.7 µg/mL and 3.5 µg/mL, respectively. This is the first report to determine the blood concentration of desvenlafaxine in a fatal intoxication caused by an overdose of desvenlafaxine single formulation.

Omics analysis unveils changes in the metabolome and lipidome of Caenorhabditis elegans upon polydopamine exposure.

Polydopamine (PDA) is an insoluble biopolymer with a dark brown-black color that forms through the autoxidation of dopamine. Because of its outstanding biocompatibility and durability, PDA holds enormous promise for various applications, both in the biomedical and non-medical domains. To ensure human safety, protect health, and minimize environmental impacts, the assessment of PDA toxicity is important. In this study, metabolomics and lipidomics assessed the impact of acute PDA exposure on Caenorhabditis elegans (C. elegans). The findings revealed a pronounced perturbation in the metabolome and lipidome of C. elegans at the L4 stage following 24 hours of exposure to 100 µg/mL PDA. The changes in lipid composition varied based on lipid classes. Increased lipid classes included lysophosphatidylethanolamine, triacylglycerides, and fatty acids, while decreased species involved in several sub-classes of glycerophospholipids and sphingolipids. Besides, we detected 37 significantly affected metabolites in the positive and 8 in the negative ion modes due to exposure to PDA in C. elegans. The metabolites most impacted by PDA exposure were associated with purine metabolism, biosynthesis of valine, leucine, and isoleucine; aminoacyl-tRNA biosynthesis; and cysteine and methionine metabolism, along with pantothenate and CoA biosynthesis; the citrate cycle (TCA cycle); and beta-alanine metabolism. In conclusion, PDA exposure may intricately influence the metabolome and lipidome of C. elegans. The combined application of metabolomics and lipidomics offers additional insights into the metabolic perturbations involved in PDA-induced biological effects and presents potential biomarkers for the assessment of PDA safety.

Alkaloids and Lignans from the Aerial Parts of Rauvolfia tetraphylla Inhibit NO Production in LPS-activated RAW 264.7 Cells.

Three previously undescribed compounds named rauvolphyllas A-C (1-3), along with thirteen known compounds, 18ß-hydroxy-3-epi-α-yohimbine (4), yohimbine (5), α-yohimbine (6), 17-epi-α-yohimbine (7), (E)-vallesiachotamine (8), (Z)-vallesiachotamine (9), 16S-E-isositsirikine (10), Nb -methylisoajimaline (11), Nb -methylajimaline (12), ajimaline (13), (+)-lyoniresinol 3α-O-ß-D-glucopyranoside (14), (+)-isolarisiresinol 3α-O-ß-D-glucopyranoside (15), and (-)-lyoniresinol 3α-O-ß-D-glucopyranoside (16) were isolated from the aerial parts of Rauvolfia tetraphylla L. Their chemical structures were elucidated based on the extensive spectroscopic interpretation of HR-ESI-MS, 1D and 2D NMR spectra. The absolute configurations of 2 and 3 were determined by experimental ECD spectra. Compounds 5, 6, 7, and 11-13 exhibited nitric oxide production inhibition activity in LPS-activated RAW 264.7 cells with the IC50 values of 79.10, 44.34, 51.28, 33.54, 37.67, and 28.56 µM, respectively, compared to that of the positive control, dexamethasone, which showed IC50 value of 13.66 µM. The other isolates were inactive with IC50 values over 100 µM.

Inhibition by components of Glycyrrhiza uralensis of 3CLpro and HCoV-OC43 proliferation.

Coronavirus disease 2019 (COVID-19) is an infectious disease caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). 3CLpro is a key enzyme in coronavirus proliferation and a treatment target for COVID-19. In vitro and in silico, compounds 1-3 from Glycyrrhiza uralensis had inhibitory activity and binding affinity for 3CLpro. These compounds decreased HCoV-OC43 cytotoxicity in RD cells. Moreover, they inhibited viral growth by reducing the amounts of the necessary proteins (M, N, and RDRP). Therefore, compounds 1-3 are inhibitors of 3CLpro and HCoV-OC43 proliferation.

