Development and validation of a reversed-phase HPLC method for quantification of 1'-acetoxychavicol acetate content in a nanostructured lipid carrier formulation

Abstract 1'-acetoxychavicol acetate (ACA)-loaded nanostructured lipid carriers (NLCs) were formulated for prostate cancer therapy and to determine the optimal therapeutic dose, we developed a rapid, specific, and accurate reversed-phase high-performance liquid chromatography (RP-HPLC) method to quantify the ACA content in NLCs. The method was validated according to International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use (ICH) guidelines. Chromatographic separation of ACA from the lipid components was performed with an Agilent 1220 Infinity LC system and ultraviolet detector using an Agilent Poroshell C18 column (4.6 x 250.0 mm). The mobile phase consisted of acetonitrile and water (80:20 [v/v]) with a flow rate of 0.8 mL/min in isocratic mode. Linearity of the standard curve was assessed at an ACA concentration range of 5-200 µg/mL, and a 1/x weighted linear regression was adopted for the calibration curve. The calculated limits of detection and quantification were 0.59 µg/mL and 1.79 µg/mL, respectively. The mean percent recovery of ACA was 100.02% (relative SD, 2%), and the coefficients of variation for intraday and interday assays were within the values required by the ICH. We also demonstrated robustness of the method by altering the mobile phase ratio and flow rate. Furthermore, we proved specificity of the method for ACA by comparing chromatograms of the blank NLC and ACA-NLC. Hence, we effectively used this validated method to determine the drug-loading capacity and entrapment efficiency of the NLCs.

Nanostructured lipid carrier containing hydroxymethylnitrofurazone: preparation, characterization and in vitro and in vivo evaluation of leishmanicidal activity / Carreador lipídico nanoestruturado contendo hidroximetilnitrofural: preparação, caracterização físico-química e avaliação in vitro e in vivo da atividade leishmanicida

Leishmaniasis, a neglected tropical disease (NTD), is a set of diseases caused by obligatory parasitic protozoa of the genus Leishmania. And it has cutaneous and visceral eishmaniasis as its main forms. Treatment includes pentavalent antimonials. These drugs have several disadvantages, such as the need for parenteral administration, use of high dosages, long duration of treatment, severe toxicity, resistance and variable efficacy. The candidate for hydroxymethylnitrofural drug (NFOH), a prodrug derived from nitrofural, showed high activity in cell cultures infected with Trypanosoma cruzi and less toxicity when compared to nitrofural. Due to its low solubility in water and reduced bioavailability, NFOH has failed the in vivo efficacy tests. Nanostructured drug delivery systems have the potential to overcome these challenges due to their evident advantages: greater therapeutic efficacy, less toxicity, modified drug release and increased gastrointestinal absorption of drugs with low water solubility. The objective of this project will be the preparation and evaluation of the physicochemical characteristics of a nanostructured lipid carrier containing hydroxymethylnitrofural (NLC-NFOH). The NFOH showed the highest solubility in Miglyol® 840 among the tested liquid lipids. For solid lipids, Gelucire® 50/13 and Precirol® ATO5 proved to be more suitable for the solubilization of NFOH. The optimized NLC-NFOH consisted of these three lipids. These lipids were selected using a quick Technobis Crystal 16TM methodology, microscopy and DSC. Different lipid selection tools provided scientific knowledge relevant to the development of NLC. The NLC-NFOH had an average z of 198.6 ± 5.4 nm, a PDI of 0.11 ± 0.01 and a zeta potential of -13.7 ± 0.7 mV. This study allowed a design space development approach of the first NLC-NFOH with the potential to treat leishmaniasis orally. The development of a sensitive bioanalytical method using HPLC and evaluation of some analytical figures of merit for the validation allowed the quantification of NFOH and NF. The bioanalytical method for analysis of NFOH and NF use Zorbax SB-C18, 5µm, (4.6x250mm) HPLC column. The mobile phase was consisted of acetonitrile:water (20:80 v/v) with flow rate of 1.2 ml/min, at UV detection of 370 nm. The linearity of NFOH and NF was found in the range 0.0253.0 µg/ml with a correlation coefficient of r > 0.98. The precision was 2.44 to 13.77% for NFOH and 2.61 to 18.42%; the accuracy was 2.66 to 14.28% for NFOH and 2.09 to 19.06% for NF. The method showed to be suitable for effectively evaluation of NFOH is serum. NLC-NFOH (2.8 mg/kg) was administered to animals by gavage, and the blocking flow of the chylomicrons model was performed with an intraperitoneal injection of cycloheximide. The presence of NFOH in serum was evaluated with and without cycloheximide. The cytotoxicity assay of NLC-NFOH and blank-NLC showed more than 90% viable cells at the maximum concentration used (2560 µM). NFOH and NF were detected at 1h after the gavage of DMSO-NFOH or NLC-NFOH, without the pretreatment with cycloheximide. The concentration found for DMSO-NFOH and NLC-NFOH were 0.0316 and 0.0291 µg/mL, respectively. The NLC presented the NFOH absorption by the lymphatic system, demonstrated by blocking chylomicrons flow

