Lefamulin: The First Systemic Pleuromutilin Antibiotic.

OBJECTIVE: To review the pharmacology, microbiology, efficacy, and safety of lefamulin. DATA SOURCES: A literature search was performed using PubMed and Google Scholar (2010 to end-April 2020) with the search terms BC-3781 and lefamulin. Other resources included abstracts presented at recent conferences, prescribing information, and the manufacturer's and Food and Drug Administration websites. STUDY SELECTION AND DATA EXTRACTION: All relevant English-language articles of studies assessing the efficacy and safety of lefamulin were included. DATA SYNTHESIS: Lefamulin is a pleuromutilin antibiotic with activity against Staphylococcus aureus, Streptococcus pneumoniae, and atypical bacteria. Lefamulin, given at the dose of 150 mg intravenously or 600 mg orally on an empty stomach every 12 hours for 5 to 7 days, was proven noninferior to moxifloxacin for the treatment of community-acquired bacterial pneumonia (CABP). Common adverse reactions include injection site reactions, hepatic enzyme elevation, gastrointestinal upset, hypokalemia, insomnia, and headache. Lefamulin is associated with QT prolongation, and concomitant use with CYP3A substrates that prolong the QT interval is contraindicated. Lefamulin may cause fetal harm. RELEVANCE TO PATIENT CARE AND CLINICAL PRACTICE: Lefamulin is a novel antibiotic with a unique mechanism of action. It represents an alternative option to ß-lactams and macrolides in the treatment of adults with CABP and an alternative option to amoxicillin and doxycycline in the outpatient setting given the rise in resistance to macrolides and safety concerns with fluoroquinolones. Nausea, vomiting, and diarrhea may limit the tolerability of the oral formulation. CONCLUSIONS: Lefamulin is the first systemic pleuromutilin antibiotic that has proven safe and effective for adults with CABP.

Antibacterial Activity and Pharmacokinetic Profile of a Promising Antibacterial Agent: 22-(2-Amino-phenylsulfanyl)-22-Deoxypleuromutilin.

A new pleuromutilin derivative, 22-(2-amino-phenylsulfanyl)-22-deoxypleuromutilin (amphenmulin), has been synthesized and proved excellent in vitro and in vivo efficacy than that of tiamulin against methicillin-resistant Staphylococcus aureus (MRSA), suggesting this compound may lead to a promising antibacterial agent to treat MRSA infections. In this study, the effectiveness and safety of amphenmulin were further investigated. Amphenmulin showed excellent antibacterial activity against MRSA (minimal inhibitory concentration = 0.0156~8 µg/mL) and performed time-dependent growth inhibition and a concentration-dependent postantibiotic effect (PAE). Acute oral toxicity test in mice showed that amphenmulin was a practical non-toxic drug and possessed high security as a new drug with the 50% lethal dose (LD50) above 5000 mg/kg. The pharmacokinetic properties of amphenmulin were then measured. After intravenous administration, the elimination half-life (T1/2), total body clearance (Clß), and area under curve to infinite time (AUC0→∞) were 1.92 ± 0.28 h, 0.82 ± 0.09 L/h/kg, and 12.23 ± 1.35 µg·h/mL, respectively. After intraperitoneal administration, the T1/2, Clß/F and AUC0→∞ were 2.64 ± 0.72 h, 4.08 ± 1.14 L/h/kg, and 2.52 ± 0.81 µg·h/mL, respectively, while for the oral route were 2.91 ± 0.81 h, 6.31 ± 2.26 L/h/kg, 1.67 ± 0.66 µg·h/mL, respectively. Furthermore, we evaluated the antimicrobial activity of amphenmulin in an experimental model of MRSA wound infection. Amphenmulin enhanced wound closure and promoted the healing of wound, which inhibited MRSA bacterial counts in the wound and decreased serum levels of the pro-inflammatory cytokines TNF-α, IL-6, and MCP-1.

Pharmacokinetics and tolerability of lefamulin following intravenous and oral dosing.

