Anil Kumar Dwivedi

Inhalable Microparticles Containing Drug Combinations to Target Alveolar Macrophages for Treatment of Pulmonary Tuberculosis

AbstractPurpose: Drug therapy of tuberculosis (TB) requires long-term oral administration of multiple drugs for curing as well as preventing and/or combating multi-drug resistance. Persistent, high blood levels of antitubercular drugs resulting from prolonged oral administration of anti-TB drugs may be neither necessary nor sufficient to kill mycobacteria residing in macrophages (Mφ). Inhalable biodegradable microparticles containing two of the first-line anti-TB drugs, isoniazid (H), and rifampicin (R), were prepared and tested for (i) phagocytosis by mouse Mφ, (ii) administration as a dry powder inhalation to rats, and (iii) targeting alveolar Mφ with high drug doses when administered to rats. Methods:poly(D-L lactic acid) microparticles were prepared by emulsion methods and their drug content and size distribution determined. These were tested for uptake by murine Mφ in culture and resultant intracellular drug concentrations determined by high performance thin-layer chromatography (HPTLC). Rats were administered an inhalation of microparticles using an inhalation chamber developed in the lab. The extent of microparticle delivery in vivo was examined by flow-cytometry. Drug concentrations in the blood and in alveolar Mφ were estimated by high-performance liquid chromatography after oral, vascular, intratracheal, and inhalation administration. Results: Inhalable microparticles could be prepared and were taken up by cultured Mφ. Large numbers of particles could be delivered to the bronchiopulmonary system through a 2-min exposure to fluidized particles. The intracellular drug concentrations resulting from vascular delivery of soluble drugs were found to be lower than those resulting from particle inhalation. Conclusions: Inhalable microparticles containing multiple anti-TB drugs offer promises of dose and dosing-frequency reduction, toxicity alleviation, and targeting Mφ-resident persistent mycobacteria.

Kaempferol has osteogenic effect in ovariectomized adult Sprague–Dawley rats

Kaempferol (K), a flavonol, is known to have anti-osteoclastogenic effect. We here show that K, from 0.2 to 5.0 microM, increased mineralized nodules in rat primary osteoblasts. K also significantly attenuated adipocyte formation from bone marrow cells (BMCs). A single oral dose of 1 mg/kg body weight of K in Sprague-Dawley (180-200 g) rats resulted in a peak serum level of 2.04+/-0.8 nM in 30 min (Tmax), suggesting its rapid absorption. The Cmax of K in bone marrow was 0.684 nM after 90 min. Rats were ovariectomized (OVx) along with sham-operated rats and left for 4 weeks. Daily oral administration of K (5 mg/kg body weight) was then started to one group of OVx rats, and continued for 10 weeks. K levels were found to be 0.311 and 0.838 nM at the end of 4 and 10 weeks, respectively. K exhibited no estrogenicity at the uterine level. The K-treated group exhibited significantly higher bone mineral density (BMD) in the trabecular regions (femur neck, proximal tibia and vertebrae) and lower serum ALP (bone turnover marker) compared with the OVx rats. The compressive energy of the vertebrae was significantly higher in the OVx+K-treated group compared with the OVx group. K treatment of OVx rats resulted in the increase in osteoprogenitor cells as well as inhibition of adipocyte differentiation from BMCs compared with the OVx group. Together we show that K is non-estrogenic in vivo and exerts bone anabolic activity with attendant inhibition of bone marrow adipogenesis.

Effects of Egb 761 on bone mineral density, bone microstructure, and osteoblast function: Possible roles of quercetin and kaempferol.

The effects of standardized and concentrated extract of Ginkgo biloba, Egb 761, were studied on estrogen deficiency-induced bone loss in ovariectomized (OVx) rats rendered osteopenic. Upon osteopenia development, Egb 761 was administered at a dose of 100mgkg(?1)day(?1) by oral gavage to OVx rats whereas control group received vehicle. Following 5 weeks of treatment, the OVx+Egb 761 group (n=12) of rats exhibited significantly higher whole body BMD and lower bone turnover markers (serum alkaline phosphatase and osteocalcin) than OVx rats that were given vehicle (n=12). BMD levels in excised bones were also found to be higher in both trabecular (most robustly in lumbar vertebrae) and cortical bones of OVx+Egb 761 compared with OVx+vehicle group. Egb 761 did not exhibit estrogen agonistic activity at the uterine level. Microcomputed tomography demonstrated that OVx+Egb 761 group had better bone microarchitectural parameters compared with OVx+vehicle group. Moreover, OVx+Egb 761 group had higher femoral mRNA levels of osterix, type I collagen and osteocalcin compared with OVx+vehicle group. Determination of levels of three flavonoids of Egb 761, that are known to have bone conserving property, in serum and bone marrow suggests that kaempferol and quercetin, and not rutin, likely mediate the beneficial actions observed with Egb 761 treatment. These results show for the first time that oral administration of Egb 761 restores bone mass in aged OVx rats.