A leishmaniose, uma doença tropical negligenciada (DTN), é um conjunto de doenças causadas por protozoários parasitas obrigatórios do gênero Leishmania. E tem como formas principais a leishmaniose cutânea e visceral. O tratamento inclui antimoniais pentavalentes. Esses fármacos apresentam várias desvantagens, como necessidade de administração parenteral, uso de altas dosagens, longa duração do tratamento, toxicidade grave, resistência e eficácia variável. O candidato ao fármaco hidroximetilnitrofural (NFOH), um pró-fármaco derivado do nitrofural, apresentou alta atividade em culturas de células infectadas pelo Trypanosoma cruzi e menor toxicidade quando comparado ao nitrofural. Devido à sua baixa solubilidade em água e biodisponibilidade reduzida, o NFOH falhou nos testes de eficácia in vivo. Os sistemas nanoestruturados de liberação de fármacos têm potencial para superar esses desafios devido às suas vantagens evidentes: maior eficácia terapêutica, menor toxicidade, liberação modificada do fármaco e aumento da absorção gastrointestinal de fármacos com baixa solubilidade em água. O objetivo deste projeto será a preparação e avaliação das características físico-químicas de um carreador lipídico nanoestruturado contendo hidroximetilnitrofural (NLC-NFOH). O NFOH apresentou a maior solubilidade no Miglyol® 840 entre os lipídios líquidos testados. Para lipídios sólidos, Gelucire® 50/13 e Precirol® ATO5 se mostraram mais adequados para a solubilização de NFOH. O NLC-NFOH otimizado consistiu desses três lipídios. Esses lipídios foram selecionados usando Technobis Crystal 16TM, microscopia e DSC. Diferentes ferramentas de seleção de lipídios forneceram conhecimento científico relevante para o desenvolvimento de NLC. O NLC-NFOH teve z-average de 198,6 ± 5,4 nm, PDI de 0,11 ± 0,01 e potencial zeta de -13,7 ± 0,7 mV. Este estudo permitiu o desenvolvimento por abordagem de Design Space do primeiro NLC-NFOH com potencial para tratar a leishmaniose por via oral. O desenvolvimento de um VIII método bioanalítico sensível utilizando HPLC e a avaliação de algumas figuras analíticas de mérito para a validação permitiram a quantificação de NFOH e NF em soro. O método bioanalítico para análise de NFOH e NF usou coluna de HPLC Zorbax SB-C18, 5 µm, (4,6 x 250 mm). A fase móvel foi constituída por acetonitrila: água (20:80 v / v) com vazão de 1,2 ml / min, com detecção no UV de 370 nm. A linearidade de NFOH e NF foi encontrada na faixa de 0,0253,0 µg / ml com um coeficiente de correlação de r> 0,98. A precisão foi de 2,44 a 13,77% para NFOH e 2,61 a 18,42%; a precisão foi de 2,66 a 14,28% para NFOH e 2,09 a 19,06% para NF. O método mostrou-se adequado para avaliação efetiva do NFOH no soro. NLC-NFOH (2,8 mg / kg) foi administrado aos animais por gavagem, e o modelo de bloqueio do fluxo de quilomícrons foi realizado com injeção intraperitoneal de cicloheximida. A presença de NFOH no soro foi avaliada com e sem cicloheximida. O ensaio de citotoxicidade de NLC-NFOH e brancoNLC mostrou mais de 90% de células viáveis na concentração máxima utilizada (2560 µM). NFOH e NF foram detectados 1h após a gavagem de DMSO-NFOH ou NLC-NFOH, sem o pré-tratamento com cicloheximida. As concentrações encontradas para DMSO-NFOH e NLC-NFOH foram 0,0316 e 0,0291 µg / mL, respectivamente. O NLC apresentou a absorção do NFOH pelo sistema linfático, demonstrada pelo bloqueio do fluxo dos quilomícrons

Design and Antitumor Activity Evaluation of n-Dodecanol Modified Docetaxel Prodrug Nanostructured Lipid Carrier / 中国药学杂志