Identification and Quantitation of the Bioactive Components in Wasted Aralia elata Leaves Extract with Endothelial Protective Activity.

Aralia elata, a renowned medicinal plant with a rich history in traditional medicine, has gained attention for its potential therapeutic applications. However, the leaves of this plant have been largely overlooked and discarded due to limited knowledge of their biological activity and chemical composition. To bridge this gap, a comprehensive study was conducted to explore the therapeutic potential of the 70% ethanol extract derived from Aralia elata leaves (LAE) for the treatment of cardiovascular disease (CVD). Initially, the cytotoxic effects of LAE on human umbilical vein endothelial cells (HUVECs) were assessed, revealing no toxicity within concentrations up to 5 µg/mL. This suggests that LAE could serve as a safe raw material for the development of health supplements and drugs aimed at promoting cardiovascular well-being. Furthermore, the study found that LAE extract demonstrated anti-inflammatory properties in HUVECs by modulating the PI3K/Akt and MAPK signaling pathways. These findings are particularly significant as inflammation plays a crucial role in the progression of CVD. Moreover, LAE extract exhibited the ability to suppress the expression of adhesion molecules VCAM-1 and ICAM-1, which are pivotal in leukocyte migration to inflamed blood vessels observed in various pathological conditions. In conjunction with the investigation on therapeutic potential, the study also established an optimal HPLC-PDA-ESI-MS/MS method to identify and confirm the chemical constituents present in 24 samples collected from distinct regions in South Korea. Tentative identification revealed the presence of 14 saponins and nine phenolic compounds, while further analysis using PCA and PLS-DA allowed for the differentiation of samples based on their geographical origins. Notably, specific compounds such as chlorogenic acid, isochlorogenic acid A, and quercitrin emerged as marker compounds responsible for distinguishing samples from different regions. Overall, by unraveling its endothelial protective activity and identifying key chemical constituents, this research not only offers valuable insights for the development of novel treatments but also underscores the importance of utilizing and preserving natural resources efficiently.

Chiral separation and molecular modeling study of decursinol and its derivatives using polysaccharide-based chiral stationary phases.

Plant-based bioactive substances have long been used to treat inflammatory ailments, owing to their low toxicity and cost-effectiveness. To enhance plant treatment by eliminating undesirable isomers, optimizing the chiral separation techniques in pharmaceutical and clinical studies is important. This study reported a simple and effective method for chiral separation of decursinol and its derivatives, which are pyranocoumarin compounds with anti-cancer and anti-inflammatory properties. Baseline separation (Rs >1.5) was achieved using five different polysaccharide-based chiral stationary phases (CSPs) that differed in chiral origin, chiral selector chemistry, and preparation technique. To separate all six enantiomers simultaneously, n-hexane and three alcohol modifiers (ethanol, isopropanol, and n-butanol) were used as mobile phases in the normal-phase mode. The chiral separation ability of each column with various mobile phase compositions was compared and discussed. As a result, amylose-based CSPs with linear alcohol modifiers demonstrated superior resolution. Three cases of elution order reversal caused by modifications of CSPs and alcohol modifiers were observed and thoroughly analyzed. To elucidate the chiral recognition mechanism and enantiomeric elution order (EEO) reversal phenomenon, detailed molecular docking simulations were conducted. The R- and S-enantiomers of decursinol, epoxide, and CGK012 exhibited binding energies of -6.6, -6.3, -6.2, -6.3, -7.3, and -7.5 kcal/mol, respectively. The magnitude of the difference in binding energies was consistent with the elution order and enantioselectivity (α) of the analytes. The molecular simulation results demonstrated that hydrogen bonds, π-π interactions, and hydrophobic interactions have a significant impact on chiral recognition mechanisms. Overall, this study presented a novel and logical approach of optimizing chiral separation techniques in the pharmaceutical and clinical industries. Our findings could be further applied for screening and optimizing enantiomeric separation.

Lipid class-dependent alterations of Caenorhabditis elegans under harmane exposure.