Protective effect of fruits of Morinda citrifolia L. on scopolamine induced memory impairment in mice: A behavioral, biochemical and cerebral blood flow study

ETHNOPHARMACOLOGICAL RELEVANCE Noni (Morinda citrifolia L.) is widely used for different illnesses including CNS disorders. Recently Noni has been reported to prevent amyloid beta induced memory impairment in mice. However, the influence of Noni on cholinergic system has not been explored so far. Therefore, present study was designed to investigate effect of Noni fruit on memory, cerebral blood flow (CBF), oxidative stress and acetylcholinesterase (AChE) activity in scopolamine induced amnesia model. MATERIALS AND METHODS Mice were orally treated with ethanolic extract of Noni fruit and chloroform, ethyl acetate and butanol fractions of ethanolic extract for three days. Scopolamine was administered 5 min prior to acquisition trial and memory function was evaluated by passive avoidance test. CBF was measured by laser doppler flowmetry. AChE activity and oxidative stress parameters were estimated in mice brain at the end of behavioral studies. Further, effect of ethanolic extract and its fractions (5-400 μg/ml) on AChE activity was measured in vitro. RESULTS Scopolamine caused memory impairment along with reduced CBF, increased AChE activity and oxidative stress in mice brain. Ethanolic extract of Noni fruits and its chloroform and ethyl acetate fractions significantly improved memory and CBF. However, butanol fraction had no effect. Further, increased oxidative stress and AChE activity following scopolamine was significantly attenuated by ethanolic extract of Noni and its fractions. Moreover ethanolic extract and its fractions showed dose dependent inhibition of AChE activity in vitro. CONCLUSION These observations suggest that Noni may be useful in memory impairment due to its effect on CBF, AChE and oxidative stress.

Mechanism of action of some acrylophenones, quinolines and dithiocarbamate as potent, non-detergent spermicidal agents

Some suitably substituted acrylophenones, quinolines and dithiocarbamate were synthesized as new generation, non-detergent spermicides and were studied for their mechanism of action in comparison with various known spermicides belonging to several different classes of chemical compound. Nonoxynol-9, benzalkonium chloride, Sapindus saponins, verapamil, emetine and tartaric acid were used as reference molecules to study the effect of new spermicides on human sperm motility parameters (using computer-assisted semen analyzer), plasma membrane integrity, lipid peroxidation and defense system against reactive oxygen species (ROS). Results have indicated that sperm plasma membrane remains the primary site of action of most of the spermicides, though the effect may be predominantly on the physiological integrity rather than the structural integrity in case of the new compounds. Lipid peroxidation may play an important role in disrupting sperm membrane physiology that may or may not be accompanied with a detrimental effect on the defense system of the human spermatozoa against the ROS.

Pharmacokinetics study of arteether loaded solid lipid nanoparticles: an improved oral bioavailability in rats.

Arteether (ART), an artemisinin derivative, is a life saving drug for multiple drug resistant malaria. It has a deliverance effect in Falciparum malaria and cerebral malaria. We have prepared solid lipid nanoparticles (SLN) by high pressure homogenization (HPH) technique. ART-loaded SLN (ART-SLN) has been produced reproducibly with homogeneous particle size. ART-SLN was characterized for their size measured by Zetasizer Nano-ZS, Malvern, UK and by high resolution transmission electron microscopy (HR-TEM) and which was found to be 100 ± 11.2 nm. The maximum percentage entrapment efficiency (%EE) determined with the high-performance liquid chromatography (HPLC) has been found to be 69 ± 4.2% in ART-SLN-3. The release pattern from ART-SLN revealed that the release of ART is slow but time-dependent manner, which is desirable as it will help to protect the acid degradation of ART in stomach. The percentage cytotoxicity of blank SLN has been found within the acceptable range. The pharmacokinetics results indicated that ART-SLN-3 absorption has been significantly enhanced in comparison to ART in aqueous suspension and ART in ground nut oil (GNO) in rats. The % relative bioavailability (RB%) of ART-SLN to the ART in GNO and ART in aqueous suspension in rats was 169.99% and 7461%, respectively which was found to be significantly high in both the cases. From the results, it can be concluded that ART-SLN offers a new approach to improve the oral bioavailability of ART.