OBJECTIVE To design N dodecanol modified docetaxel(DTX) prodrug, prepare nanostructured lipid carrier(NLC) and investigate in vitro antitumor activity and in vivo pharmacodynamic. METHODS Nanostructured lipid carrier (DNLC) encapsulating n-dodecanol-modified DTX prodrug was prepared by ultrasonic method. The formulation was optimized by single-factor experiment and response surface optimization. The accumulated rates of DTX degraded from DNLC in different media was evaluated by high performance liquid chromatography (HPLC). The morphology of DNLC was observed by transmission electron microscopy (TEM). The particle size and PDI of DNLC were determined by Malvern particle size analyzer. The long-term stability of the preparations was investigated. In vitro cytotoxicity of DNLC was measured by MTT method. In vivo pharmacodynamics of DNLC were performed in 4T1 tumor xenograft balb/c mice using saline and DTX-Sol as control. RESULTS n-Dodecanol-modified DTX prodrug was synthesized and used to prepare DNLC. The optimal formulation was as following: mass ratio of emulsifier to co-emulsifier (Km) of 1∶3, solid-liquid lipid ratio of 1.43∶1, drug-lipid ratio of 1∶10, the emulsifier concentration of 60 mg•mL-1, the temperature of 70 ℃ and the stirring speed of 800 r•min-1. DNLC had a round appearance and a uniform spherical shape. And the particle size and PDI remained substantially stable within 30 d. The accumulated rates of DTX degraded from DNLC in PBS (pH 7.4), PBS (pH 7.4) containing 10 mmol•L-1 DTT and 10 mmol•L-1 H2O2 was (9.07±0.01)%, (21.52±0.35)% and (96.72±4.12)% at 24 h, respectively. After incubation of DTX-Sol and DNLC with 4T1 cells for 72 h, IC50 of DTX-Sol and DNLC were (1.2±0.2) and (13.2±4.3)nmol•L-1, respectively. The cytotoxicity of DTX-Sol group was stronger than that of DNLC group. At the end of the pharmacodynamics, the tumor volumes of the mice in saline, DTX-Sol and DNLC groups were (1 930.39±215.20), (1 013.64±138.65), and (765.16±177.43)mm3, respectively. And the change percentage of body weight in saline, DTX-Sol and DNLC groups were (-19.69±4.44)%, (-14.85±3.61)% and (-2.61±1.70)%. There were significant differences in tumor volume and body weight between the DNLC and DTX-Sol group (P<0.05). CONCLUSION The prepared DNLC shows good stability, redox sensitivity, obvious anti-tumor effect and lower toxicity. These RESULTS could provide a new experimental basis for the development of DTX prodrug loaded nano-drug delivery system.

Effect of ginsenoside Rg_3 nanostructured lipid carrier modified by pullulan on promoting absorption and its anti-tumor evaluation in vitro / 中国中药杂志

Ginsenoside Rg_3 is widely used in clinical practice as an anti-tumor adjuvant drug, but its application is limited due to its poor oral absorption. In this study, we intended to construct a ginsenoside Rg_3 nanostructured lipid carrier modified by the pullulan(PUL-Rg_3-NLC) to improve the adhesion properties of ginsenoside Rg_3, promote the drug uptake and improve the anti-tumor efficacy. PUL-Rg_3-NLC was characterized by morphology, particle size and Zeta potential. In vivo adhesion characteristics were evaluated by oral gavage tests, and the results were verified from multiple perspectives in combination with in vitro uptake behavior and in vitro pharmacodynamics. The results showed that PUL-Rg_3-NLC, with a particle size of(102±1.89) nm, was characterized by gastric adhesion and could be retained in gastric tissues for a long time, and its uptake by BGC-823 cells was promoted mainly through the pathway mediated by the caveolin-mediated endocytosis. In vitro MTT, cell apoptosis, wound-healing assay and invasion assay all showed some anti-tumor effects. Therefore, PUL-Rg_3-NLC can significantly promote the adhesion of Rg_3 in the stomach, promote the uptake of drugs by gastric cancer cells, and improve the anti-tumor effect. This study can provide some reference for the adjuvant treatment of gastric cancer.

Design And Development Of Nanostructured Lipid Carrier Containing Triamcinolone Acetonide

Objective: The aim of present study was to prepare nanostructured lipid carriers (NLCs) based Triamcinolone acetonide (TA). Methods: Nanostructured lipid carriers (NLCs) consisted of solid lipid and liquid lipid are a new type of lipid nanoparticles, prepared by using solvent diffusion and high pressure homogenization methods, which offer the advantage of improved drug loading capacity and release properties. Glyceryl monostearate selected as the solid lipid, capmul MCM C8 as the liquid lipid, polyvinyl Alcohol (PVA) as the surfactant. NLCs dispersion was characterized by particle size analysis, zeta potential, scanning electron microscopy (SEM), differential scanning calorimetry, and an in vitro release study. Results: Optimized NLCs loaded with TA were exhibited spherical shape with particle size 286.1 nm, polydispersity index 0.317, zeta potential-21.9 mV and entrapment efficiency 86.19% respectively. The result of differential scanning calorimetry (DSC) showed that drug was dispersed in NLCs in a crystalline state. In vitro release studies revealed that drug release of optimized batch was 8.34 % and 88.84% at 1h and 8h respectively. The release kinetics of the optimized NLCs best fitted the peppas-korsmeyer model. Furthermore, morphological investigations by SEM showed that optimized batch exhibit a spherical shape and a smooth surface. Conclusion: Thus, the results indicated that successfully prepared TA-loaded NLCs and could potentially be exploited as a carrier with improved drug loading capacity and sustained drug release. The present results demonstrated that these systems could be a promising platform for inflammatory diseases, in particular for psoriasis topical therapy.

