ABSTRACT Polymeric nanoparticles have demonstrated a great potential for biomedical application, especially in drug delivery systems acting as nanocarriers. The polymers obtained from living organisms, known as biopolymers, are promising materials for these purposes since they present biodegradability, biocompatibility and low immunogenicity, which are features required for in vivo applications. In this work, polymeric nanoparticles were prepared from mixtures of biopolymers such as chitosan, collagen and gelatin by ionic reticulation with sodium tripolyphosphate. Picric acid as a model drug was encapsulated into the nanoparticles. Chitosan of 50.4 kDa of molecular weight and 81 % of deacetylation degree was prepared through the deacetylation and depolymerization of chitin extracted from squid pens (Loligo sp). The composition, size distribution, and surface charge of the prepared nanoparticles were evaluated. Moreover, the influence of the nanoparticles mass and picric acid concentration were analyzed during the loading process by determining of the loaded content and the incorporation efficiency. Maximum loaded contents of picric acid were 7.16 % and 6.45 % for chitosan/gelatin nanoparticles and chitosan/collagen nanoparticles, respectively. In vitro release tests showed a quick picric acid release in chitosan/gelatin nanoparticles. The nanoparticles prepared with collagen took longer time to release the picric acid. Kinetic studies, using mathematical models, revealed that the release takes place through a diffusion mechanism.

RESUMEN Las nanopartículas poliméricas han demostrado un gran potencial para aplicaciones biomédicas, especialmente en sistemas de liberación de fármacos actuando como nanocargadores. Los polímeros obtenidos de organismos vivos, conocidos como biopolímeros, son materiales interesantes para estos propósitos desde que presentan biodegradabilidad, biocompatibilidad y baja inmunogenicidad, en cual tiene características requeridas para aplicaciones in vivo. En este trabajo, nanopartículas poliméricas fueron preparados a partir de mezclas de biopolímeros como el quitosano, colágeno y gelatina por reticulación iónica con tripolifosfato de sodio. Quitosano de 50,4 kDa de masa molar y 81 % de grado de desacetilación fue preparado a través de la desacetilación y despolimerización de la quitina extraída a partir de pluma de calamar (Loligo sp). La composición, distribución de tamaño y carga superficial de las nanopartículas preparadas también fue evaluada. Además, la influencia de la masa de las nanopartículas y la concentración de ácido pícrico durante el proceso de cargado fue analizado mediante la determinación del contenido cargado y la eficiencia de la incorporación. El máximo contenido cargado de ácido pícrico fue de 7.16% y 6.45% para nanopartículas de quitosano/gelatina y quitosano/colágeno, respectivamente. Los ensayos de liberación in vitro mostraron una rápida la liberación de ácido pícrico en las nanopartículas de quitosano/gelatina. Las nanopartículas preparadas con colágeno liberaron el ácido pícrico en mayor tiempo. Los estudios cinéticos usando modelos matemáticos mostraron que la liberación se lleva a cabo por un mecanismo de difusión.

Biomaterials with the hydroxyapatite and biopolymers such as chitosan derived of crustaceans are is an alternative for bone repair. Carbon nanotubes have been a focus of interest because they can ameliorate the biomechanical properties of biomaterials. The objective of this study was to evaluate these materials in the repair of cranial defects in rats. The animals were divided in groups: without implant (G1), implanted with the chitosan/carbon nanotube membrane (G2), and chitosan/nanotube membrane mineralized with hydroxyapatite (G3). The animals were sacrificed 5 weeks after surgery and the skulls were removed for analysis of the defect area. The results showed absence of chronic inflammatory and little bone neoformation in the defect area of all groups. In G2 and G3 there was lack of reabsorption of the biomaterial that were encapsulated by connective tissue. In conclusion, the biomaterials were biocompatible, but their specific physicochemical properties did not indicate a considerable osteoregenerative capacity.