OBJECTIVES: To explore the pharmacokinetics (PK) of oral and intravenous (iv) lefamulin after single and multiple doses, and the effect of food on bioavailability. METHODS: Lefamulin PK was examined in four studies. In Study 1, PK was assessed in patients with acute bacterial skin and skin structure infections who received repeated iv lefamulin q12h (150 mg). In Study 2, a four-period crossover study, healthy subjects received a single dose of oral lefamulin [immediate-release (IR) tablet, 1 × 600 mg] in a fasted and fed state, oral lefamulin (capsule, 3 × 200 mg) in a fasted state, and iv lefamulin in a fasted state. In Study 3, a three-period crossover study, healthy males received a single oral lefamulin dose (IR) in the following states: fasted, fasted followed by a high-calorie meal 1 h post-dose, and fed. Study 4 had two parts; in part A, healthy males received a single lefamulin dose (IR) in a fasted and fed state; in part B, subjects received repeated doses of lefamulin (IR, q12h) or placebo. Adverse events (AEs) were recorded in each study. RESULTS: Single and repeated dosing of iv and oral lefamulin resulted in comparable exposure. Intravenous and oral lefamulin (given fasted or with a meal 1 h post-dose) resulted in bioequivalence. Bioequivalence was not established between oral lefamulin in the fed state and iv or oral administration in the fasted state. All AEs were mild or moderate in severity, no serious AEs were reported, and no participant discontinued because of an AE. CONCLUSIONS: The PK of lefamulin supports successful switch from iv to oral therapy; lefamulin was generally well tolerated.

Prediction of lefamulin epithelial lining fluid penetration after intravenous and oral administration using Phase 1 data and population pharmacokinetics methods.

OBJECTIVES: Lefamulin is a semi-synthetic intravenous and oral pleuromutilin antibiotic with activity against pathogens commonly associated with community-acquired bacterial pneumonia. Using data from two Phase 1 studies, a population pharmacokinetics (PPK) model for lefamulin in plasma and epithelial lining fluid (ELF) was constructed. METHODS: Plasma pharmacokinetic (PK) data from a crossover, bioavailability, food-effect study and plasma and ELF PK data from a tissue penetration study in normal healthy volunteers were used to construct a PPK model for lefamulin. Model development involved refinement of a previous PPK model for intravenous and oral administration, followed by application of the model to plasma and ELF data from the tissue penetration study. The ELF penetration ratio of lefamulin was determined using model-based simulations. RESULTS: The PPK analysis data set contained 1103 plasma and 12 ELF lefamulin concentrations from 32 subjects. A three-compartment model with non-linear protein binding and two parallel absorption processes provided precise and unbiased estimated plasma concentration-time profiles. The absorption rate was slower and bioavailability was decreased after a high-fat/high-calorie meal. ELF data were well described using first-order rate constants into and out of the ELF compartment. The median predicted lefamulin total-drug ELF AUC0-24/free-drug plasma AUC0-24 ratio was ∼5:1 after intravenous or oral administration. CONCLUSIONS: The final PPK model allowed precise characterization of plasma and ELF exposures after intravenous and oral administration. The high ELF penetration ratio suggests that the penetration of lefamulin into the effect site is rapid and extensive, irrespective of route of administration.

Introduction: lefamulin and pharmacokinetic/pharmacodynamic rationale to support the dose selection of lefamulin.

Lefamulin is the first semisynthetic pleuromutilin being developed for oral and intravenous administration. The drug selectively inhibits prokaryotic ribosomal protein synthesis by binding to the peptidyl transferase centre via four H-bonds and other interactions, resulting in an 'induced fit' that tightens the binding pocket around lefamulin. This unique mechanism of action has been associated with a low probability of cross-resistance to other antimicrobial classes commonly used to treat community-acquired bacterial pneumonia (CABP). This Supplement, entitled 'Pharmacokinetic and pharmacodynamic analyses and dose rationale for lefamulin, a novel pleuromutilin antibiotic, for the treatment of community-acquired bacterial pneumonia', is intended to be a valuable resource for both clinicians and researchers. It provides the essential pharmacokinetic and pharmacodynamic data on lefamulin that were used to support the optimal dose selection of lefamulin for the safe and effective treatment of CABP in adults.