Optimization of prescription process of cantharidin nanostructured lipid carrier by central composite design-response surface methodology / 中草药

Objective: To integrate the toxic component of cantharidin (CTD) into a novel nanostructured lipid carrier (NLC) and optimize the cantharidin nanostructured lipid carrier (CTD-NLC) formulation process to reduce the toxicity of CTD and enhance its targeting. Methods: CTD-NLC was prepared by emulsified ultrasonic dispersion method. The encapsulation efficiency was determined by dialysis method. The average particle size, particle size distribution (polydispersity index, PDI), Zeta potential, encapsulation efficiency, and drug loading were taken as indicators. Univariate investigation and central composite design-response surface methodology (CCD-RSM) were used to optimize the prescription process of CTD-NLC. Multivariate quadratic fitting was used to evaluate the model equation between indicators and factors. The fitted equation was analyzed by the variance analysis and the optimal prescription was predicted by the resonse surface. Results: The optimized CTD-NLC prescriptions were as follow: mass of total lipid was 453.66 mg, solid to liquid lipid ratio of 1:2, total stable dose of 16.9 mg/mL, ratio of Pluronic F68 to egg yolk lecithin (Lipoid E PC S) of 3.88:1, with ultrasound for 30 min (working 2 s, stopping 2 s). The prepared CTD-NLC was clear clarification in appearance with light blue opalescence, the average particle size was (85.99 ± 0.49) nm, PDI was 0.280 ± 0.002, Zeta potential was (-8.21 ± 0.24) mV, encapsulation efficiency was (98.57 ± 0.05)%, and drug loading was (0.65 ± 0.01)%. Conclusion: The fitting model established by CCD-RSM is accurate and reliable. The optimized CTD-NLC distribution is concentrated, with high encapsulation efficiency and good physical stability. It lays a foundation for the subsequent in vitro and in vivo studies of CTD-NLC.

Acute Toxicity Study of Intravenous Administration of Thymoquinone-Loaded Nanostructured Lipid Carrier (TQ-NLC) in Sprague Dawley Rats

@#Introduction: Thymoquinone (TQ), a bioactive compound from Nigella sativa is known for its various medicinal properties. Due to the low solubility of TQ, nanostructured lipid carrier (NLC) has been used as a delivery system to improve its efficacy. Nevertheless, the effect of TQ-NLC when administered intravenously is unclear. This study investigated the acute toxicity profile of intravenous administration of TQ-NLC in an in vivo model. Methods: Twelve female Sprague dawley rats were assigned randomly into two groups (n=6); a control and a treatment group that received normal saline and 25 mg/kg TQ-NLC, respectively, via intravenous injection. The rats were observed for 14 days for any alterations to their usual physical conditions such as behaviour and mortality, body weight, food intake, organ-to-body weight ratio, and haematological, biochemical and histopathological profile. Results: There were no significant changes (p>0.05) in the body weight, food intake, organ-to-body weight ratio, and haematological, biochemical and histopathological profile between TQ-NLC treatment and the control group. However, inflammation was observed at the site of injection on the rat’s tail. Conclusion: Intravenous administration of TQ-NLC (25 mg/kg) did not exert acute toxic effect in female Sprague dawley rats. The data can be used as a basis to further develop TQNLC as a potential therapeutic drug.

In vitro dissolution and anti-vitiligo activity of isopsoralen nanostructured lipid carriers / 中草药

Objective To evaluate the in vitro dissolution characteristic of IPRN-NLC and to study its effects on B16F10 cells proliferation, melanin synthesis, and tyrosinase activity. Methods The dynamic dialysis was employed to compare the in vitro dissolution of IPRN and IPRN-NLC; MTT assay was used to detect the proliferation of B16F10; The tyrosinase activity was determined by L-DOPA-oxidation; The melain content was determined by GENMED Cell Melanin Quantitative Assay Kit. Results The accumulation dissolution of IPRN-NLC was 67.31% within 72 h, which showed sustained release; While the dissolution of IPRN-suspension, IPRN-physical mixture, and IPRN-DMSO were 53.34%, 90.30%, and 98.67%, respectively. The IPRN-NLC could significantly promote the proliferation, tyrosinase activity and melanin content compared with IPRN DMSO groups (P < 0.05) at the same concentration. Conclusion IPRN-NLC could increase the solubility of the drug with sustained release, and showed good cell biology intermiscibility, which could significantly increase the effects on B16F10 cells.

