The spun-out silk fiber consists of two fibroin filaments that are cemented together by sericin coating. The serine-rich sericins, which make 20-30% of the cocoon silk proteins in Bombyx mori, are dissolved in hot water during silk fiber reeling from the cocoon. The sericin extract is usually discarded. Only small amounts are currently used in cosmetics and lately also as replacement of bovine serum products in the cell culture media. The use in culture media is hindered by poor standardization of the extracts. To overcome this problem, we attempted preparation of several recombinant proteins based on partial sequences of one out of the three sericin genes of Bombyx mori, sericin 3. Translation product of the Ser3 gene is extremely serine-rich with a relatively high representation of the aspartate, asparagin, glutamate, and glutamine. Using specific primers we have prepared a cDNAs of 267, 279, 525, 672, and 528 bp, respectively, derived from the Ser3 gene. The cDNAs were cloned and expressed as fusion proteins with hexahistidine in Escherichia coli. The proteins were purified by affinity chromatography and analyzed by acrylamide electrophoresis. Ser3 gene contain repetitive motifs rich in serine and including some charged amino acids.

A fibroína de seda é uma proteína sintetizada pela espécie Bombyx mori, popularmente conhecida como bicho-da-seda. O casulo de Bombyx mori é composto por fibras de fibroína e pela sericina, responsável por unir os fios de fibroína. A fibroína é um polímero natural bastante versátil e pode ser processada de maneira a formar materiais como filmes, microesferas e hidrogéis. Os hidrogéis são redes tridimensionais formadas por macromoléculas e capazes de absorver grande quantidade de água sem perder sua integridade estrutural. Devido à características como biocompatibilidade, elevado teor de água e boas propriedades de difusão de oxigênio e nutrientes, os hidrogéis são amplamente utilizados em pesquisa biomédica. A fibroína é biodegradável, termicamente estável, altamente cristalina, flexível, resistente à tração, além de insolúvel em água e na maioria dos solventes orgânicos. A fibroína derivada das sedas Frison Extra e Meada 21 Denier foi separada da sericina pelo processo de degomagem, onde as fibras de seda foram imersas em soluções alcalinas e submetidas à aquecimento com posterior remoção de sericina. Os fios de fibroína foram dispersos nas soluções de CaCl2:H2O, LiCl:H2O e LiCl:EtOH:H2O. Diferenças nos tempos de dispersão para amostras distintas degomadas ou não foram observadas, bem como para cada um dos sistemas salino/solvente empregados. Após o processo de dispersão da fibroína as dispersões obtidas foram caracterizadas quanto à textura e reologia. As amostras não degomadas apresentaram maior firmeza e aumento do módulo elástico G\'. Algumas amostras contendo sericina ou etanol apresentaram comportamento newtoniano. Em uma segunda etapa, as amostras foram dialisadas para a produção do hidrogel e em seguida liofilizadas. Não houveram diferenças entre os tempos de gelificação para amostras distintas. Finalmente objetivou-se a caracterização dos hidrogéis obtidos por meio de ensaios termogravimétricos, difratométricos, espectrofotométricos e microscópicos. Todos os hidrogéis de fibroína de seda apresentaram alta resistência térmica, com presença predominante da conformação em folha-β da fibroína. Morfologicamente, os hidrogéis obtidos a partir de dispersões em LiCl:EtOH:H2O e CaCl2:EtOH:H2O apresentaram aspecto de rede enovelada, enquanto que os hidrogéis oriundos de dispersões em LiCl:H2O apresentaram estrutura lamelar. / The silk fibroin is a protein synthesized by Bombyx mori species, popularly known as silkworm silk. The Bombyx mori cocoon is composed of fibroin fibers and the sericin protein, which is responsible for joining the fibroin yarns. The fibroin is a quite versatile natural polymer and can be processed to form materials such as films, microspheres and hydrogels. Hydrogels are three-dimensional networks formed by macromolecules and capable of absorbing large quantities of water without losing their structural integrity. Due to biocompatibility, ability to mimic biological tissues, high water content and good diffusion properties of oxygen and nutrients, hydrogels are widely used in biomedical research. The fibroin is biodegradable, thermally stable, highly crystalline, flexible and tensile resistant. Besides, it is insoluble in water and in most organic solvents. The fibroin from Frison Extra and Meada 21 Denier silks was separated from the sericin by the degumming process, whereby the silk fibers were immersed in alkaline solutions and subjected to heating with subsequent dissolution and removal of sericin. Fibroin yarns were dispersed in various solutions: CaCl2:H2O; LiCl:H2O; and LiCl:EtOH:H2O. It was observed that according to the fibroin type and solvent solution used the time to achieve dispersion varied. Texture and rheology were determined for every sample after dispersion. All non-degummed samples had shown increased firmness and elastic module G\'. It was observed that the samples presented a wide behavior range, some that contained sericin or ethanol presented Newtonian behavior. Subsequently the samples were dialyzed to obtain the hydrogels and lyophilized. All samples (with exception of two) have formed hydrogels in a 24 hours period. The lyophilized hydrogels were analyzed by thermogravimetry, diffractometry, spectrophotometry and microscopy. All silk fibroin hydrogels presented high thermic resistance with β-sheet predominance. Morphologically, all hydrogels obtained from LiCl:EtOH:H2O and CaCl2:EtOH:H2O solutions had shown entangled aspect wile hydrogels from LiCl:H2O solutions had shown lamellar structures.