Pharmacokinetic/pharmacodynamic target attainment analyses to support intravenous and oral lefamulin dose selection for the treatment of patients with community-acquired bacterial pneumonia.

OBJECTIVES: Lefamulin is a semi-synthetic intravenous (iv) and oral pleuromutilin antibiotic active against community-acquired bacterial pneumonia (CABP) pathogens. Pharmacokinetic/pharmacodynamic (PK/PD) target attainment analyses were carried out to evaluate lefamulin 150 mg iv q12h and 600 mg orally q12h under fed and fasted conditions for the treatment of patients with CABP. METHODS: The analyses undertaken used a population PK model based on Phase 1 PK data, non-clinical PK/PD targets for efficacy and in vitro surveillance data for Streptococcus pneumoniae (SP) and Staphylococcus aureus (SA), and Monte Carlo simulation. Percentage probabilities of PK/PD target attainment by MIC on day 1 were determined using median total-drug epithelial lining fluid (ELF) and free-drug plasma AUC:MIC ratio targets associated with 1 and 2 log10 cfu reductions from baseline. RESULTS: Percentage probabilities of attaining the total-drug ELF AUC:MIC ratio target for a 1 log10 cfu reduction from baseline for SP were ≥99.2% at the MIC90 of 0.12 mg/L and 96.7%, 82.1% and 96.3% for iv and oral dosing regimens under fed and fasted conditions, respectively, at the MIC99 of 0.25 mg/L. Percentage probabilities of attaining the free-drug plasma AUC:MIC target for the same endpoint at the SP MIC99 were 100% for each regimen. For the SA MIC90 of 0.12 mg/L and AUC:MIC ratio targets for the same endpoint, percentage probabilities were 92.7%-100% for iv and oral dosing regimens. CONCLUSIONS: These data provide support for lefamulin 150 mg iv q12h and 600 mg orally q12h for the treatment of patients with CABP and suggest that doses may not need to be taken under fasted conditions.

In vivo pharmacodynamics of lefamulin, the first systemic pleuromutilin for human use, in a neutropenic murine thigh infection model.

OBJECTIVES: To characterize the pharmacokinetics (PK) and pharmacodynamics (PD) of lefamulin in the neutropenic murine thigh infection model to ascertain (i) which PK/PD index best correlates with efficacy and (ii) whether the magnitude of the index that drives efficacy varies for different pathogens. METHODS: We evaluated the in vivo PK/PD of lefamulin against five Streptococcus pneumoniae and five Staphylococcus aureus strains using a neutropenic murine thigh infection model. The relationships between bacterial burden in the thigh of normal and neutropenic mice after 24 h of lefamulin treatment and various PK/PD indices were determined. RESULTS: The kinetics of the three doses was linear by AUC. Rate of killing was maximal at concentrations near the MIC; suppression of regrowth was dose dependent, with a post-antibiotic effect of 3.0-3.5 and 1.0-1.5 h against S. pneumoniae and S. aureus, respectively. The efficacy of lefamulin correlated most strongly with the AUC0-24/MIC ratio; coefficient of determination was 79.9% for S. pneumoniae and 78.3% for S. aureus. The magnitude of the 24 h AUC/MIC of total drug required ranged from 9.92 to 32.1 for S. pneumoniae and 40.2 to 82.5 for S. aureus, corresponding to free drug values (∼20% free fraction) of 1.98-6.42 and 8.04-16.5, respectively. CONCLUSIONS: Lefamulin, the first systemically available pleuromutilin in humans, exhibits time- and concentration-dependent killing. The presence of white blood cells had only a slight effect in enhancing the activity of the drug, indicating a leucocyte-independent effect. The identified driver of efficacy, the AUC0-24/MIC ratio and the ratios determined against various S. aureus and S. pneumoniae strains, will inform further non-clinical and clinical trials.

Pharmacokinetics/pharmacodynamics of lefamulin in a neutropenic murine pneumonia model with Staphylococcus aureus and Streptococcus pneumoniae.