Research on monoclonal antibody OX26 modified salvianolic acid B/baicalin nanostructured lipid carriers / 中草药

Objective To prepare, characterize, and study cellular uptake of transferrin receptor monoclonal antibody OX26 modified nanostructured lipid carrier loaded with salvianolic acid B and baicalin (Sal B/BA-NLC). Methods Sal B/BA-NLC was prepared by emulsification-solvent evaporation method. OX26 was thiolated with 2-iminothiolane hydrochloride and then conjugated to the surface of Sal B/BA-NLC. The morphology, particle size, Zeta potential, and encapsulation efficiency (EE) were evaluated for the physicochemical properties, and OX26 modified Sal B/BA-NLC was verified by differential scanning calorimetry (DSC) and nuclear magnetic resonance spectroscopy (NMR). Coumarin-6 (C6) was used as the fluorescent probe instead of baicalin and salvianolic acid B to prepare the formulations in cellular uptake study. The cellular uptake study was conducted by brain microvascular endothelial cells bEnd.3 using high content cell imaging analysis system. Results The prepared OX26 modified Sal B/BA-NLC had particle size of (27.50 ± 3.37) nm, PDI of 0.39 ± 0.04, and Zeta potential of (-7.06 ± 1.85) mV. The DSC and NMR results indicated that the drug was encapsulated in the nanostructured lipid carrier in an amorphous form. The results of cell uptake showed that the fluorescence intensities of the three solutions in bEnd.3 cells were: OX26-C6-NLC > C6-NLC > C6-SL. Conclusion The prepared OX26 modified Sal B/BA-NLC has smaller particle size, uniform distribution, and high EE. The OX26-modified NLC group had a higher intake than the solution group and the unmodified NLC group.

Optimization of Paeonol NLC Thermosensitive Gel Formulation by Box-Behnken Response Surface Methodology / 中国中医药信息杂志

Objective To optimize the formulation of paeonol nanostructured lipid carrier (NLC) thermosensitive in situ gel through Box-Behnken response surface method; To investigate its release properties in vitro to provide references for the study of transdermal drug delivery system. Methods Taking mass fraction of poloxamer 407 and poloxamer 188 as the factors, the gelling temperature as the index, the mathematical relationship between the gelling temperature and two factors was established by binomial model and multivariate linear regression model. The Box-Behnken response surface method was used to optimize the formulation of paeonol NLC thermosensitive in situ gel, and the in vitro release characteristics of the preparation was investigated. Results There was a credible quantitative relationship between the gelling temperature and the 2 factors, and the binomial model was more reliable than the multivariate linear model. The best prescriptions of paeonol NLC thermosensitive in situ gel were 22.90% poloxamer 407 and 3.34% poloxamer 188; gelling temperature was (33.4±0.1)℃, and the cumulative release amount of paeonol in situ gel in 24 h was 51.19%. Conclusion This method is suitable for the formulation optimization of paeonol NLC thermosensitive in-situ gel, and the established mathematical model has good predictability. The optimized formulation can provide references for the development of paeonol transdermal preparation.

Construction of artesunate nanoparticles modified by hyaluronic acid and cell-penetrating peptides and its inhibitory effect on cancer cells in vitro / 中国中药杂志

Hyaluronic acid (HA) and cell-penetrating peptide (CPP) R6H4-SA modified artesunate nanostructured lipid carrier (HA-R6H4-NLC/ART) for anti-tumor therapy was prepared. The physicochemical properties and in vitro drug release of HA-R6H4-NLC/ART were evaluated, and the uptake and cytotoxicity of liver cancer HepG2 cells were studied. The results showed that HA-R6H4-NLC/ART was spherical like in appearance, and the average particle size was about 160 nm. In vitro release experiments showed that the drug delivery system had sustained release characteristics. Cell results showed that, in slightly acidic environment, pH sensitive CPP R6H4-SA mediated cellular uptake of nanoparticles was significantly higher than that of non-sensitive peptide R8-SA. Meanwhile, HA-R6H4-NLC/ART had a targeting effect on HepG2 cells, and the HA receptor saturation experiment showed that the endocytosis of HA-R6H4-NLC/ART was mediated by the HA receptor on the cell surface. As compared with the unmodified or R6H4-SA single modified group, HA and R6H4-SA co-modified HA-R6H4-NLC/ART significantly improved the cell uptake and had a stronger anti-tumor effect under the conditions of the slightly acid environment and hyaluronidase degradation. The above results showed that hyaluronic acid and CPP R6H4-SA co-modified artesunate nanostructured lipid carrier, which can effectively identify and penetrate the tumor cell membrane into the cell, is a potentially efficient targeting delivery system for anti-tumor drugs.

Acute and chronic toxicity of 0.5% podophyllotoxin-loaded nanostructured lipid carriers to vaginal mucosa in rabbits and rats / 南方医科大学学报