The peak and valley problems caused by oral administration, injection or other conventional methods, call for developing systems that can deliver therapeutics more effectively. As one of the techniques, diffusion-controlled drug release membranes have significant interest due to great ease with which they can be designed to achieve near-zeroth-order release kinetics. Since diffusion is the rate-limiting step in these systems, determining the permeability and diffusivity of drug molecules in the membrane is therefore important in evaluating drug release performance. This study focuses on the Membrane Permeation Controlled Release (MPC) system, which involves a non-porous (dense) membrane, comprising of two biopolymers, sericin and chitosan. Ciprofloxacin hydrochloride and (+)-cis-diltiazem hydrochloride were used as hydrophilic model drugs, and nitro-2-furaldehyde semicarbazone (Nitrofurazon) was used as a hydrophobic model drug. Permeation experiments were carried out in a semi-infinite reservoir/receptor system to simulate in-vitro drug release. The intrinsic permeability and diffusivity (P, D) of the drugs through the membranes were determined using a modified time-lag method based on short time permeation and mass balance method based on long time permeation. The partition coefficients Kd of the drugs in the membranes and the swelling degree of the membranes were determined by sorption/desorption experiments. The diffusivities of the drugs were also determined from the sorption/desorption kinetics. Over the experimental ranges tested, the drug concentration and membrane cross-linking did not have significant effects on these parameters presumably due to the relatively low drug concentrations and mild crosslinkings of the membranes. The diffusivity coefficients of ciprofloxacin hydrochloride, (+)-cis-diltiazem hydrochloride and nitrofurazon in the membranes were found to be in the range of (2.0-2.6)×〖10〗^(-9)±2.6×〖10〗^(-10) cm2/s, (2.5-2.6) ×〖10〗^(-9)±1.1×〖10〗^(-10) and (38-134) ×〖10〗^(-9)±33.1×〖10〗^(-9) (cm2/s), respectively, and their permeability coefficients were in the range of (24-29)×〖10〗^(-8),(51-52) ×〖10〗^(-8) and (131-169) ×〖10〗^(-8) (cm2/s), respectively. The partition coefficients were determined to be around 0.91±0.21, 25±0.12 and 26±0.31, respectively. The diffusivity coefficients determined from sorption experiments for ciprofloxacin hydrochloride, diltiazem hydrochloride and nitrofurazon were found to be in the range of (3.2-7.6) ×〖10〗^(-9)±6.3×〖10〗^(-8), (6-10) ×〖10〗^(-9)±2.6×〖10〗^(-8) and (15-18) ×〖10〗^(-9)±2.7×〖10〗^(-7) (cm2/s), respectively. Also the diffusivity coefficients determined from sorption experiments for ciprofloxacin hydrochloride, diltiazem hydrochloride and nitrofurazon were in the range of (20-47) ×〖10〗^(-9), (12-24) ×〖10〗^(-9) and (11-20) ×〖10〗^(-9) (cm2/s), respectively. Nonetheless the differences in the diffusivities calculated from permeation and sorption/desorption experiments are considered to be acceptable, in view of the different experimental techniques used in this work, for the purpose of comparison of the membrane diffusivity and permeability.

