Direct spectrofluorimetric methods as alternatives to compendial ones used for the quality control of biopharmaceuticals: development, validation and application

Abstract A simple, rapid, precise, accurate and sustainable spectrofluorimetric method (SFM) was developed, validated and applied for the determination of 4-aminobenzoic acid and aromatic amino acids (phenylalanine, tryptophan and tyrosine). These compounds are used in biopharmaceutical formulations and therefore must be analyzed by quality control laboratories to meet the criteria established in pharmacopoeias. In general, potentiometric titration (PT) is described in the compendia as the official analytical technique. However, this method showed low sensitivity and selectivity, and moreover was performed with a non-aqueous solvent (acetic acid), which led to higher consumption of reagents and consequently to the formation of residues. Therefore, the SFM was developed in aqueous medium at pH 7.2 using phosphate buffer. It was successfully validated according to the ICH guidelines and showed good linearity range (r>0.999), specificity, accuracy and precision (within and between days) and robustness. The test results were compared between the SFM and PT using raw material samples, while according to the F- and t-tests at 95% confidence level, no statistical difference was found between the methods

Pullulan as a stabilizer agent of polymeric nanocapsules for drug delivery

ABSTRACT Polymeric stabilizers have received attention in the preparation of nanostructured systems due to their ability to enhance formulation stability. Considering this, the objective of this work was to prepare poly(ε-caprolactone) nanocapsules using the pullulan as a polymeric stabilizer. The nanocapsules were prepared using the interfacial deposition method of preformed polymers and they were characterized in terms of pH, average diameter, polydispersity index, zeta potential, beclomethasone dipropionate content, encapsulation efficiency, photostability and drug release profiles. The formulations showed physicochemical characteristics consistent with nanocarriers for drug delivery such as: average diameter lower than 270 nm, polydispersity indexes lower than 0.2, negative zeta potential (-22.7 to -26.3 mV) and encapsulation efficiencies close to 100%. In addition, the nanocapsules were able to delay the beclomethasone dipropionate photodegradation under UVC radiation and by the dialysis bag diffusion technique, the nanocapsules were able to prolong the drug release. Thus, pullulan could be considered an interesting excipient to formulate polymeric nanocapsules.

Gallic acid as the major antioxidant in pequi (Caryocar brasiliense Camb. ) fruit peel / Ácido gálico como principal antioxidante da casca do fruto do pequi (Caryocar brasiliense Camb. )

ABSTRACTIn this study, ethanol-water extracts of pequi fruit peel were fractionated in order to identify and quantify the major antioxidant present in it. The fractions were subjected to liquid-liquid phase extraction and silica-gel column chromatography, and antioxidant activity was monitored using the 2,2-diphenyl-1-picrylhydrazyl radical-scavenging assay. The purity of the fractions was evaluated using thin-layer chromatography and high-performance liquid chromatography (HPLC). The substance with antioxidant property was identified through the analysis in a liquid chromatography-mass spectroscopy fragmentation and was quantified using HPLC. After the Silica-gel fractionation, it was identified a fraction with high antioxidant activity and purity, which contained gallic acid as the main compound. The gallic acid was found at the amount of 26.54 ± 1.13 mg/g of the dry mass of the pequi fruit peel. Because the quantifications were performed using crude ethanol-water extract, it was suspected that gallic acid was present in a free form. Thus, pequi fruit peel may serve as an attractive alternative of feedstock for natural antioxidant production. Moreover, the results obtained in this study emphasize the value of the pequi plant, and suggests improved opportunities for families that use this fruit`s products.

RESUMOExtratos hidroetanólicos da casca do fruto do pequi foram fracionados a fim de identificar e quantificar o principal antioxidante presente. Frações do extrato foram submetidas ao particionamento líquido-líquido e fracionamento em coluna de sílica gel. As atividades antioxidantes das frações foram monitoradas usando ensaio de redução do radical 2,2-difenil-1-picrilhidrazila e a pureza das frações foi avaliada em cromatografia de camada delgada e cromatografia líquida de alta eficiência (CLAE). A substância com propriedades antioxidantes foi identificada através da análise em sistema de cromatografia líquida associada à espectrometria de massas e foi quantificada em HPLC. Após o fracionamento identificou-se uma fração com alta atividade antioxidante e pureza, contendo ácido gálico como o composto principal. Ácido gálico foi encontrado em concentrações de 26,54 ± 1,13 mg/g de massa seca. Devido às quantificações terem sido realizadas no extrato hidroetanólico bruto, acredita-se que o ácido gálico esteja presente na forma livre. Assim, a casca do fruto pequi pode servir como interessante alternativa de matéria prima para a produção desse antioxidante natural. Além disso, esse resultado enfatiza o valor da planta do pequi e sugere oportunidades para as famílias que utilizam produtos de pequi.