Altered lipid patterns in Caenorhabditis elegans (C. elegans) resulting from exposure to harmane remain to be explored. In this study, untargeted lipidomics was carried out to elucidate the effects of acute exposure to harmane on the lipidome of C. elegans. Exposure to the compound was evaluated based on the reproduction ability of the worms at 0.1 and 1 µg/mL. No significant effects of harmane were observed at these concentrations. Furthermore, we found that the modulatory effects of harmane on the lipidome of C. elegans at 1 µg/mL were lipid class dependent. In particular, harmane-treated worms were enriched in triglycerides and fatty acids, regardless of the degree of saturation. Glycerophospholipids were generally down-regulated. Furthermore, functional analyses suggested that there was a reduction in lipid membrane bilayer-related terms, and in some related to the mitochondria, and endoplasmic reticulum of C. elegans when treated with harmane. Lipid droplets and storage appeared to be up-regulated. In conclusion, our findings suggest that harmane exposure affects the lipidome of C. elegans in a sophisticated manner. Further investigations are required to elucidate the molecular mechanisms underlying these lipid pattern changes.

Caenorhabditis elegans: a model organism in the toxicity assessment of environmental pollutants.

The unfavorable effects of environmental pollutants are becoming increasingly evident. In recent years, Caenorhabditis elegans (C. elegans) has been used as a powerful terrestrial model organism for environmental toxicity studies owing to its various advantages, including ease of culture, short lifespan, small size, transparent body, and well-characterized genome. In vivo bioassays and field studies can analyze and evaluate various toxic effects of the toxicants on the model organism, while emerging technologies allow profound insights into molecular disturbances underlying the observed phenotypes. In this review, we discuss the applications of C. elegans as a model organism in environmental toxicity studies and delineate apical assays such as lifespan, growth rate, reproduction, and locomotion, which are widely used in toxicity evaluation. In addition to phenotype assays, a comprehensive understanding of the toxic mode of action and mechanism can be achieved through a highly sensitive multi-omics approach, including the expression levels of genes and endogenous metabolites. Recent studies on environmental toxicity using these approaches have been summarized. This review highlights the practicality and advantages of C. elegans in evaluating the toxicity of environmental pollutants and presents the findings of recent toxicity studies performed using this model organism. Finally, we propose crucial technical considerations to escalate the appropriate use of C. elegans in examining the toxic effects of environmental pollutants.

Inotodiol, an antiasthmatic agent with efficacy and safety, preferentially impairs membrane-proximal signaling for mast cell activation.

While inhaled corticosteroids (ICSs) are the mainstay of asthma treatment, due to compliance, drug safety, and resistance issues, new medications to replace ICSs are in high demand. Inotodiol, a fungal triterpenoid, showed a unique immunosuppressive property with a preference for mast cells. It exerted a mast cell-stabilizing activity equally potent to dexamethasone in mouse anaphylaxis models when orally administered in a lipid-based formulation, upgrading bioavailability. However, it was four to over ten times less effective in suppressing other immune cell subsets, depending on the subsets, than dexamethasone showing invariably potent inhibition. Accordingly, inotodiol affected the membrane-proximal signaling for activating mast cell functions more profoundly than other subsets. Inotodiol also effectively prevented asthma exacerbation. Importantly, considering the no-observed-adverse-effect level of inotodiol was over 15 times higher than dexamethasone, its therapeutic index would be at least eight times better,implying that inotodiol is a viable option for replacing CSs in treating asthma.

The anti-obesity effect of mulberry leaf (Mori Folium) extracts was increased by bioconversion with Pectinex.

Mulberry leaf (Mori Folium) extract (MLE) is known to have anti-obesity effects. In this study, the enhanced effects of MLE after bioconversion treatment using Pectinex (BMLE) on obesity were explored, and the underlying mechanisms were investigated using the active components, neochlorogenic acid (5-CQA) and cryptochlorogenic acid (4-CQA), whose amounts were increased by bioconversion of MLE. Both MLE and BMLE inhibited lipid accumulation in 3T3-L1 adipocytes without cytotoxicity and suppressed the expression of CCAAT/enhancer-binding protein alpha (C/EBPα). In addition, MLE and BMLE decreased high-fat diet-induced adipose tissue mass expansion. Notably, BMLE significantly increased antiadipogenic and anti-obesity effects compared to MLE in vitro and in vivo. The active ingredients increased by bioconversion, 5-CQA and 4-CQA, inhibited the protein levels of C/EBPα and the mRNA levels of stearoyl-CoA desaturase 1 (Scd1). These findings provide new insights into the therapeutic possibility of using bioconversion of MLE, by which upregulation of 5-CQA and 4-CQA potently inhibits adipogenesis.