This study compares a wound dressing based on bacterial cellulose/collagen (BC/COL) hydrogel in rat dorsum with commercial collagenase ointment and untreated wound. The hydrogel was characterized by Scanning Electron Microscopy (SEM), Thermogravimetric Analysis (TGA), Fourier transformed - Infrared spectroscopy (FT-IR) and X-ray diffraction (XRD). According to the In vivo test and macroscopic evaluation, BC/COL hydrogel showed a better repair of wounds and promoted statistically significant differences of tissue repair between treatments on the 7th day after surgery. Better quality, quantity and orientation evaluation of collagen fibers (p=0.0001) were observed in the BC/COL hydrogel and collagenase ointment groups in relation to the control group. The BC/COL hydrogel promoted better wound healing than collagenase and the control group, therefore, it can be considered a potential wound dressing for skin regeneration.

Biomaterials are used as a promising alternative to bone grafts, including bioceramics whose composition resembles that of bone and fibrin sealants due to their hemostatic properties. The objective was to evaluate the repair of cranial defects in 40 rats, grafted with hydroxyapatite and a new fibrin sealant derived from snake venom. The animals were divided into four groups: C (control, no graft); Ha (hydroxyapatite); FS (fibrin sealant), and HaFS (hydroxyapatite and fibrin sealant). The animals were euthanized 2 and 6 weeks after surgery and wound area were submitted to analysis. After 2 weeks, immature bone was formed from the borders of the defect and in groups Ha and HaFS, few hydroxyapatite particles were surrounded by new bone. After 6 weeks, the new bone was mature and surrounded several hydroxyapatite particles, without connective tissue interposition and the volume of new bone was higher in HaFS group. The hydroxyapatite in combination with the new fibrin sealant accelerates bone repair.

Objetivou-se, neste trabalho, obter biomembranas de colágeno tratadas em solução alcalina por 72 horas (GE), a partir de centros tendinosos diafragmáticos de equinos e comparar sua biocompatibilidade com membranas conservadas em solução de glicerina a 98% (GG) e membranas não tratadas (GC). As membranas foram implantadas na fáscia interna do músculo reto do abdome de equinos e retiradas, juntamente com os tecidos adjacentes, aos sete, 63 e 126 dias pós-operatórios para a preparação de lâminas histológicas. O estudo histomorfométrico das lâminas revelou processo inflamatório mais intenso para os implantes GG e CC e cicatrização mais rápida para os implantes GE. Concluiu-se que as biomembranas colagênicas tratadas em solução alcalina são mais biocompatíveis do que biomembranas conservadas em glicerina 98%.

The objective of this research was obtain collagen biomembranes treated in alkaline solution for 72 hours (GE) from tendineous diaphragmatic center of equines and compare its biocompatibility with membranes preserved in a glycerin solution 98% (GG) and membranes do not treated (GC). The membranes were implanted in the internal fascia of recto abdominis muscle of equines and removed, with adjacent tissues, seven, 63 and 126 days postoperative for the preparation of histological slides. The histomorphometric study revealed more intense inflammatory process to GG and CC implants and faster healing for GE implants. It was concluded that the collagen biomembranes treated in alkaline solution is more biocompatible than biomembranes preserved in 98% glycerin.

The effect of different propionaldehyde ratios on the properties of bis-GMA-based comonomers and copolymers diluted with propoxylated bis-GMA (CH3bis-GMA) was evaluated. Five experimental comonomers were prepared combining bis-GMA with CH3bis-GMA and propionaldehyde at 0, 2, 8, 16, 24 mol%. Light polymerization was effected with the use of 0.2 wt. (%) each of camphorquinone and N,N-dimethyl-p-toluidine. Resin degrees of conversion (%DC) were evaluated by FT-IR spectrophotometry and Tg by Differential Scanning Calorimeter. Complex viscosity (η*), the effect of temperature on η*, and Microhardness (H) for dry and wet samples were also determined. Data were analyzed by Student's t-test, one-way ANOVA and Tukey-Kramer test (α = 0.05). The group with 24 mol% additive had a significant increase in %DC and H, and the lowest comonomer Tg and η*. No remarkable variation was noted in copolymers Tg s. All resins presented Newtonian behavior of viscosity, which linearly decreased with increased temperature. The η* decreased sigmoidally as the additive ratio increased.