OBJECTIVES: To present results of preclinical studies that supported further development of lefamulin for treating patients with community-acquired bacterial pneumonia (CABP). METHODS: The effect of bovine lung surfactant on the antibacterial activity of lefamulin against Streptococcus pneumoniae and Staphylococcus aureus was determined by broth microdilution assay. In vitro accumulation of lefamulin was evaluated in J774 mouse macrophages. Pharmacokinetics was assessed in female BALB/c (Bagg albino) mice treated with subcutaneous lefamulin (35 or 70 mg/kg). In neutropenic lung infection experiments, BALB/c mice received intraperitoneal cyclophosphamide before challenge with single S. pneumoniae or S. aureus strains; subcutaneous lefamulin (1.25-160 mg/kg) was given twice daily post-infection. Hill models described relationships between AUC/MIC ratios and changes in log10 cfu. RESULTS: Lung surfactant did not significantly increase lefamulin MIC values against test strains. Lefamulin uptake in macrophages was rapid (a plateau was reached in ∼3 h). In mice, distribution of lefamulin [plasma to epithelial lining fluid (ELF)] was rapid, showing an ∼2-fold increase in lefamulin exposure in the ELF during the 5.5 h period. Median plasma AUC/MIC ratios associated with 1 and 2 log10 cfu reductions from baseline were 1.37 and 2.15, respectively, for S. pneumoniae and 2.13 and 6.24 for S. aureus. Corresponding ELF results were 14.0 and 22.0 for S. pneumoniae and 21.7 and 63.9 for S. aureus. CONCLUSIONS: Overall, lefamulin displays desirable pharmacokinetic/pharmacodynamic relationships that are predictive of the clinical effectiveness of lefamulin and other antibacterial agents used to treat CABP.

Interaction of deoxyschizandrin and schizandrin B with liver uptake transporters OATP1B1 and OATP1B3.

1. Deoxyschizandrin and schizandrin B have diverse pharmacological effects, including hepatoprotective activity. We aim to study their hepatic uptake and their effects on the hepatic uptake of other clinical drugs mediated by OATP1B1 and OATP1B3. 2. Deoxyschizandrin exhibited a high affinity for OATP1B1 with Km of 17.61 ± 0.43 µM but a low affinity for OATP1B3. Similarly, schizandrin B also showed a strong affinity for OATP1B1 with Km of 18.45 ± 1.23 µM but a weak affinity for OATP1B3. 3. Atorvastatin and rifampicin could inhibit the uptake of deoxyschizandrin and schizandrin B mediated by OATP1B1. 4. Intriguingly, both deoxyschizandrin and schizandrin B significantly promoted the uptake of atorvastatin (with EC50 of 50.58 ± 8.08 and 24.70 ± 5.82 µM, respectively) and rosuvastatin (with EC50 of 13.46 ± 2.70 and 8.99 ± 4.73 µM, respectively) mediated by OATP1B1. Deoxyschizandrin could markedly promote the uptake of fluvastatin but inhibit the uptake of sodium taurocholate (TCNa) mediated by OATP1B1. 5. The promotion on hepatic uptake of statins mediated by OATP1B1 might lead to enhanced efficacy of cholesterol lowering and reduced risk of myopathy for hyperlipidemia patients when given statins together with deoxyschizandrin or schizandrin B.

Pharmacokinetics and liver uptake of three Schisandra lignans in rats after oral administration of liposome encapsulating ß-cyclodextrin inclusion compound of Schisandra extract.