OBJECTIVE@#To test the acute and chronic toxicity of topical application of 0.5% podophyllotoxin-loaded nanostructured lipid carriers (POD-NLC) to the vaginal mucosa.@*METHODS@#Twelve New Zealand rabbits were randomized into 3 groups and subjected to daily topical applications of normal saline (control group), 0.5% podophyllotoxin tincture (POD-T) or 0.5% POD-NLC on the vaginal mucosa for 10 consecutive days, and the pathological changes in the mucosa were graded using the Eckstein scoring system.The acute toxicity of POD-NLC was tested in 20 SD female rats, which received intravaginal administration of POD-NLC or vehicle for 3 times within 24 h; After 14 days of continuous observation, the rats were dissected for calculating the viscera coefficient.For testing the chronic toxicity of POD-NLC, 80 SD female rats were randomized into 4 groups and subjected to daily intravaginal administration of the vehicle or POD-NLC at low, moderate or high doses for 13 consecutive weeks.The rats were weighed once a week and at the end of the experiment, 2/3 of the rats from each group were sacrificed to collect blood samples, calculate the viscera coefficient, and examine the pathological changes in the liver.The remaining 1/3 rats were observed for another 2 weeks without further drug treatment and the same examinations were performed.@*RESULTS@#In the rabbits, 0.5% POD-NLC elicited only mild irritation while POD-T caused moderate irritation of the vaginal mucosa.In the acute toxicity test, the organ coefficients were comparable between the rats treated with the vehicle and POD-NLC (>0.05).Long-term intravaginal administration of POD-NLC did not produce significant changes in the behavior, activity, body weight, blood biochemical profiles or organ coefficient as compared with the vehicle control group (>0.05).@*CONCLUSIONS@#Intravaginal administration of 0.5% POD-NLC causes very mild irritation without obvious acute or chronic toxicity to the vaginal mucosa in rabbits and rats.

Buparvaquone nanostructured lipid carrier development: physicochemical and in vitro leishmanicidal performances / Desenvolvimento de carreadores lipídicos nanoestruturados contendo buparvaquona: caracterização físico-química e avaliação da atividade leishmanicida in vitro

Leishmaniases is a group of diseases caused by parasites of the genus Leishmania. The estimated number of deaths from visceral leishmaniases ranges from 20,000 to 50,000 annually. The most common treatment over the past 60 years has been pentavalent antimonials. Besides the doubtful effectiveness, they present several disadvantages such as the need for parenteral administration, large doses, long treatment, severe toxicity and parasite resistance. Buparvaquone (BPQ), a drug used for veterinary treatment of theileriosis, showed promising activity against Leishmania spp. However, due to its low aqueous solubility and bioavailability, it has failed in in vivo tests. The use of nanotechnologies has the potential to overcome these drawbacks due to the following advantages: increase in drug water-solubility, increase in therapeutic efficacy and treatment toxicity reduction. Therefore, the present work aimed the development, optimization, physical-chemical evaluation and in vitro performances of nanostructured lipid carriers (NLC) for BPQ encapsulation. The NLC preparation was performed by high pressure homogenization, and surface response and factorial design were applied to formulation optimization. In vitro dissolution profiles were evaluated in phosphate buffer pH 7.4 with tween 80 0.07% w/v or sodium dodecyl sulfate 1% w/v and simulated body fluid pH 7.4. Cytotoxicity was evaluated in mouse peritoneal macrophages and leishmanicidal activity in L. infantum amastigotes. Six optimized NCL were prepared and they showed solubility improvement from 1.5- fold to 611-fold when compared with free BPQ, depending on the formulation and medium. Dissolution profiles showed the NLC formulation suitability for BPQ regarding oral administration, the release could reach 83.29% of a 4mg dose in 30 minutes for formulation of 175.1 nm, while the free drug could be dissolved only 2.89% of the same dose after 4 hours. Moreover, formulation of 230.7 nm showed 81.42% of drug release in in phosphate buffer pH 7.4 with dodecyl sulfate 1.0% w/v after 30 minutes, while BPQ did not dissolved. Cytotoxicity assay showed the safety of all formulations. The iv CC50 values were close to 500 µM, while the IC50 against amastigotes was only 456.5 nM for free BPQ. Developed NLCs showed an increase in IC50 from 2.0 to 3.1-fold when compared to free drug in the in vitro leishmanicidal evaluation. Therefore, the NLC containing BPQ are a promising alternative for the treatment of leishmaniases as oral and parenteral drug dosage forms. Additionally, they have a potential use for lymphatic targeted drug delivery, which can be an innovative approach for this neglected disease.