Network Pharmacology-Based Investigation on Therapeutic Mechanisms of the Angelica dahurica Radix and Ligusticum chuanxiong Rhizoma Herb Pair for Anti-Migraine Effect.

Migraines are a common neurological disorder characterized by desperate throbbing unilateral headaches and are related to phonophobia, photophobia, nausea, and vomiting. The Angelica dahurica Radix and Ligusticum chuanxiong Rhizoma herb pair (ALHP) has been used to treat migraines for centuries in traditional Chinese medicine (TCM). However, the physiological mechanisms of migraine treatment have not yet been elucidated. In this study, a total of 50 hub targets related to the effect of 28 bioactive compounds in ALHP on anti-migraine were obtained through network pharmacology analysis. GO and KEGG analyses of the hub targets demonstrated that ALHP treatment of migraines significantly involved the G-protein-coupled receptor signaling pathway, chemical synaptic transmission, inflammatory response, and other biological processes. According to the degree of gene targets in the network, ACE, SLC3A6, NR3CI, MAPK1, PTGS2, PIK3CA, RELA, GRIN1, GRM5, IL1B, and DRD2 were found to be the core gene targets. The docking results showed a high affinity for docked conformations between compounds and predicted targets. The results of this study suggest that ALHP could treat migraines by regulating immunological functions, diminishing inflammation, and improving immunity through different physiological pathways, which contributes to the scientific base for more in-depth research as well as for a more widespread clinical application of ALHP.

Inhibitory Activity of Quaternary Isoquinoline Alkaloids on Soluble Epoxide Hydrolase.

The quaternary isoquinoline alkaloids of palmatine (1), berberine (2), and jatrorrhizine (3) were evaluated in terms of their ability to inhibit soluble epoxide hydrolase (sEH). They had similar inhibitory activities, with IC50 values of 29.6 ± 0.5, 33.4 ± 0.8, and 27.3 ± 0.4 µM, respectively. Their respective Ki values of 26.9, 46.8, and 44.5 µM-determined by enzyme kinetics-indicated that they inhibited the catalytic reaction by binding noncompetitively with sEH. The application of computational chemistry to the in vitro results revealed the site of the receptor to which the ligand would likely bind. Accordingly, three alkaloids were identified as having a suitable basic skeleton for lead compound development of sEH inhibitors.

Extraction and Concentration of Waste Pueraria lobata Stems with Antioxidants and Anti-Melanogenesis Activity as a Novel Skin Whitening Agent for Natural Cosmetic Prototypes.

The root of Pueraria lobata (Willd.) is used commercially in different products, including dietary supplements, cosmetics, and teas, but its stem part is rarely used and studied. Therefore, this study evaluated the antioxidant and anti-melanogenesis activities of the bioactive fraction of P. lobata stem and investigated whether the activated carbon decolorization technique would have an impact on its activity and chemical composition. We observed that the dichloromethane fraction of P. lobata stem (DCM-PLS) has excellent antioxidant and anti-melanin synthesis activity at a concentration of 50 µg/mL. For the investigation of the anti-melanogenesis mechanism, we evaluated the mRNA expression of tyrosinase, which was depressed by the DCM-PLS. Daidzin was identified as the main active ingredient in DCM-PLS by using a high-performance liquid chromatography-diode array detector-hyphenated with tandem mass spectrometry. In addition, the activated carbon decolorization technology has no negative impact on the main components and bioactivity of DCM-PLS. DCM-PLS also did not induce any skin response in the human skin safety test. Collectively, DCM-PLS could be used as a natural type of skin-whitening agent in skin care products.