Schisandra chinensis fructus (SCF) is widely used traditional Chinese medicine, which possesses hepato-protective potential. Schisandrin (SD), schisantherin (ST), and γ-schizandrin (SZ) are the major bioactive lignans. The main problem associated with the major bioactive lignans oral administration is low oral bioavailability due to the lignans' poor aqueous solubility and taste. The aim of the present research work was to develop liposome (SCL) encapsulated ß-cyclodextrin (ß-CD) inclusion complex loaded with SCF extract (SCF-E). The SD, ST, and SZ were selected as effective candidates to perform comparisons of liver targeting among the solution (SES), ß-cyclodextrin inclusion compound (SCF-E-ß-CD), liposome (SEL), and SCL of SCF-E to characterize the pharmacokinetic behaviors and liver targeting in rats. The ß-CD inclusion complex (SCF-E-ß-CD) was used to improve the solubility. The concentrations were determined using high-performance liquid chromatography (HPLC) and analyzed by DAS3.0. The pharmacokinetic results indicate that the plasma concentration-time courses were fitted well to the one-compartment model with the first weighing factor. The half-life period (t1/2) and area under the concentration-time curve (AUC) of the three components in SCL were the largest. The SCL exhibit a relatively high liver targeting effect. The results would be helpful for guiding the clinical application of this herbal medicine.

A Rapid UPLC-MS Method for Quantification of Gomisin D in Rat Plasma and Its Application to a Pharmacokinetic and Bioavailability Study.

Gomisin D, a lignan compound isolated from Fructus Schisandra, is a potential antidiabetic and anti-Alzheimer's agent. Recently, gomisin D was used as a quality marker of some traditional Chinese medicine (TCM) formulas. In this study, a rapid ultra-performance liquid chromatography/tandem mass spectrometry method (UPLC-MS/MS) was developed and validated to quantify gomisin D in rat plasma for a pharmacokinetic and bioavailability study. Acetonitrile was used to precipitate plasma proteins. Separations were performed on a BEH C18 column with a gradient mobile phase comprising of acetonitrile and water (0.1% formic acid). An electrospray ionization source was applied and operated in the positive ion mode. The multiple reaction monitoring mode (MRM) was utilized to quantify gomisin D and nomilin (internal standard, IS) using the transitions of m/z 531.2 → 383.1 and m/z 515.3 → 161.0, respectively. The calibration curve was linear over the working range from 1 to 4000 ng/mL (R² = 0.993). The intra- and interday precision ranged from 1.9% to 12.9%. The extraction recovery of gomisin D was in the range of 79.2-86.3%. The validated UPLC-MS/MS method was then used to obtain the pharmacokinetic characteristics of gomisin D after intravenous (5 mg/kg) and intragastric (50 mg/kg) administration to rats. The bioavailability of gomisin D was 107.6%, indicating that this compound may become a promising intragastrical medication. Our results provided useful information for further preclinical studies on gomisin D.

A Validated HPLC-MS/MS Assay for 14-O-[(4,6-Diaminopyrimidine-2-yl)thioacetyl] Mutilin in Biological Samples and Its Pharmacokinetic, Distribution and Excretion via Urine and Feces in Rats.

14-O-[(4,6-Diaminopyrimidine-2-yl)thioacetyl] mutilin (DPTM), a novel pleuromutilin candidate with a substituted pyrimidine moiety, has been confirmed to possess excellent antibacterial activity against Gram-positive bacteria. To illustrate the pharmacokinetic profile after intravenous (i.v.), intramuscular (i.m.) and oral (p.o.) administrations with DPTM, as well as tissue distribution and excretion via urine and feces in vivo, a specific, sensitive and robust HPLC-MS/MS method was first developed to determine DPTM in rat plasma, various tissues, urine and feces. The plasma, tissues, urine and feces samples were treated by protein precipitation with acetonitrile using tiamulin fumarate as an internal standard (IS). This method which was achieved on an HPLC system detector equipped with an ESI interface, was sensitive with 5 ng/mL as the lower limit of detection and exhibited good linearity (R² > 0.9900) in the range of 5⁻4000 ng/mL for plasma, various tissues, urine and feces, as well as intra-day precision, inter-day precision and accuracy. The matrix effects ranged from 94.2 to 109.7% with RSD ≤ 9.4% and the mean extraction recoveries ranged from 95.4 to 109.5% in plasma, tissue homogenates, urine and feces (RSD ≤ 9.9). After i.v., i.m. and p.o. administrations, DPTM was rapidly absorbed and metabolized in rats with the half-life (t1/2) of 1.70⁻1.86, 3.23⁻3.49 and 4.38⁻4.70 for 10, 25 and 75 mg/kg doses, respectively. The tissue distribution showed that DPTM was diffused into all the tested tissues, especially into the intestine and lung. Excretion via urine and feces studies demonstrated that DPTM was mainly excreted by feces after administration.

