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Abstract In this study, the manufacturing process of lamivudine (3TC) and zidovudine (AZT) tablets (150+300 mg respectively) was evaluated using statistical process control (SPC) tools. These medicines are manufactured by the Fundação para o Remédio Popular "Chopin Tavares de Lima" (FURP) laboratory, and are distributed free of charge to patients infected with HIV by the Ministry of Health DST/AIDS national program. Data of 529 batches manufactured from 2012 to 2015 were collected. The critical quality attributes of weight variation, uniformity of dosage units, and dissolution were evaluated. Process stability was assessed using control charts, and the capability indices Cp, Cpk, Pp, and Ppk (process capability; process capability adjusted for non-centered distribution; potential or global capability of the process; and potential process capability adjusted for non-centered distribution, respectively) were evaluated. 3TC dissolution data from 2013 revealed a non-centered process and lack of consistency compared to the other years, showing Cpk and Ppk lower than 1.0 and the chance of failure of 2,483 in 1,000,000 tablets. Dissolution data from 2015 showed process improvement, revealed by Cpk and Ppk equal to 2.19 and 1.99, respectively. Overall, the control charts and capability indices showed the variability of the process and special causes. Additionally, it was possible to point out the opportunities for process changes, which are fundamental for understanding and supporting a continuous improvement environment.

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Leishmaniasis, a neglected tropical disease, is prevalent in 98 countries with the occurrence of 1.3 million new cases annually. The conventional therapy for visceral leishmaniasis requires hospitalization due to the severe adverse effects of the drugs, which are administered parenterally. Buparvaquone (BPQ) showed in vitro activity against leishmania parasites; nevertheless, it has failed in vivo tests due to its low aqueous solubility. Though, lipid nanoparticles can overcome this holdback. In this study we tested the hypothesis whether BPQ-NLC shows in vivo activity against L. infantum. Two optimized formulations were prepared (V1: 173.9 ± 1.6 nm, 0.5 mg of BPQ/mL; V2: 232.4 ± 1.6 nm, 1.3 mg of BPQ/mL), both showed increased solubility up to 73.00-fold, and dissolution up to 83.29%, while for the free drug it was only 2.89%. Cytotoxicity test showed their biocompatibility (CC50 >554.4 µM). Besides, the V1 dose of 0.3 mg/kg/day for 10 days reduced the parasite burden in 83.4% ±18.2% (p <0.05) in the liver. BPQ-NLC showed similar leishmanicidal activity compared to miltefosine. Therefore, BPQ-NLC is a promising addition to the limited therapeutic arsenal suitable for leishmaniasis oral administration treatment.

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This study investigated the feasibility of polysaccharide-coated poly(n-butyl cyanoacrylate) (PBCA) nanoparticles for oral delivery of acyclovir (ACV). PBCA nanoparticles were obtained by the emulsion polymerization method. Chitosan was chemically modified to obtain N,N,N-trimethylchitosan (TMC), which was used to coat the nanoparticles (PBCA-TMC). Nanoparticles were characterized by dynamic light scattering, zeta potential, differential scanning calorimetry (DSC), atomic force microscopy (AFM), cytotoxicity, and the effect on the transepithelial electrical resistance (TEER) of the Caco-2 cells. The size of the coated nanoparticles (296.2 nm) was significantly larger than uncoated (175.0 nm). Furthermore, PBCA nanoparticles had a negative charge (-11.7 mV), which was inverted to highly positive values (+36.5 mV) after coating. DSC analysis suggested the occurrence of the coating, which was confirmed by AFM images. The MTT assay revealed concentration-dependent cytotoxicity for the core-shell nanoparticles. Additionally, PBCA-TMC caused a significant but reversible decrease in the Caco-2 cell monolayer TEER. Entrapped ACV (PBCA-ACV-TMC), a Biopharmaceutical Classification System class III drug substance, increased approximately 3.25 times the Papp of ACV in the Caco-2 permeability assay. The nanoparticles were also able to provide in vitro ACV controlled release using media with different pH values (1.2; 6.8; 7.4). Accordingly, this new core-shell nanoparticle showed the potential to improve the oral delivery of ACV.