OBJETIVO: Avaliar os aspectos clínicos e vascularização corneal (VC) induzida pelo enxerto interlamelar das membranas de colágeno nativo (MCN) e de colágeno aniônico (MCA) em córneas de coelhos. MÉTODOS: Um fragmento com 0,25 x 0,25 cm de MCN (grupo 1) e MCA (grupo 2) foi realizado no olho direito (olho tratado) por enxertia interlamelar. No olho esquerdo (olho controle) foi realizado apenas um túnel estromal. No olho direito (olho controle) foi realizado apenas um túnel estromal. Dezesseis coelhos foram utilizados e foram divididos em dois grupos experimentais composto por oito animais cada. A avaliação clínica foi realizada aos 1, 3, 7, 15 e 30 dias de pós-operatório. A análise da vascularização corneal foi realizada após 30 dias pelo Sistema de analisador de imagens Leica Qwin-550®. RESULTADOS: Após 7 dias, a vascularização corneal do limbo em direção central da córnea observada foi de 2,25 ± 0,71 mm (MCN) e 1,0 ± 1,69 mm (ACM), respectivamente. Após 15 dias a vascularização corneal aumentou em ambos os grupos (5,25 ± 1,03 mm - MCN; 2,0 ± 2,39 mm - MCA), diminuindo até o 30º dia (2,25 ± 2,10 mm - MCN; 0,75 ± 2,12 mm - MCA). A análise estatística indicou que as médias das distâncias dos vasos do limbo em direção ao enxerto observadas após 7 e 15 dias não diferiram estatisticamente (p=0,17), e 15 e 30 dias de pós-operatório houve tendência a diferir estatisticamente (p=0,09). Os olhos controles não apresentaram nenhuma alteração. CONCLUSÃO: As membranas de colágeno nativo e de colágeno aniônico induzem a vascularização corneal quando aplicadas na córnea de coelhos por meio de enxertia interlamelar, mas membrana de colágeno ativo induz a pequena vascularização corneal quando comparada à membrana de colágeno aniônico. Embora estudos adicionais sejam necessários, os resultados encontrados no presente estudo demonstraram que as membranas de CN e CA possam ser úteis em ceratoplastias. Estas membranas consistem em mais uma opção de enxerto para o tratamento cirúrgico de reparo da córnea em coelhos e outros animais, quando não há resolução com outras formas de tratamento médico e cirúrgico.

PURPOSE: To evaluate the corneal vascularization (CV) and the clinical aspects induced by interlamellar graft with native (NCM) and anionic (ACM) collagen membranes in rabbits corneas. METHODS: An interlamellar graft with a 0.25 x 0.25 cm NCM (group 1) or ACM (group 2) fragment was performed in the right eye (treated eye). In the left eye, an estromal tunnel was done (control eye). Sixteen rabbits were used, and they were subdivided into two experimental groups of eight animals each. The clinical evaluation was performed at the 1st, 3rd, 7th, 15th and 30th postoperative days. Corneal vascularization analysis was performed after 30 days by the Images Analizator System Leica Qwin-550®. RESULTS: After 7 days, corneal vascularization was observed at about 2.25 ± 0.71 mm (NCM) and at about 1.0 ± 1.69 mm (ACM), respectively, from the limbus in direction to the central cornea. After 15 days, CV increased in both groups (5.25 ± 1.03 mm - NCM; 2.0 ± 2.39 mm - ACM) and then progressively decreased until day 30 (2.25 ± 2.10 mm - NCM; 0.75 ± 2.12 mm - ACM). The statistical analysis indicated that the averages of the distances from the limb vessels to the grafts observed after 7 and 15 days had not differed statistically (p=0.17), and after 15 and 30 postoperative days had a tendency to differ statistically (p=0.09). The control eyes did not present any changes. CONCLUSION: The interlamellar graft with native and anionic collagen membranes induced corneal vascularization when applied to rabbit corneas, but anionic collagen membrane induced a smaller corneal vascularization when compared to native collagen membrane. Although further studies are required, the results found in this study demonstrated the usefulness of interlamellar graft with native and anionic collagen membranes in keratoplasties. These membranes consists in one more graft option for the surgical treatment of corneal repair in rabbits and others animals, when other forms of medical and surgical treatment are not effective.

