Quantification of IDP-73152, a novel antibiotic, in plasma from mice, rats and humans using an ultra-high performance liquid chromatography/tandem mass spectrometry method for use in pharmacokinetic studies.

IDP-73152, a novel inhibitor of a bacterial peptide deformylase, was recently approved as a new, investigational drug in Korea for the clinical management of infections caused by Gram positive bacteria. The objective of this study was to develop/validate a simple and robust analytical method for the determination of IDP-73152 in plasma samples from rodents and humans, and to assess the feasibility of the assay for use in pharmacokinetic studies using animal models. Plasma samples were processed using a standard method for protein precipitation and an aliquot of the extract then injected onto an UHPLC-MS/MS system. The drug and IDP-117293, an internal standard, were analyzed in the positive ion-mode by electrospray ionization and quantified by monitoring the transition at m/z 555.2→245.2 for IDP-73152 and 563.3→253.1 for the internal standard, respectively. The lower and upper limit of the assay was determined to be 5 and 10000ng/ml, respectively, with an acceptable linearity (R>0.999) in the response-concentration relationship. Validation parameters, including accuracy, precision, dilution, recovery, matrix effect and stability were found to be within the acceptable ranges recommended by the assay validation guidelines of the United States FDA. The method was successfully applied to the quantification of IDP-73152 in plasma from mice/rats that had received a single oral administration of 80mg/kg IDP-73152, in the form of the mesylate salt. These findings suggest that the validated assay can be used in preclinical and clinical pharmacokinetic studies of IDP-73152.

Identification of novel aminopiperidine derivatives for antibacterial activity against Gram-positive bacteria.

We have previously reported amidopiperidine derivatives as a novel peptide deformylase (PDF) inhibitor and evaluated its antibacterial activity against Gram-positive bacteria, but poor pharmacokinetic profiles have resulted in low efficacy in in vivo mouse models. In order to overcome these weaknesses, we newly synthesized aminopiperidine derivatives with remarkable antimicrobial properties and oral bioavailability, and also identified their in vivo efficacy against methicillin-resistant Staphylococcus aureus (MRSA), vancomycin-resistant Enterococcus (VRE) and penicillin-resistant Streptococcus pneumoniae (PRSP).

Formulation of a modified-release pregabalin tablet using hot-melt coating with glyceryl behenate.

A modified-release (MR) tablet of the anti-anxiety drug pregabalin (PRE) was prepared by hot-melt coating PRE with glyceryl behenate (GB) as a release retardant and compressing to form a matrix with microcrystalline cellulose (MCC) as a hydrophilic diluent. GB-coated PRE had a size in the range of 177-290 µm with good to acceptable flowability. Tablet hardness decreased slightly as GB content increased. PRE release from the tablet matrices was successfully modified by altering the ratio of MCC and GB, and it was found that dissolution- or diffusion-controlled release depended on the amount of GB used. Drug release was pH-independent. An accelerated stability test on the most promising MR tablet at 40°C and 75% relative humidity for 6 months showed no significant changes in PRE content, and the occurrence of total impurities--including PRE-lactam--was within acceptable limits. After oral administration of the selected MR tablet or a commercial IR capsule (Lyrica) to healthy human volunteers, pharmacokinetic parameters including Tmax, Cmax, AUC0-24, and T1/2 were compared. The confidence interval of AUC0-24 was within the adequate range, but that of Cmax was inadequate. This study demonstrated the potential use of GB for PRE-containing MR formulations.

Bioconversion of vitamin D3 to calcifediol by using resting cells of Pseudonocardia sp.