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Nanocrystals and lipid-based nanosystems have the potential to play a crucial role in a significant shift in the treatment of ophthalmic diseases. These drug delivery systems allow overcoming the barriers imposed by anatomy and physiology of the organ of vision. This review aims to present new perspectives for these innovative preparations, emphasising the applications of the nanocrystal and lipid-based nanosystem while outlining their advantages and the drawbacks. The in vivo performance of the lipid-based nanosystems was highlighted. Lipid-based nanosystems and nanocrystals showed a prolonged effect, improved ocular bioavailability, upper therapeutic efficacy, higher permeation, prolonged residence time, and sustained drug release, compared to the current applications. Well-established and innovative developments updates of these systems are highlighted herein.

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OBJECTIVES: This study aimed to describe the preparation and in vitro evaluation of a surface-modified nanostructured lipid carrier (NLC) using chitosan and dextran for co-delivery of buparvaquone (BPQ) and polymyxin B (PB) against leishmaniasis. METHODS: The NLC was prepared using high-pressure homogenisation. Polymyxin B binding and surface modification with biopolymers were achieved by electrostatic interaction. In vitro cytotoxicity was assessed in mouse peritoneal macrophages, and leishmanicidal activity in amastigotes of Leishmania infantum. RESULTS: The performance attributes of BPQ-NLC, BPQ-NLC-PB[A-] (anionic) and BPQ-NLC-PB[C+] (cationic) were respectively: Z-average 173.9 ± 1.6, 183.8 ± 4.5 and 208.8 ± 2.6 nm; zeta potential -19.6 ± 1.5, -20.1 ± 1.1 and 31.1 ± 0.8 mV; CC50 583.4 ± 0.10, 203.1 ± 0.04 and 5.7 ± 0.06 µM; IC50 229.0 ± 0.04, 145.7 ± 0.04 and 150.5 ± 0.02 nM. The NLC in vitro leishmanicidal activity showed up to 3.1-fold increase when compared with free BPQ (P < 0.05, α = 0.05). CONCLUSIONS: The developed NLC proved to be a promising formulation with which to overcome the drawbacks of current leishmaniasis treatment by the co-delivery of two alternative drugs and a macrophage targeting modified surface.

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Dextran-coated poly (n-butyl cyanoacrylate) nanoparticles (PBCA-NPs) were prepared and were evaluated for enhanced delivery of a promising anti-Leishmania drug candidate, hydroxymethylnitrofurazone (NFOH), to phagocytic cells. Currently available chemotherapy for leishmaniasis, such as pentavalent antimonials, presents low safety and efficacy. Furthermore, widespread drug resistance in leishmaniasis is rapidly emerging. To overcome these drawbacks, the use of nanosized delivery systems can reduce systemic drug toxicity and increase the drug concentration in infected macrophages, therefore improving treatment of leishmaniasis. PBCA-NPs containing NFOH (PBCA-NFOH-NPs) were prepared by an anionic emulsion polymerisation method. The z-average and polydispersity index (PDI) were determined by photon correlation spectroscopy, the zeta potential by microelectrophoresis and the entrapment efficiency by HPLC. Cytotoxicity was determined using macrophages from BALB/c mice. Efficacy tests were performed using Leishmania amazonensis promastigotes and amastigotes. The z-average of PBCA-NFOH-NPs was 151.5 ± 61.97 nm, with a PDI of 0.104 ± 0.01, a zeta potential of -10.1 ± 6.49 mV and an entrapment efficiency of 64.47 ± 0.43%. Efficacy in amastigotes revealed IC50 values of 0.33 µM and 31.2 µM for the nanostructured and free NFOH, respectively (95-fold increase). The cytotoxicity study indicated low toxicity of the PBCA-NFOH-NPs to macrophages. The selectivity index was 370.6, which is 49-fold higher than free NFOH (7.6). Such findings indicated that improved efficacy could be due to NP internalisation following site-specific drug delivery and reactivation of immune protective reactions by the NP components. Thus, PBCA-NFOH-NPs have the potential to significantly improve the treatment of leishmaniasis, with reduced systemic side effects.