Objetivou-se, neste trabalho, avaliar as alterações físico-químicas proporcionadas pelo tratamento de centros tendinosos diafragmáticos homólogos em solução alcalina seguida de liofilização, para implantação na fáscia interna do músculo reto do abdome de eqüinos. As amostras foram tratadas em períodos de 24, 48, 72, 120 e 144 horas, liofilizadas e analisadas quanto à homogeneidade, flexibilidade e resistência à sutura das amostras. Posteriormente foram caracterizadas por calorimetria exploratória diferencial e microscopia eletrônica de varredura. Para a implantação nos eqüinos, foram utilizadas amostras tratadas por 72 horas seguidas de liofilização, amostras conservadas em glicerina 98% e amostras apenas liofilizadas, que foram retiradas após uma, nove e 18 semanas para avaliar a existência de aderências. Verificou-se que a homogeneidade e a flexibilidade são diretamente proporcionais ao aumento do tempo de tratamento em solução alcalina, enquanto que a resistência é inversamente proporcional ao aumento de tempo, sendo o tratamento por 72 horas intermediário para estas características. A calorimetria exploratória diferencial mostrou que o tratamento não desnatura o colágeno presente nas amostras. Na microscopia eletrônica de varredura, observou-se que o aumento de tempo de tratamento proporciona expansão de zonas menos densas do material. Em relação à formação de aderências, as amostras apenas liofilizadas apresentaram grau máximo na formação da classificação proposta, seguida pelas amostras conservadas em glicerina 98% com grau médio e as amostras tratadas em solução alcalina e liofilizadas, que foram classificadas em grau mínimo. Concluiu-se que o tratamento por 72 horas seria mais apropriado para implantação e que a integração tissular com a parede abdominal foi melhor em relação às amostras apenas liofilizadas e às conservadas em glicerina.

The objectives in this research were to evaluate physicist-chemistries alterations in equines diaphragmatic tendineous centers implants submitted to alkaline solution and lyophilization treatment. The samples had been treated in alkaline solution for 24, 48, 72, 120 and 144 hours and lyophilized. It was analyzed the homogeneity, flexibility and resistance to the suture. After that, they had been submitted to the differential scanner calorimetry and to the scanner electronic microscopy. Samples of 72 hours in alkaline treatment, conserved in glycerin 98% and only lyophilized were implanted in recto abdominis internal fascia of equines. They were removed one, nine and 18 weeks post-implantation to evaluate the existence of tacks. It was verified that the homogeneity and flexibility are directly proportional to the increase of time of treatment in alkaline solution while the resistance is inversely proportional to the time increase, being 72 hour treatment intermediate for these characteristics. The differential scanner calorimetry showed that the treatment do not denature the present collagen in samples. The electronic microscopy showed that the increase of treatment time provides expansion of less dense zones of the material. In relation to the formation of tacks, the samples only lyophilized had presented maximum degree in the proposal classification followed for the conserved in glycerin 98% samples with average degree and the treated in alkaline solution and lyophilized samples had been classified in minimum degree. It was concluded that the treatment for 72 hours would be more appropriate for implantation and that the tissue integration with abdominal wall was better in relation to the samples lyophilized only and to the conserved in glycerin ones.