Study on the pharmacokinetics of deoxyschizandrin and schizandrin in combination with epigallocatechin gallate, a component of green tea, in rats.

1. Green tea is commonly used worldwide due to its potential positive health benefits. We have examined the effects of epigallocatechin gallate (EGCG), the most abundant catechin in green tea, on the pharmacokinetics of deoxyschizandrin (DSD) and schizandrin (SD), which are the representative lignans in popular traditional Chinese medicines Fructus schisandrae, in rats. 2. The effects on the transport in Caco-2 cells and metabolism in human liver microsomes (HLMs) of DSD and SD by EGCG were determined to analyze their interactions thoroughly. 3. In pharmacokinetic studies, rats were divided into four groups. Each group was orally treated with DSD alone (Group 1), DSD combined with EGCG (Group 2), SD alone (Group 3) and SD combined with EGCG (Group 4). The pharmacokinetic parameters of DSD and SD in rats were determined by UPLC-MS/MS. 4. The in vivo results indicated that EGCG had no significant influence on the pharmacokinetic behaviors of DSD or SD in rats, which were in accordance with the in vitro transport and metabolism studies. However, there were marked differences between male and female rats among Cmax, AUC0-t, AUC0-∞ of DSD and SD. This disparity suggested that gender differences might exist in the pharmacokinetic processes of DSD or SD in rats.

Investigation of pharmacokinetics, tissue distribution and excretion of schisandrin B in rats by HPLC-MS/MS.

Schisandrin B has received much attention owing to its various biological activities. The present study was aimed at the formulation development of schisandrin B and investigation of the pharmacokinetic profiles, distribution and excretion of schisandrin B in Sprague-Dawley rats. In this study, micronized schisandrin B particles with particle size of 10-20 µm were chosen as the research object. Chromatographic separation was carried out on a BDS Hypersil C18 column (50 × 2.1 mm, i.d. 3.5 µm). Schisandrin B and deoxyschizandrin (internal standard) were detected without interference in the multiple reaction monitoring mode with positive electrospray ionization. The pharmacokinetic parameters were calculated by a noncompartmental method. The area under concentration-time curve and the maximum concentration showed a significant difference in gender. The calculated absolute oral bioavailability of schisandrin B was ~55.0% for female rat and 19.3% for male rat. Schisandrin B exhibited linear pharmacokinetics properties within the range of the tested oral dose (10, 20 and 40 mg/kg). After oral administration of schisandrin B, it was extensively distributed in ovary and adipose tissue. The result also showed very low urinary, biliary and fecal excretion of schisandrin B implying that schisandrin B was excreted mainly in the forms of metabolites.

Development and validation of an LC-MS/MS method for the simultaneous quantification of seven constituents in rat plasma and application in a pharmacokinetic study of the Zaoren Anshen prescription.

A sensitive, specific and accurate liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed and validated for the simultaneous determination of seven constituents of the Zaoren Anshen prescription (ZAP) in rat plasma after oral administration of the ZAP: spinosin, salvianic acid A, 6'''-feruloylspinosin, protocatechualdehyde, salvianolic acid B, schisandrin and deoxyschisandrin. The plasma samples and the internal standard (IS) sulfamethoxazole were extracted using acetonitrile. Chromatographic separation was performed with an Agilent HC-C18 column using a gradient elution profile and a mobile phase consisting of 0.01% formic acid in water (A) and acetonitrile (B). The analytes were quantified simultaneously in a single run using an ion trap mass spectrometer operated in the multiple reaction monitoring mode and electrospray ion-source polarity in the positive and negative modes. The calibration curves for spinosin, salvianic acid A, 6'''-feruloylspinosin, protocatechualdehyde, salvianolic acid B, schisandrin and deoxyschisandrin were linear over the concentration ranges of 2.90-1160, 2.50-1000, 1.80-720, 0.65-260, 2.50-1000, 8.00-1600 and 1.30-520 ng/mL, respectively. The intra- and inter-day precisions in terms of relative standard deviation were <18.9%, and the accuracies in terms of relative error were within ±14.2%. Consequently, the proposed method was successfully applied to the pharmacokinetic analysis of these seven major active compounds in rats administered ZAP. These results will facilitate research aiming to predict the effectiveness of the optimal dose of ZAP and might be beneficial for the therapeutic use of ZAP in the clinical setting.