Phenolic Profile and Fingerprint Analysis of Akebia quinata Leaves Extract with Endothelial Protective Activity.

In contrast to the stem and fruit of Akebia quinata, A. quinata leaves as a source rich in phenolic compounds with potentially beneficial pharmacological activities have been largely overlooked. To develop and use A. quinata leaves as a resource, we evaluated its potential as a cardiovascular-protective agent. Herein, we investigated the effects and potential mechanisms of A. quinata leaves extract on lipopolysaccharide (LPS)-induced inflammatory responses in human umbilical vein endothelial cells. We found that A. quinata leaves extract pretreatment of 10 µg/mL significantly attenuated LPS-induced protein expression of intercellular adhesion molecule-1, vascular cell adhesion molecule-1. Furthermore, this extract also suppressed LPS-induced phosphorylation of nuclear factor-κB p65. In order to elucidate the chemical profiles of the samples, the HPLC fingerprint was established, and prominent peaks were identified via HPLC-electrospray ionization-mass spectrometry. Multivariate statistical analyses, including hierarchical cluster analysis, principal component analysis, and partial least-squares discriminant analysis, were performed to evaluate the clustering of the samples. It was found that isochlorogenic acid C was a key marker for the classification of A. quinata leaves from the Gongju and Muju city in Korea. Collectively, this study not only suggested the potential of A. quinata leaves as a novel therapeutic candidate for inflammatory cardiovascular disease but also developed a quality control method for A. quinata leaves, which could help to expand the application of A. quinata.

Effect of citric acid and heat treatment on the content of less-polar ginsenosides in flower buds of Panax ginseng.

Ginseng flower bud (GFB), as an inexpensive part of Panax ginseng, attracted significant attention as a beneficial functional food with medicinal potentials due to its high content of ginsenosides. A few studies focused on the utilization of heat treatment and citric acid treatment to process ginseng flowers, converting its polar ginsenosides into rare ginsenosides to improve its biological activities. Thus, in this study, we compared the changes of ginsenosides in GFB after citric acid and heat treatment by HPLC method. The results revealed that less-polar ginsenoside, Rg6 and F4, increased to 1.01 and 0.27% by heat treatment, respectively. Further, ginsenoside F2 increased to 1.13% with 1 M citric acid treatment. Furthermore, based on the combination of these two processing methods for the first time, the conversion rate of less-polar ginsenosides surged to 80%. The content of ginsenoside Rg3(s) and Rg5 increased to 1.509 and 1.871%, respectively, by simultaneous heat and citric acid treatment. Therefore, a processing approach that simultaneously performs heat and citric acid treatments has been proposed, and this considerably inexpensive and convenient processing method could be applied to the processing of GFBs and produce less-polar ginsenosides.

Special Issue Editorial: Isolation and Analysis of Characteristic Compounds from Herbal and Plant Extracts.

Herbal and plant extracts exhibit various types of properties and activities that have been applied in the medicinal field to treat diseases and achieve better health [...].

Anti-Wrinkle Effect of Isatis indigotica Leaf Extract: Evaluation of Antioxidant, Anti-Inflammation, and Clinical Activity.

Isatis indigotica leaf is an oriental herbal medicine that has been known for various pharmacological effects. However, its anti-wrinkle activity has not been fully evaluated. Therefore, we evaluated the anti-wrinkle effect of I. indigotica leaf extract on human skin. The purified extract inhibited 85.4% of 2,2-diphenyl-1-1picrylhydrazyl and 72.2% of 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt radicals at a concentration of 1 mg/mL. Nitrite production was reduced by 30% after treatment with 50 µg/mL of extract. Three fractions from the extract downregulated the mRNA expression of matrix metalloproteinase-1 and -3 and upregulated the expression of interleukin 4. Among the three fractions, fraction 2 exhibited the highest activity. The major component of the extract was identified as 3,4,5-trimethoxycinnamic acid by liquid chromatography coupled with mass spectrometry. Molecular docking was conducted to predict the binding mechanism of 3,4,5-trimethoxycinnamic with matrix metalloproteinase-1 and -3, and their binding energies were -5.20 and -4.89 kcal/mol, respectively. In a clinical trial, five roughness values of visiometer and visual score were significantly reduced in treated groups compared with the placebo group after 8 weeks. I. indigotica leaf extract inhibits wrinkle formation, and could be a potential anti-wrinkle agent. This is the first clinical trial demonstrating its anti-wrinkle activity.