OBJECTIVES: Resting cells of Pseudonocardia sp. KCTC 1029BP were used for the bioconversion of vitamin D3 to calcifediol which is widely used to treat osteomalacia and is industrially produced by chemical synthesis. RESULTS: To obtain the maximum bioconversion yield of calcifediol by the microbial conversion of vitamin D3, a two-step optimization process was used, including the Plackett-Burman and the central composite designs. Six variables, namely agitation speed, aeration rate, resting cell concentration, vitamin D3 concentration, temperature, and pH, were monitored. Of these, aeration rate, resting cell concentration, and temperature were selected as key variables for calcifediol production and were optimized using the central composite design. Optimal bioconversion conditions obtained were as follows: aeration rate of 0.2 vvm, resting cell concentration of 4.7% w/v, and temperature of 33 °C. CONCLUSION: Using the optimal conditions, 356 mg calcifediol l(-1) was obtained with a bioconversion yield of 59.4% in a 75 l fermentor. These are the highest values reported to date.

Whole cell bioconversion of vitamin D3 to calcitriol using Pseudonocardia sp. KCTC 1029BP.

Calcitriol is an important drug used for treating osteoporosis, which can be produced from vitamin D3. The current method of producing calcitriol from vitamin D3 during cultivation of microbial cells results in low yields of calcitriol and high purification costs. Therefore, in this study, the steps of cell cultivation and bioconversion of vitamin D3 to calcitriol were separated. Cells of Pseudonocardia sp. KCTC 1029BP were utilized as a whole cell catalyst to produce a high level and yield of calcitriol from vitamin D3. In addition, the effects of bioconversion buffers, cyclodextrins, and metal salts on the production of calcitriol were comparatively examined and selected for incorporation in the bioconversion medium, and their compositions were statistically optimized. The optimal bioconversion medium was determined as consisting of 15 mM Trizma base, 25 mM sodium succinate, 2 mM MgSO4, 0.08% ß-cyclodextrin, 0.1% NaCl, 0.2% K2HPO4, and 0.03% MnCl2. Using this optimal bioconversion medium, 61.87 mg/L of calcitriol, corresponding to a 30.94% mass yield from vitamin D3, was produced in a 75-L fermentor after 9 days. This calcitriol yield was 3.6 times higher than that obtained using a bioconversion medium lacking ß-cyclodextrin, NaCl, K2HPO4, and MnCl2. In conclusion, utilizing whole cells of Pseudonocardia sp. KCTC 1029BP together with the optimal bioconversion medium markedly enhanced the production of calcitriol from vitamin D3.

Effects of Polygonatum sibiricum rhizome ethanol extract in high-fat diet-fed mice.

CONTEXT: The rhizome of Polygonatum sibiricum Redoute (Liliaceae) has long been used to treat diabetes-associated complications. However, the pharmacological mechanism of P. sibiricum on metabolic disorders is not clear. OBJECTIVE: This study investigates the effect of an ethanol extract of P. sibiricum rhizomes (designated ID1216) on obesity conditions including weight loss in high-fat (HF) diet-fed mice and explores the potential underlying mechanisms. METHODS: To identify the metabolic impact of the P. sibiricum rhizome extract, HF diet-fed mice were administered ID1216 orally at doses of 250 and 1000 mg/kg/d for 10 weeks, and various factors related to metabolic syndrome were analyzed. We also examined the effects of ID1216 on the expression of genes involved in adipogenesis and lipolysis in 3T3-L1 cells, as well as genes associated with energy homeostasis in C2C12 myocytes. RESULTS: ID1216 administration led to significant decreases in body weight gain (37.5%), lipid accumulation in adipose tissues (52.8%), and the levels of plasma triglycerides (26.4%) and free fatty acids (28.1%) at a dose of 250 mg/kg/d, compared with the vehicle-treated group, as well as improved insulin resistance. In addition, ID1216 was found to regulate the expression of genes related to adipogenesis and fatty acid oxidation in 3T3-L1 cells and enhance the expression of genes that modulate energy homeostasis in C2C12 myocytes. CONCLUSIONS: ID1216 may be a promising therapeutic agent for improving obesity conditions through the sirtuin-1 and peroxisome proliferator-activated receptor γ coactivator-1α pathway.

ID1201, the ethanolic extract of the fruit of Melia toosendan ameliorates impairments in spatial learning and reduces levels of amyloid beta in 5XFAD mice.