OBJETIVO: Estudar a vascularização corneal (VC) induzida pela membrana de quitosana (MQ) implantada por enxertia interlamelar na córnea de coelhos. MÉTODOS: Foram utilizados 16 coelhos. No olho esquerdo procedeu-se enxertia interlamelar de um fragmento de 0,25 x 0,25 cm de MQ (olho tratado). No olho direito realizou-se apenas a microbolsa estromal (olho controle). Avaliaram-se clinicamente os animais aos 1, 3, 7, 15 e 30 dias de pós-operatório. Aos 30 dias mensurou-se a VC pelo Sistema Analisador de Imagens LEICA QWIN-550®. RESULTADOS: Aos sete dias observou-se VC a 1,5±0,93 mm do limbo em direção ao eixo visual. Aos 15 dias houve aumento da VC (4,75±3,20 mm), que se manteve aos 30 dias (4,25±4,10 mm). Os olhos controles não apresentaram quaisquer alterações oculares. Houve diferença estatística (p<0,05) entre as áreas corneais vascularizadas dos olhos tratados e controles aos 15 e 30 dias de pós-operatório. CONCLUSÕES: A MQ induziu angiogênese corneal quando aplicada à córnea de coelhos por enxertia interlamelar, a qual persistiu de forma leve até 30 dias de pós-operatório. Embora estudos adicionais sejam necessários a MQ poderá ser mais uma opção de membrana para enxertos em ceratoplastias.

PURPOSE: To evaluate corneal vascularization (CV) induced by interlamellar graft chitosan membrane (CM) in rabbit cornea. METHODS: An interlamellar graft with a 0.25 x 0.25 cm CM fragment was performed in the left eye (treated eye). In the right eye, an estromal tunnel was done (control eye). The clinical evaluation was done at 1, 3, 7, 15 and 30 days postoperatively. CV analysis was after 30 days by the Images Analizator System LEICA QWIN-550®. RESULTS: After 7 days, CV at 1.5±0.92 mm from the limbus in direction of the cornea axial area was observed. After 15 days CV increased (4.75±3.19 mm), remaining until day 30 (4.25±4.06 mm). The control eyes did not present any changes. There was a statistical differences of the vascularizated corneal areas between control and treated eyes from the 15th to the 30th postoperative day. CONCLUSION: The chitosan membrane induced corneal angiogenesis when applied to rabbit cornea through an interlamellar graft, which was maintained at low levels until 30 days postoperatively. Although further studies are necessary, the results found here demonstrated the usefulness of chitosan membrane in keratoplasties.

Biodegradable polymer blends were obtained using collagen and chitosan. Membranes of collagen and chitosan in different proportions (3:1, 1:1 and 1:3) were prepared by mixing their acetate solutions (pH 3.5) at room temperature. The blends were characterized by differential scanning calorimetry (DSC), thermogravimetric analysis (TGA), Fourier Transform infrared (FTIR) spectroscopy, specific viscosity, water absorption and stress-strain assays. The results showed that chitosan did not interfere in the structural arrangement of the collagen triple helix and the properties of the blends can be controlled by varing the proportion of the collagen and the chitosan.

This work describes the selective hydrolysis of carboxyamide groups of asparagine and glutamine of collagen matrices for the preparation of negatively charged collagen biomaterials. The reaction was performed in the presence of chloride and sulfate salts of alkaline and alkaline earth metals in aqueous dimethylsulfoxide solution and, selectively hydrolysis of carboxyamide groups of collagen matrices was promoted without cleavage of the peptide bond. The result is a new collagen material with controlled increase in negative charge content. Although triple helix secondary structure of tropocollagen was preserved, significative changes in thermal stabilities were observed in association with a new pattern of tropocollagen macromolecular association, particularly in respect microfibril assembly, thus providing at physiological pH a new type of collagen structure for biomaterial preparation, characterized by different charge and structural contents .

Anionic collagen: calcium phosphate composite was obtained by controlled mixing of collagen and calcium phosphate until the consistence of a past. Material was characterized by a Ca/P ratio of 1.55, with a X-ray diffraction pattern similar to that for hydroxyapatite. Differential Scanning Calorimetry showed that the protein is not denatured under the processing conditions. Scanning Electronic Microscopy showed that the mineral phase are regularly covered with collagen fibers, indicating that anionic collagen is efficient in the preparation of stable form of calcium phosphate ceramic paste.