Evaluation of Anti-Melanogenesis Activity of Enriched Pueraria lobata Stem Extracts and Characterization of Its Phytochemical Components Using HPLC-PDA-ESI-MS/MS.

The root of Pueraria lobata (Willd.) is a widely used herbal medicine worldwide, whereas the stem of the plant is discarded or used as feed for livestock. To reuse and exploit the stem of P. lobata as a resource, we investigated its potential as a skin-whitening agent. We found that the developed, enriched P. lobata stem (PLS) extract significantly inhibited melanin production in the 3-isobutyl-1-methylxanthine-induced B16/F10 cells at a concentration of 50 µg/mL. To further confirm the mechanism of the antimelanogenic effect of the enriched PLS extracts, we examined the mRNA expression of tyrosinase, which was suppressed by the extracts. To standardize and implement effective quality control of the enriched PLS extracts, its major chemical constituents were identified by high-performance liquid chromatography-photodiode array-electrospray ionization-mass spectrometry. In total, 12 constituents were identified. In silico analysis showed that the main constituents, puerarin and daidzin, had excellent binding affinities for human tyrosinase. Collectively, our results suggest that the PLS extracts could be used as anti-pigmentation agents.

Metabolomic Studies for the Evaluation of Toxicity Induced by Environmental Toxicants on Model Organisms.

Environmental pollution causes significant toxicity to ecosystems. Thus, acquiring a deeper understanding of the concentration of environmental pollutants in ecosystems and, clarifying their potential toxicities is of great significance. Environmental metabolomics is a powerful technique in investigating the effects of pollutants on living organisms in the environment. In this review, we cover the different aspects of the environmental metabolomics approach, which allows the acquisition of reliable data. A step-by-step procedure from sample preparation to data interpretation is also discussed. Additionally, other factors, including model organisms and various types of emerging environmental toxicants are discussed. Moreover, we cover the considerations for successful environmental metabolomics as well as the identification of toxic effects based on data interpretation in combination with phenotype assays. Finally, the effects induced by various types of environmental toxicants in model organisms based on the application of environmental metabolomics are also discussed.

A Novel Bioanalytical Method for Determination of Inotodiol Isolated from Inonotus Obliquus and Its Application to Pharmacokinetic Study.

In this study, we developed a bioanalytical method using liquid chromatography coupled to triple quadrupole tandem mass spectrometry (LC-MS/MS) to apply to a pharmacokinetic study of inotodiol, which is known for its anti-cancer activity. Plasma samples were prepared with alkaline hydrolysis, liquid-liquid extraction, and solid-phase extraction. Inotodiol was detected in positive mode with atmospheric pressure chemical ionization by multiple-reaction monitoring mode using LC-MS/MS. The developed method was validated with linearity, accuracy, and precision. Accuracy ranged from 97.8% to 111.9%, and the coefficient of variation for precision was 1.8% to 4.4%. The developed method was applied for pharmacokinetic study, and the mean pharmacokinetic parameters administration were calculated as follows: λz 0.016 min-1; T1/2 49.35 min; Cmax 2582 ng/mL; Cl 0.004 ng/min; AUC0-t 109,500 ng×min/mL; MRT0-t 32.30 min; Vd 0.281 mL after intravenous administration at dose of 2 mg/kg and λz 0.005 min-1; T1/2 138.6 min; Tmax 40 min; Cmax 49.56 ng/mL; AUC0-t 6176 ng×in/mL; MRT0-t 103.7 min after oral administration. The absolute oral bioavailability of inotodiol was 0.45%, similar to nonpolar phytosterols. Collectively, this is the first bioanalytical method and pharmacokinetic study for inotodiol.