A previous study has demonstrated the anti-amyloidogenic effects of the ethanolic extract of Meliae Fructus (ID1201) using cell lines with stably expressed human Swedish mutant APP695 and ß-secretase 1, and 5Xfamilial AD (FAD) mice carrying five mutations. Here, we investigated the effects of ID1201 on cognitive impairment in 5XFAD mice. Daily administration of ID1201 was commenced at 3 months of age and continued for 3 months. Mice were serially trained in cued/response and place/spatial training tasks in the Morris water maze. After this training, testing for strategy preference was conducted. Non-transgenic control mice with vehicle treatment, vehicle-treated 5XFAD, and ID1201-treated 5XFAD mice showed equivalent performance in cued/response training. However, as training progressed to the subsequent place/spatial learning, vehicle-treated control and ID1201-treated 5XFAD mice differed significantly from vehicle-treated 5XFAD mice in measures of spatial learning (search error and adaptive spatial learning strategy). In the strategy preference test that followed, control mice preferred a place/spatial strategy relative to vehicle-treated 5XFAD mice, but differences between ID1201-treated 5XFAD mice and vehicle-treated 5XFAD mice were not significant. Additionally, ID1201 treatment reduced hippocampal levels of insoluble Aß42 and increased cortical levels of soluble amyloid precursor protein α. These results indicate that ID1201 may possess potential as a therapeutic agent for Alzheimer's disease by decreasing Aß deposits.

In vitro virucidal effect of mouthrinse containing C31G on seasonal influenza viruses.

C31G is a potent antimicrobial agent and can disrupt the microbial membrane by the alkyl portion of the molecule. The objective of this study was to evaluate the virucidal effectiveness of C31G and mouthrinse containing C31G (Sense-Time) on seasonal influenza viruses. Evaluation of the virucidal activity against influenza viruses was performed with end-point titration in 10-day-old chicken embryos and Madin-Darby canine kidney cells. In vitro studies demonstrated that C31G and Sense-Time inhibited the growth of seasonal influenza viruses even in the presence of 5% organic material. Gargling with C31G or Sense-Time would enhance oropharyngeal hygiene, which would be helpful for reducing influenza transmission.

Anti-amyloidogenic effects of ID1201, the ethanolic extract of the fruits of Melia toosendan, through activation of the phosphatidylinositol 3-kinase/Akt pathway.

Amyloid beta (Aß) peptides, which are generated from amyloid precursor protein (APP), are thought to play a major role in the pathogenesis of Alzheimer's disease (AD). This study investigated the anti-amyloidogenic effects of the ethanolic extract of Meliae Fructus (ID1201) using human embryonic kidney 293 cells with stably expressed human wild-type or Swedish mutant APP695 and ß-secretase 1. ID1201 treatment enhanced the non-amyloidogenic metabolism of APP; increases in soluble APPα levels and decreases in soluble APPß and Aß levels resulted from the α-secretase activation through the phosphatidylinositol 3-kinase (PI3K)/Akt pathway. In addition, ID1201-treated 5×familial AD (FAD) mice with 5 mutations in APP and presenilin 1 showed reduced levels of Aß and amyloid plaques in the brain relative to those of 5×FAD mice with vehicle treatments. These results indicate that ID1201 possesses anti-amyloidogenic effects via the activation of the PI3K/Akt pathway, suggesting that it is a potential therapeutic agent for AD.

Optimization of medium components for high-molecular-weight hyaluronic acid production by Streptococcus sp. ID9102 via a statistical approach.