Leishmanioses são um grupo de doenças causadas por parasitas do gênero Leishmania. O número estimado de óbitos por leishmaniose visceral varia entre 20.000 e 50.000 por ano. O tratamento mais comum nos últimos 60 anos tem sido os antimônios pentavalentes. Além da eficácia duvidosa, eles apresentam várias desvantagens, como a necessidade de administração parenteral, altas doses, tratamento prolongado, toxicidade severa e resistência parasitária. Buparvaquona (BPQ), um fármaco usado para tratamento veterinário da teileriose, mostrou atividade promissora contra Leishmania donovani. No entanto, devido à sua baixa solubilidade e biodisponibilidade aquosa, falhou em testes in vivo. O uso das nanotecnologias tem o potencial de superar esses obstáculos devido às seguintes vantagens: aumento da solubilidade em água, aumento da eficácia terapêutica e redução da toxicidade do tratamento. Portanto, o presente trabalho objetivou o desenvolvimento, otimização, avaliação físico-química e avaliação do desempenho in vitro de carreadores lipídicos nanoestruturados (NLC) para o encapsulação da BPQ. A preparação do NLC foi realizada por homogeneização de alta pressão e superfície de resposta e planejamento fatorial foram aplicados à otimização das formulações. Os perfis de dissolução in vitro foram avaliados em tampão fosfato pH 7.4 com tween 80 a 0.07% p/v ou dodecilsulfato de sódio 1.0% p/v e fluido corporal simulado pH 7.4. A citotoxicidade foi avaliada em macrófagos peritoneais de camundongos e atividade leishmanicida em amastigotas de L. infantum. Foram preparados quatro NCL otimizados e mostraram melhora da solubilidade de 1,5 a 611 vezes quando comparado com a BPQ livre, dependendo da formulação e do meio. Os perfis de dissolução mostraram a adequação da formulação NLC para BPQ em relação à administração oral. A dissolução pode atingir 83,29% de uma dose de 4.0 mg em 30 minutos para a formulação de 175,1 nm, enquanto o fármaco livre dissolveu apenas vi 2,89% da mesma dose após 4 horas. Além disso, a formulação de 230,7 nm mostrou 81,42% de liberação do fármaco em tampão fosfato pH 7.4 com dodecil sulfato de sódio 1.0% p/v após 30 minutos, enquanto o BPQ não se dissolveu. O teste de citotoxicidade mostrou a segurança de todas as formulações. Os valores CC50 foram próximos de 500 µM, enquanto o IC50 em amastigotas foi de apenas 456,5 nM para BPQ livre. Os NLC desenvolvidos mostraram um aumento no IC50 de 2,0 a 3,1 vezes quando comparado ao;fármaco livre na avaliação leishmanicida in vitro. Logo, as NLC contendo BPQ são uma alternativa promissora para o tratamento de leishmanioses como formas farmacêuticas oral e parenteral. Além disso, eles têm um uso potencial para a sítio-específico ao sistema linfático, o que pode ser uma abordagem inovadora para esta doença negligenciada.

Preparation and Formulation Optimization of Nanostructured Lipid Carriers of Adefovir Dipivoxil / 中国药房

OBJECTIVE:To prepare nanostructured lipid carrier of adefovir dipivoxil(ADV-NLC),and optimize the formula-tion. METHODS:Using stearic acid and glycerin monostearate as solid lipid,oleic acid as liquid lipid,Gemini surfactant and poly-sorbate 80 as emulsifier,sodium dodecyl sulfate (SDS) as stabilizer,solvent dispersion ultrasonic method was used to prepare ADV-NLC. And using particle size,polydispersity index,Zeta potential,encapsulation efficiency as indexes,single factor test was conducted to screen Gemini surfactant-polysorbate 80 ratio,emulsifier dosage(ratio of emulsifier to water phase),drug-lipid ratio, solid-liquid lipid ratio. RESULTS:The formula was as follow as 3% emulsifier (Gemini surfactant-polysorbate 80 ratio of 1:2), 4.5% drug-lipid ratio,solid-liquid lipid ratio of 6:5. The average particle size of the prepared ADV-NLC was(48.83±2.65)nm, polydispersity index<0.3,Zeta potential was(-28.7±1.8)mV,encapsulation efficiency was(77.65±0.03)%(n=3). CONCLU-SIONS:ADV-NLC is successfully prepared,and the formulation is reasonable and feasible.

Preparation of Oxcarbazepine Nanostructured Lipid Carriers and Their Physicochemical Properties / 中国药学杂志

OBJECTIVE: To prepare oxcarbazepine nanostructured lipid carriers (OXC-NLC), and investigate their physicochemical properties. METHODS: OXC-NLC was prepared by emulsification solvent evaporation method, and the optimum prescription was screened by orthogonal design. Scanning electron microscope was used to observe the morphology of OXC-NLC. Granulometer was applied to determine the particle size and distribution. DSC was adopted for phase analysis. RESULTS: OXC-NLC prepared under the optimum conditions was mostly spherical grains, with an average particle size of (63.04 ± 2.05) nm, which were distributed evenly, and the Zeta potential was (-33.52 ± 0.34) mV. DSC results indicated that the drug was dispersed in nanoparticles in an amorphous state, with entrapment efficiency of (98.16 ± 1.59)% and drug-loading capacity of (4.27 ± 0.70)%. The in vitro drug release was 27.31% in 8 h, which was followed by a sustained release. CONCLUSION: The prepared OXC-NLC has uniform particle size distribution, high encapsulation efficiency, and sustained release effect. This processing technology is simple, reliable, and highly reproducible.