Pharmacokinetics of Schizandrin and Its Pharmaceutical Products Assessed Using a Validated LC-MS/MS Method.

Schisandra chinensis has been used as an important component in various prescriptions in traditional Chinese medicine and, more recently, in Western-based medicine for its anti-hepatotoxic effect. The aim of this study was to develop a selective, rapid, and sensitive ultra-performance liquid chromatography-tandem mass spectrometry method for pharmacokinetic studies of schizandrin in rats. Liquid-liquid extraction was used for plasma sample preparation. A UHPLC reverse-phase C18e column (100 mm × 2.1 mm, 2 µm) coupled with a mobile phase of methanol-0.1% formic acid (85:15, v/v) was used for sample separation. A triple quadrupole tandem mass spectrometer was used to detect the analytes in the selected reaction monitoring mode. The linear range of schizandrin in rat plasma was 5.0-1000 ng/mL (r² > 0.999), with a lower limit of quantification of 5 ng/mL. The method was validated with regard to accuracy, intra-day and inter-day precision, linearity, stability, recovery, and matrix effects in rat plasma, which were acceptable according to the biological method validation guidelines developed by the FDA. This method was successfully applied to a pharmacokinetic study after oral administration of 3 g/kg and 10 g/kg of Schisandra chinensis products, which yielded a maximum concentration of schizandrin of 0.08 ± 0.07 and 0.15 ± 0.09 µg/mL, respectively. A parallel study design was used to investigate the oral bioavailability of single compound of schizandrin and the herbal extract, the single compound of pure schizandrin (10 mg/kg, i.v.), pure schizandrin (10 mg/kg, p.o.), and the herbal extract of Schisandra chinensis (3 g/kg and 10 g/kg, p.o.) were given individually. The dose of Schisandra chinensis (3 g/kg) equivalent to schizandrin (5.2 mg/kg); the dose of Schisandra chinensis (10 g/kg) equivalent to schizandrin (17.3 mg/kg). The result demonstrated that the oral bioavailability of schizandrin was approximately 15.56 ± 10.47% in rats, however the oral bioavailability of herbal extract was higher than single compound. The method was successfully applied to the pharmacokinetic study of pure schizandrin after oral administration of its pharmaceutical industry products in rats.

Deoxyschizandrin Loaded Liposomes on the Suppression Lipid Accumulation in 3T3-L1 Adipocytes.

Deoxyschizandrin (DS) is a bioactive benzocyclooctadiene lignan found in the fruit of Schisandra chinensis. However, poor bioavailability and non-specificity of DS frequently caused low therapeutic efficacy. In the present study, DS-liposome (DS-lipo) was implemented to enhance the hepatic targeting and inhibition effects on adipocyte differentiation in 3T3-L1 cells. The formulations enabled encapsulation of as much as 24.14% DS. The DS-lipo prepared was about 73.08 nm, as measured by laser light scattering (LLS) morphology. In the visual field of a scanning electron microscope (SEM), the liposomes were spherical with similar size and uniform dispersion. Fluorescence live imaging study exhibited hepatic targeting of liposomes in vivo. Furthermore, High-Content Analysis (HCS) imaging microassay analyses revealed DS-lipo and DS reduced cytoplasmic lipid droplet in 3T3-L1 adipocytes, with the IC50 value of 8.68 µM and 31.08 µM, respectively. The lipid droplet accumulation inhibition rate of 10 µM DS-lipo was above 90%, which was even superior to the effect of 30 µM DS solution. The current findings suggest that DS-lipo was a therapeutic strategy for alleviating lipid-associated diseases and nonalcoholic fatty liver disease (NAFLD).