A naturally occurring naringenin derivative exerts potent bone anabolic effects by mimicking oestrogen action on osteoblasts

BACKGROUND AND PURPOSE Naringenin and its derivatives have been assessed in bone health for their oestrogen‐‘like’ effects but low bioavailability impedes clinical potential. This study was aimed at finding a potent form of naringenin with osteogenic action.

HPLC and HPTLC assays for the antimalarial agents Chloroquine, Primaquine and Bulaquine

A combination Kit for antirelapse treatment of P. vivax malaria, consisting of Chloroquine phosphate tablets and Bulaquine capsules has been recently developed, and marketed under the trade name Aablaquine. Bulaquine is prepared from Primaquine. Several methods of analysis are reported for each drug separately as well two drugs in combination but no method for simultaneous estimation of these three drugs is known. Therefore, the present study was undertaken to develop a sensitive and reproducible high performance liquid chromatographic as well as high performance thin layer chromatographic assay method for the simultaneous estimation of Chloroquine, Primaquine and Bulaquine.

Chitosan-based macrophage-mediated drug targeting for the treatment of experimental visceral leishmaniasis

The potential of chitosan microparticles as a carrier of doxorubicin for the treatment of visceral leishmaniasis was evaluated by macrophage-mediated drug targeting approach. Cationic charge of doxorubicin was masked by complexing it with dextran sulphate (a poly anion) in order to facilitate its incorporation into cationic chitosan microparticles. Prior to in vitro and in vivo studies, characterization studies were carried out systematically: particle size (∼1.049 µm), surface morphology (fluorescence microscopy – spherical structured microparticles), Fourier transform infrared spectroscopy (to characterize effective cross-linking) and differential scanning calorimetry. In vitro studies were carried out in J774.1 in order to check the effective endocytotic uptake of microparticles by macrophages. In vivo studies were conducted in Syrian golden hamsters as per well-established protocols and the results drawn from in vivo studies displayed substantial reduction in leishmanial parasite load for doxorubicin-encapsulated chitosan microparticles: ∼78.2 ± 10.4%, when compared to the control (free doxorubicin): 33.3 ± 2.4%.

Design and synthesis of ERα/ERβ selective coumarin and chromene derivatives as potential anti-breast cancer and anti-osteoporotic agents

Several new coumarin and chromene prototype derivatives have been synthesised and evaluated for their ERα and ERβ selective activity. Coumarin prototype compounds 18 & 19 were found to be ERα selective and the most active, exhibiting potential antiproliferative activity against both ER +ve & ER −ve breast cancer cell lines. The surprise finding of the series, however, are the novel prototype III chromenes 45 & 46, with aroyl substitution at the 6th position. Both the compounds have shown potent antiproliferative activity against both the breast cancer cell lines, promote alkaline phosphatase activity, enhance osteoblast mineralization in vitro, significantly decrease ERE–ERα dependent transactivation and induce ERβ activity. This specific upregulation of ERβ isoform activity of compound 45 may be responsible for the antiosteoporotic activity at picomolar concentration. In addition, both the compounds were also devoid of any estrogenic activity, which correlates to their antiestrogenic behaviour in the two breast cancer cell lines. Assessment of selectivity using specific SiRNAs for ERα and ERβ revealed that most of the compounds showed ERα and ERβ-mediated action, except compound 28, which showed selectivity to ERα only. Computational docking analysis of active compounds 18 and 45 was conducted to correlate the interaction with the two receptors and it was found that the docked conformations of the coumarin prototype, compound 18 at ERα and ERβ active sites were more or less superimposable on each other. However, the unique orientation of the aminoalkoxy side chain of novel chromene (prototype III) compound 45 in the ERβ binding cavity may be responsible for its potential biological response.