Preparation and Transdermal Absorption in vitro of Piroxicam Nanostructured Lipid Carrier / 中国药师

Objective: To prepare piroxicam nanostructured lipid carrier and investigate its transdermal absorption behavior in vitro. Methods:Piroxicam nanostructured lipid carrier was prepared by a melt-emulsion ultrasonication and low temperature-solidifica-tion method. The physicochemical properties such as appearance, morphology, particle size distribution, PdI and zeta potential of pi-roxicam nanostructured lipid carrier were evaluated. The transdermal absorption in vitro was investigated using Franz diffusion cells. Results:Piroxicam nanostructured lipid carrier was clear and transparent with small spherical shape as seen under a transmission elec-tron microscope. The particle size distribution, PdI and zeta potential was (106. 4 ± 31. 6) nm, (0. 217 ± 0. 07) and ( -31. 6 ± 2. 5) mV, respectively. Piroxicam nanostructured lipid carrier had higher cumulative transdermal amount in 12 h than piroxicam solution. Conclusion:The nanostructured lipid carrier can remarkably improve piroxicam permeation into skin, which provides reference for the new dosage form for the topical use of piroxicam.

Preparation of isopsoralen loaded nanostructured carrier and its in vitro transdermal permeation characteristics / 中国中药杂志

To increase the permeation and retention of isopsoralen in skin, and improve its bioavailability.Isopsoralen loaded nanostructure liquid carrier (IPRN-NLC) was prepared by high pressure homogenization andoptimized by orthogonal experiment with the encapsulation efficiency, drug loading and average particle size as the evaluation indexes. The in vitro transdermal permeation of IPRN-NLC was evaluated by Franze diffusion cells.The results showed that solid-liquid lipid ratio of optimum IPRN-NLC formulation was 7∶3,drug-lipid ratio of 1∶30, 1% surfactant. Under these conditions, IPRN-NLC had an average encapsulation of （90.25±0.73）%,drug loading of （1.56±0.27）% and an average particle size of (305±1.57) nm.The in vitro transdermal permeation results showed that IPRN-NLC could increase the amount of IPRN permeated though skin, with 3 times of the epidermal retention as compared with IPRN solution. From the results we can know that the IPRN-NLC prepared by high pressure homogenization can improve the permeation andaccumulation of IPRN in the skin, with wide application prospects in the field of transdermal administration.

Preparation and evaluation of dioscin nanostructured lipid carriers / 中国中药杂志

In this report, a heat and high-pressure homogenization method was used to prepare dioscin nanostructured lipid carriers, and the formulation of dioscin nanostructured lipid carriers was optimized by central composite design-response surface methodology. In vitro evaluation data showed that the preparation of dioscin nanostructured lipid carriers under optimal process by central composite design-response surface methodology had a spherical shape and homogeneous size distribution, with a particle size of (90.9±0.6) nm, a polydispersity index of (0.253±0.07), Zeta potential of (-45.7±0.5) mV, encapsulation efficiency of (90.2±0.5)%, and the drug loading of (23.30±0.10)%. These results clearly indicate that the preparation of dioscin nanostructured lipid carriers made with the heat and high-pressure homogenization method have very good physical and chemical properties, suitable for therapeutic applications.

Oral absorption of asiatic acid nanoparticles modified with PEG / 中国中药杂志

A solvent diffusion method was used to prepare pegylated asiatic acid (AA) loaded nanostructured lipid carriers (p-AA-NLC), and the ligated intestinal circulation model was established to observe the absorption and distribution in small intestine. The concentration of AA in bile after oral administration of p-AA-NLC was detected by HPLC in healthy SD rats to indirectly evaluate the oral absorption promoting effect of PEG-modified namoparticles. The results showed that the penetration of p-AA-NLC was enhanced significantly and the transport capacity was increased greatly in small intestinal after PEG modification. As compared with the normal nanoparticles (AA-NLC), the Cmax of the drug excretion was increased by 76%, the time to reach the peak (tmax ) was decreased and the elimination half-life t1/2 was doubled in the rats after oral administration of p-AA-NLC, and the AUC0→t was 1.5 times of the AA-NLC group, indicating that the oral bioavailability of AA-NLC was significantly improved by hydrophilic modification of PEG.

Evaluation and effects on HaCaT cells in vitro of tanshinone IIA-loaded nanostructured lipid carrier / 中草药

Objective: To evaluate TIIA-NLC's quality in vitro and study its effects on the HaCaT cells. Methods: Its particle size, polydispersity index (PDI) and photostability were investigated by Zetasizer analyzer and high-performance liquid chromatography (HPLC). The in vitro release of TIIA-NLC within 72 h was measured by dialysis method; Meanwhile MTT assay was used to detect proliferation activity on HaCaT cells. Results: The average particle size, PDI and Zeta potential of TIIA-NLC was: (178 ± 9) nm, 0.183 ± 0.017 and (−27.5 ± 5.6) mV. The 72 h in vitro accumulative release rate was 52.28%, and the degradation velocity of TIIA was significantly slowed down. The TIIA-NLC inhibited the proliferation of HaCaT cells in concentration-dependent mode within a certain range. Compared with TIIA group, TIIA-NLC had more effects on the HaCaT cells. Conclusion: The prepared TIIA-NLC showed good stability, sustained release of interest, good cell biology intermiscibility, and could obviously increase the effects on the HaCaT cells.