Bioanalytical assay development and validation for simultaneous quantification of five schisandra lignans in rat primary hepatocytes based on LC-MS/MS: application to a real-time uptake study for Schisandra Lignan Extract.

Schisandra lignans, mainly including schizandrol A, schizandrol B, schisantherin A, schizandrin A, schizandrin B, etc., are the major active ingredients of Schisandra chinensis. In the present study, a robust liquid chromatography-tandem mass spectrometric (LC-MS/MS) method was developed for the simultaneous quantification of schisandra lignans in rat primary hepatocytes. Lovastatin was used as an internal standard, and chromatographic separation was achieved on a Shimadzu C18 column with a gradient elution at the flow rate of 0.2 mL/min. All of the analytes were detected in multiple reaction monitoring mode with positive electrospray ionization since the sodium adduct ion [M + Na]+ was observed as the most intensive peak in the MS spectrum. For schizandrol A, schisantherin A and schizandrin A, the dynamic range was within 2-1000 ng/mg protein, and the linear range of schizandrol B and schizandrin B was from 5 to 1000 ng/mg protein. The intra- and inter-day precision was <15% and the accuracy (relative error) ranged from -15 to 15%. No significant variation was observed in the stability tests. The validated method was then successfully applied to the time-dependent uptake study for the Schisandra Lignan Extract in rat primary hepatocytes.

Simultaneous determination of wogonin, oroxylin a, schisandrin, paeoniflorin and emodin in rat serum by HPLC-MS/MS and application to pharmacokinetic studies.

Wogonin and oroxylin A in Scutellariae Radix, schisandrin in Chinensis Fructus, paeoniflorin in Moutan Cortex and emodin in Polygoni Cuspidate Rhizome et Radix are anti-inflammatory active compounds. A method for simultaneous determination of the five compounds in rat was developed and validated using high-performance liquid chromatography with tandem mass spectrometry (HPLC-MS/MS). The separation was performed on a Symmetry C18 column (4.6 × 50 mm, 3.5 µm) with acetonitrile and 0.1% formic acid aqueous solution as the mobile phases. The detection was performed using multiple-reaction monitoring with electrospray ionization source in positive-negative ion mode. The calibration curves showed good linearity (r ≥ 0.9955). The lower limit of quantification (LLOQ) was 5 ng/mL for wogonin and schisandrin, 10 ng/mL for oroxylin A and emodin, and 15 ng/mL for paeoniflorin, respectively. The relative standard deviations of intraday and interday precisions were <11.49 and 14.28%, respectively. The extraction recoveries and matrix effects were acceptable. The analytes were stable under the experiment conditions. The validated method has been successfully applied to pharmacokinetic studies of the five compounds in rats after oral administration of Hu-gan-kan-kang-yuan capsule. This paper would be a valuable reference for pharmacokinetic studies of Chinese medicine preparations containing the five compounds.

Pharmacokinetics and protein binding of MPT0B292.

MPT0B292 was identified through screening of compounds able selectively to acetylate α-tubulins in cells and it exhibited potent anti-tumor, anti-angiogenesis and anti-metastatic effects in vitro and in vivo. Because of its poor water solubility, MPT0B292 is difficult to formulate with conventional approaches and hence difficulties are experienced in research practices. MPT0B292 was mixed with albumin in an aqueous solvent to form drug albumin nanoparticles with a size range around 333 nm. Unbound fractions of these nanoparticles were investigated in different or the same albumin concentration solutions. Unlike most drugs, the binding of MPT0B292 in human serum albumin increased with increasing drug concentrations. An analytical method was also developed and validated to determine MPT0B292 in rat plasma. This analytical method was applied successfully to the intravenous pharmacokinetic study of MPT0B292 in rats. A single dose study was regularly done to characterize the pharmacokinetic properties of the drug. Additionally, a novel i.v. infusion study was carried out to verify the extraction ratio of MPT0B292. The pharmacokinetic analysis revealed that MPT0B292 was a high extraction ratio drug with high systemic clearance, a high volume of distribution and a short half-life in rats. Copyright © 2017 John Wiley & Sons, Ltd.