Hyaluronic acid (HA), linear high-molecular-weight glycosaminoglycan produced from Streptococcus sp., has raised interest in the medical and cosmetics industries because of the various biological functions of HA. In this paper, we report on the optimization of medium components for HA production in Streptococcus sp. ID9102 (KCTC 11935BP) by two-step optimization (one-factor-at-a-time and taguchi orthogonal array design). In the first step, medium components, such as carbon, nitrogen, phosphate, and mineral sources, were selected for HA production in Streptococcus sp. ID9102 (KCTC 11935BP) using the one-factor-at-a-time method. In the second step, the concentration of the selected medium components was optimized using taguchi orthogonal array design. The design for medium optimization was developed and analyzed using MINITAB 14 software. In addition, the effect of amino acid and organic acid, such as glutamine, glutamate, and oxalic acid, was studied for HA production in Streptococcus sp. ID9102 (KCTC 11935BP). Through these processes, the optimum medium comprising 4% glucose, 0.75% yeast extract, 1.0% casein peptone, 0.25% K(2)HPO(4), 0.05% MgCl(2), 0.5% NaCl, 0.04% glutamine, 0.06% glutamate, and 0.02% oxalic acid was determined. We were able to produce HA with a molecular weight of 5.9 x 10(6) at a productivity of 6.94 g/l on pilot scale fermentation.

Production of teicoplanin from Actinoplanes teichomyceticus ID9303 by adding proline.

We evaluated the influence of amino acids in improving teicoplanin productivity. Arginine, lysine, and proline were selected for better productivity among 20 amino acids in Erlenmeyer flasks. Proline was finally chosen as the additive for maximum teicoplanin productivity in a 5-liter fermenter. We obtained the highest teicoplanin productivity, 3.12 g/l, on the eighth d in a 75-liter pilot fermenter.

Comparative QSAR studies on peptide deformylase inhibitors.

Comparative quantitative structure-activity relationship (QSAR) analyses of peptide deformylase (PDF) inhibitors were performed with a series of previously published (British Biotech Pharmaceuticals, Oxford, UK) reverse hydroxamate derivatives having antibacterial activity against Escherichia coli PDF, using 2D and 3D QSAR methods, comparative molecular field analysis (CoMFA), comparative molecular similarity indices analysis (CoMSIA), and hologram QSAR (HQSAR). Statistically reliable models with good predictive power were generated from all three methods (CoMFA r (2) = 0.957, q (2) = 0.569; CoMSIA r (2) = 0.924, q (2) = 0.520; HQSAR r (2) = 0.860, q (2) = 0.578). The predictive capability of these models was validated by a set of compounds that were not included in the training set. The models based on CoMFA and CoMSIA gave satisfactory predictive r (2) values of 0.687 and 0.505, respectively. The model derived from the HQSAR method showed a low predictability of 0.178 for the test set. In this study, 3D prediction models showed better predictive power than 2D models for the test set. This might be because 3D information is more important in the case of datasets containing compounds with similar skeletons. Superimposition of CoMFA contour maps in the active site of the PDF crystal structure showed a meaningful correlation between receptor-ligand binding and biological activity. The final QSAR models, along with information gathered from 3D contour and 2D contribution maps, could be useful for the design of novel active inhibitors of PDF.

Inhibition of lipopolysaccharide and interferon-gamma-induced expression of inducible nitric oxide synthase and tumor necrosis factor-alpha by Lithospermi radix in mouse peritoneal macrophages.

Lithospermi radix (LR, root of Lithospermum erythrorhizon Siebold. et Zuccarinii) has been used to treat various conditions, such as septic shock, eczema and burns. In this study, the effect of LR on lipopolysaccharide (LPS) and recombinant interferon-gamma (rIFN-gamma)-induced production of nitric oxide (NO) and tumor necrosis factor (TNF)-alpha were examined using mouse peritoneal macrophages. At 0.01-1 mg/ml, LR inhibited the LPS/rIFN-gamma-induced expression of inducible nitric oxide synthase (iNOS) and TNF-alpha release. To clarify the mechanism involved, the effect of LR on the activation of nuclear factor (NF)-kappaB was examined. The LPS/rIFN-gamma-induced activation of NF-kappaB was almost completely blocked by LR at 1mg/ml without cytotoxicity. These findings demonstrate that the inhibition of the LPS/rIFN-gamma-induced production of NO and TNF-alpha by LR involves the inhibition of NF-kappaB activation.