In this study, it was aimed to produce sericine/collagen composite membranes and to investigate their properties as a wound dressing. Different membrane compositions were prepared by casting and solvent evaporation method. After initial studies for optimization of ratios, membrane groups at two different thicknesses were prepared for each selected ratio and cross-linked with 3 % (w/v) glutaraldehyde (GTA). Considering the wound dressing requirements, equilibrium degree of swelling (EDS), water vapor transmission rate (WVTR), oxygen permeability, mechanical properties, in situ degradation, microbial penetration and cytotoxicity of membranes were examined. The EDS of membranes had a range of 14.91 to 4.37 (g/g) and increased significantly with the presence of sericin. There was no obvious relationship between the sericin ratio of membranes and WVTR, but the increase in membrane thickness decreased WVTR significantly. Thin and sericin containing membranes had statistically better oxygen permeabilities. Sericin deteriorated the tensile strength and elongation of membranes statistically. Cross-linked groups were resistant to hydrolytic degradation through 4 weeks of incubations. None of the membranes were penetrable to bacteria owing to their dense structure. For cytotoxicity studies, 3T3 fibroblasts and keratinocytes were seeded on membranes separately, and analyzed with MTT assays, and light microscopy and scanning electron microscopy (SEM). As regards to MTT assay, keratinocytes proliferated significantly on membranes and reached to high confluence within 7 days. Similarly, fibroblasts also showed high proliferation on membranes. Light microscopy and SEM analysis showed that both cells could attach, grow and spread on membranes. Also, cells gained their characteristic morphology after 1 day and formed flattened structure within 7 days.

The peak and valley problems caused by oral administration, injection or other conventional methods, call for developing systems that can deliver therapeutics more effectively. As one of the techniques, diffusion-controlled drug release membranes have significant interest due to great ease with which they can be designed to achieve near-zeroth-order release kinetics. Since diffusion is the rate-limiting step in these systems, determining the permeability and diffusivity of drug molecules in the membrane is therefore important in evaluating drug release performance. This study focuses on the Membrane Permeation Controlled Release (MPC) system, which involves a non-porous (dense) membrane, comprising of two biopolymers, sericin and chitosan. Ciprofloxacin hydrochloride and (+)-cis-diltiazem hydrochloride were used as hydrophilic model drugs, and nitro-2-furaldehyde semicarbazone (Nitrofurazon) was used as a hydrophobic model drug. Permeation experiments were carried out in a semi-infinite reservoir/receptor system to simulate in-vitro drug release. The intrinsic permeability and diffusivity (P, D) of the drugs through the membranes were determined using a modified time-lag method based on short time permeation and mass balance method based on long time permeation. The partition coefficients Kd of the drugs in the membranes and the swelling degree of the membranes were determined by sorption/desorption experiments. The diffusivities of the drugs were also determined from the sorption/desorption kinetics. Over the experimental ranges tested, the drug concentration and membrane cross-linking did not have significant effects on these parameters presumably due to the relatively low drug concentrations and mild crosslinkings of the membranes. The diffusivity coefficients of ciprofloxacin hydrochloride, (+)-cis-diltiazem hydrochloride and nitrofurazon in the membranes were found to be in the range of (2.0-2.6)×〖10〗^(-9)±2.6×〖10〗^(-10) cm2/s, (2.5-2.6) ×〖10〗^(-9)±1.1×〖10〗^(-10) and (38-134) ×〖10〗^(-9)±33.1×〖10〗^(-9) (cm2/s), respectively, and their permeability coefficients were in the range of (24-29)×〖10〗^(-8),(51-52) ×〖10〗^(-8) and (131-169) ×〖10〗^(-8) (cm2/s), respectively. The partition coefficients were determined to be around 0.91±0.21, 25±0.12 and 26±0.31, respectively. The diffusivity coefficients determined from sorption experiments for ciprofloxacin hydrochloride, diltiazem hydrochloride and nitrofurazon were found to be in the range of (3.2-7.6) ×〖10〗^(-9)±6.3×〖10〗^(-8), (6-10) ×〖10〗^(-9)±2.6×〖10〗^(-8) and (15-18) ×〖10〗^(-9)±2.7×〖10〗^(-7) (cm2/s), respectively. Also the diffusivity coefficients determined from sorption experiments for ciprofloxacin hydrochloride, diltiazem hydrochloride and nitrofurazon were in the range of (20-47) ×〖10〗^(-9), (12-24) ×〖10〗^(-9) and (11-20) ×〖10〗^(-9) (cm2/s), respectively. Nonetheless the differences in the diffusivities calculated from permeation and sorption/desorption experiments are considered to be acceptable, in view of the different experimental techniques used in this work, for the purpose of comparison of the membrane diffusivity and permeability.