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Buparvaquone (BPQ), a veterinary drug, was formulated as nanostructured lipid carriers (NLC) for leishmaniases treatment. The formulation design addressed poor water solubility of BPQ and lack of human drug delivery system. The DSC/TG and microscopy methods were used for solid lipids screening. Softisan® 154 showed highest BPQ solubility in both methods. The BPQ solubility in liquid lipids using HPLC revealed Miglyol® 812 as the best option. Response surface methodology (RSM) was used to identify the optimal Softisan154 : Miglyol 812 ratios (7 : 10 to 2 : 1) and Kolliphor® P188 and Tween® 80 concentration (>3.0% w/w) aiming for z-average in the range of 100-300 nm for macrophage delivery. The NLC obtained by high-pressure homogenization showed low z-averages (<350 nm), polydispersity (<0.3), and encapsulation efficiency close to 100%. DSC/TG and microscopy in combination proved to be a powerful tool to select the solid lipid. The relationship among the variables, demonstrated by a linear mathematical model using RSM, allowed generating a design space. This design space showed the limits in which changes in the variables influenced the z-average. Therefore, these drug delivery systems have the potential to improve the availability of affordable medicines due to the low cost of raw materials, using well established, reliable, and feasible scale-up technology.

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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.

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Hydroxymethylnitrofurazone (NFOH) is a new compound with potential leishmanicidal and trypanocidal activity. Despite its effectiveness, the formulators have to overcome its poor aqueous solubility. Recently, polymeric nano-scale drug delivery systems have proposed for the treatment of neglected diseases. As several studies have confirmed the advantages of such formulations, and this approach provides new analytical challenges, including the need to detect trace amounts of the drug. A suitable method was developed and validated for NFOH determination bound to poly (n-butylcyanoacrylate) (PBCA) nanoparticles. The chromatographic separation was achieved using a C18 column maintained at 25 ºC and an isocratic mobile phase consisting of water and acetonitrile: 80:20 (v/v) at a flow rate of 1.2 mL min-1 and UV-detection at 265 nm. Investigated validation parameters included selectivity, linearity, accuracy, precision and robustness (changes in column temperature, mobile phase composition and flow). The method was specific, the peak of NFOH had no interference with any nanoparticle excipients and no co-elution with main degradation product (nitrofurazone). Linearity was over the range of 0.94 13.11 μg mL-1 (r2=0.999). The method was accurate and precise, recovery of 100.7%, RSD of 0.4%; intra-day and inter-day RSD range 9.98-9.99 μg mL-1 and 0.3% to 0.5%, respectively. Robustness confirmed that method could resist the applied changes. Application of the optimized method revealed an encapsulation efficiency of 64.4% (n=3). Therefore, the method was successfully developed and validated for the determination of the encapsulation efficiency of NFOH-PBCA nanoparticles.

Hidroximetilnitrofural (NFOH) é um novo composto que possui atividade leishmanicida e tripanomicida potencial. Um método apropriado foi desenvolvido e validado para a determinação de NFOH em nanopartículas de poli(n-butil cianoacrilato) (PBCA). A separação cromatográfica foi obtida usando uma coluna C18 (5 µm de tamanho de partícula, 4,6 mm de diâmetro e 150 mm de comprimento), mantida a 25 °C, fase móvel composta de água e acetonitrila 80:20 (v/v), fluxo de 1,2 mL min- 1 e detecção por UV a 265 nm. Investigaram-se os seguintes parâmetros de validação: seletividade, linearidade, exatidão, precisão e robustez (mudanças na temperatura de coluna, proporção da fase móvel e fluxo). O método mostrou-se específico, o pico de NFOH não apresentou interferência dos picos provenientes dos excipientes das nanopartículas e separado do principal produto de degradação (nitrofural). A linearidade foi obtida na faixa de 0,94-13,11 μg mL- 1 (r2=0,999). O método mostrou exatidão (recuperação de 100,7%, DPR de 0,4 %) e precisão (intra-dia e inter-dia, 9,98-9,99 μg mL- 1 e DPR 0,3% a 0,5%, respectivamente). A robustez provou que o método pode resistir às mudanças propostas. Aplicação do método otimizado revelou eficiência de encapsulação de 64,4% (n=3). Portanto, o método foi desenvolvido e validado com sucesso para a determinação da eficiência de encapsulação de nanopartículas de NFOH-PBCA.

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