Potencial inseticida de óleos de origem vegetal sobre Grapholita molesta (Busck) (Lepidoptera: Tortricidae) / Insecticidal potential of vegetable oil extracts on Grapholita molesta (Busck) (Lepidoptera: Tortricidae)

Em decorrência do estudo das interações químicas entre insetos e herbívoros, e do avanço da pesquisa fitoquímica, é conhecida ampla variedade de produtos naturais com potencial inseticida. Grapholita molesta (Busck) (Lepidoptera: Tortricidae), mariposa-oriental, é uma das principais pragas do pessegueiro, danificando brotações e frutos. O objetivo do presente estudo foi verificar a mortalidade, atratividade e deterrência de óleos vegetais em G. molesta. Ovos e pupas de insetos provindos de criação artificial foram imersos nos óleos essenciais de Elionurus muticus (Spreng.) Kuntze e Cymbopogon winterianus Jowitt ex Bor nas concentrações de 0,25%, 0,5%, 1% e 0,5%, 1%, 5%. Os testes de atratividade foram realizados em olfatômetro do tipo "Y". A deterrência à oviposição foi testada em gaiolas sem escolha e de dupla escolha. O óleo de E. muticus aplicado em ovos de G. molesta nas três concentrações causou mortalidades maiores que 30%, diferindo significativamente dos controles (p<0,05). A mortalidade causada pelo óleo de citronela foi em torno de 70%, significativamente superior ao controle (p<0,01). O óleo de citronela aplicado a 1% em pupas causou mortalidade de 99,8%. No teste com o olfatômetro, não houve atratividade dos adultos para nenhum dos óleos. A média de ovos no substrato com o tratamento (0,33±0,33), foi significativamente menor que no controle (7,3±0,88) (p<0,01), indicando deterrência à oviposição.

As a result of the study on the chemical interactions between insects and herbivores and the advancement of phytochemical research studies, a variety of natural products with great insecticidal potential are described in the literature. The Grapholita molesta (Busck) (Lepidoptera: Tortricidae), commonly known as Oriental Fruit Moth, is a major peach pest damaging shoots and fruits. The purpose of this study was to assess the mortality, attractiveness and deterrence of vegetable oils in G. molesta. Insect eggs and pupae artificially created were immersed in the essential oils of Elionurus muticus (Spreng.) Kuntze and/or Cymbopogon winterianus Jowitt ex Bor at the concentrations of 0.25%, 0.5%, 1% and 0.5%, 1%, 5%, respectively. The attractiveness tests were performed in a Y-Tube olfactometer. Oviposition deterrence was tested in no-choice and dual-choice cages. Oil of E. muticus applied in eggs of G. molesta at three concentrations caused mortality above 30%, which differs significantly from controls (p<0.05). Mortality caused by citronella oil was approximately 70%, significantly higher than control (p<0.01). The application of citronella oil at 1% in pupae caused mortality of 99.8%. There was no attractiveness of adults in the olfactometer test for any of the oils. Citronella oil presented oviposition deterrence, as the mean number of eggs in the substrate with the treatment (0.33 ± 0.33) is significantly lower than in the control (7.3 ± 0.88) (p<0.01).

Use of Curcuma longa in cosmetics: extraction of curcuminoid pigments, development of formulations, and in vitro skin permeation studies

Curcuma longa is a ginger family aromatic herb (Zingiberaceae) whose rhizomes contain curcuminoid pigments, including curcumin, a compound known for its anti-inflammatory effects. The objective of this study was to obtain curcuminoid-rich extracts, develop topical formulations thereof, and assess the stability and skin permeation of these formulations. Curcuma longa extracts were obtained and used to develop formulations. Skin permeation studies were conducted in a modified Franz diffusion cell system, and skin retention of curcuminoid pigments was quantified in pig ear membrane. Prepared urea-containing gel-cream formulations were unstable, whereas all others had satisfactory stability and pseudoplastic rheological behavior. The amount of curcuminoid pigments recovered from the receptor solution was negligible. The skin concentration of curcuminoid pigments retained was positive (>20 µg/g of skin, mostly in the stratum corneum), considering the low skin permeability of curcumin. We conclude that development of topical formulations containing curcumin or Curcuma longa extract is feasible, as long as adjuvants are added to improve preservation and durability. The formulations developed in this study enabled penetration of curcumin limited to the superficial layers of the skin and then possibly without a risk of systemic action, thus permitting local use as a topical anti-inflammatory.

Curcuma longa é uma erva aromática, pertencente à família Zingiberaceae e seus rizomas contêm pigmentos curcuminoides, dentre eles a curcumina, conhecida por seu efeito anti-inflamatório. O objetivo deste trabalho foi obter extratos ricos nestes pigmentos, desenvolver e avaliar a estabilidade de formulações tópicas e realizar o estudo da penetração cutânea. Extratos de Curcuma longa foram obtidos e utilizados no desenvolvimento de formulações. A penetração e retenção cutâneas de pigmentos curcuminoides foi avaliada em pele de orelha de porco, utilizando célula de difusão de Franz modificada. As formulações de gel creme preparadas que continham ureia foram instáveis e as demais apresentaram estabilidade satisfatória e comportamento reológico pseudoplástico. A quantidade de pigmentos curcuminoides encontrada na solução receptora foi insignificante, enquanto que a quantidade retida na pele foi superior a 20 µg/g de pele. Concluiu-se como possível o desenvolvimento de formulações tópicas contendo curcumina ou extrato de Curcuma longa, conciliando a adição de adjuvantes para a preservação e maior durabilidade da mesma. As formulações promoveram a penetração da curcumina limitada às camadas superiores da pele, possivelmente sem o risco de ocorrência de ação sistêmica sendo possível a sua utilização para ação local com atividade anti-inflamatória.

Ensayos biologicos aplicables para el control de inocuidad de cosmeticos / Biology trials applicable for the control of inocular cosmetics

Esta monografía dedicada al ensayo biológico aplicables alos productos cosméticos así a la materia prima utilizada en la elaboración de ellos, fué realizada a fin de: difundir parte de la actividad docente, presentar consideraciones relativas a la necesidad de incluir los ensayos biológicos como parte del proceso de control de calidad de productos cosméticos, nombrar los tipos de ensayos bilogicos a ser aplicados a las diferentes formas cosméticas. Se espera que monogarfía contribuya a la formación de recursos humanos, y al uso permanente dequienes se desempeñan en el campo del control de la calidad de estos productos