A fibroína de seda é uma proteína sintetizada pela espécie Bombyx mori, popularmente conhecida como bicho-da-seda. O casulo de Bombyx mori é composto por fibras de fibroína e pela sericina, responsável por unir os fios de fibroína. A fibroína é um polímero natural bastante versátil e pode ser processada de maneira a formar materiais como filmes, microesferas e hidrogéis. Os hidrogéis são redes tridimensionais formadas por macromoléculas e capazes de absorver grande quantidade de água sem perder sua integridade estrutural. Devido à características como biocompatibilidade, elevado teor de água e boas propriedades de difusão de oxigênio e nutrientes, os hidrogéis são amplamente utilizados em pesquisa biomédica. A fibroína é biodegradável, termicamente estável, altamente cristalina, flexível, resistente à tração, além de insolúvel em água e na maioria dos solventes orgânicos. A fibroína derivada das sedas Frison Extra e Meada 21 Denier foi separada da sericina pelo processo de degomagem, onde as fibras de seda foram imersas em soluções alcalinas e submetidas à aquecimento com posterior remoção de sericina. Os fios de fibroína foram dispersos nas soluções de CaCl2:H2O, LiCl:H2O e LiCl:EtOH:H2O. Diferenças nos tempos de dispersão para amostras distintas degomadas ou não foram observadas, bem como para cada um dos sistemas salino/solvente empregados. Após o processo de dispersão da fibroína as dispersões obtidas foram caracterizadas quanto à textura e reologia. As amostras não degomadas apresentaram maior firmeza e aumento do módulo elástico G\'. Algumas amostras contendo sericina ou etanol apresentaram comportamento newtoniano. Em uma segunda etapa, as amostras foram dialisadas para a produção do hidrogel e em seguida liofilizadas. Não houveram diferenças entre os tempos de gelificação para amostras distintas. Finalmente objetivou-se a caracterização dos hidrogéis obtidos por meio de ensaios termogravimétricos, difratométricos, espectrofotométricos e microscópicos. Todos os hidrogéis de fibroína de seda apresentaram alta resistência térmica, com presença predominante da conformação em folha-β da fibroína. Morfologicamente, os hidrogéis obtidos a partir de dispersões em LiCl:EtOH:H2O e CaCl2:EtOH:H2O apresentaram aspecto de rede enovelada, enquanto que os hidrogéis oriundos de dispersões em LiCl:H2O apresentaram estrutura lamelar. / The silk fibroin is a protein synthesized by Bombyx mori species, popularly known as silkworm silk. The Bombyx mori cocoon is composed of fibroin fibers and the sericin protein, which is responsible for joining the fibroin yarns. The fibroin is a quite versatile natural polymer and can be processed to form materials such as films, microspheres and hydrogels. Hydrogels are three-dimensional networks formed by macromolecules and capable of absorbing large quantities of water without losing their structural integrity. Due to biocompatibility, ability to mimic biological tissues, high water content and good diffusion properties of oxygen and nutrients, hydrogels are widely used in biomedical research. The fibroin is biodegradable, thermally stable, highly crystalline, flexible and tensile resistant. Besides, it is insoluble in water and in most organic solvents. The fibroin from Frison Extra and Meada 21 Denier silks was separated from the sericin by the degumming process, whereby the silk fibers were immersed in alkaline solutions and subjected to heating with subsequent dissolution and removal of sericin. Fibroin yarns were dispersed in various solutions: CaCl2:H2O; LiCl:H2O; and LiCl:EtOH:H2O. It was observed that according to the fibroin type and solvent solution used the time to achieve dispersion varied. Texture and rheology were determined for every sample after dispersion. All non-degummed samples had shown increased firmness and elastic module G\'. It was observed that the samples presented a wide behavior range, some that contained sericin or ethanol presented Newtonian behavior. Subsequently the samples were dialyzed to obtain the hydrogels and lyophilized. All samples (with exception of two) have formed hydrogels in a 24 hours period. The lyophilized hydrogels were analyzed by thermogravimetry, diffractometry, spectrophotometry and microscopy. All silk fibroin hydrogels presented high thermic resistance with β-sheet predominance. Morphologically, all hydrogels obtained from LiCl:EtOH:H2O and CaCl2:EtOH:H2O solutions had shown entangled aspect wile hydrogels from LiCl:H2O solutions had shown lamellar structures.

Silk proteins have provided a source of unique and versatile building blocks in the fabrication of biomedical devices for addressing a range of applications. Critical to advancing this field is the ability to establish an understanding of these proteins in their native and engineered states as well as in developing scalable processing strategies, which can fully exploit or enhance the stability, structure, and functionality of the two constituent proteins, silk fibroin and sericin. The research outlined in this dissertation focuses on the evolution in architecture and capability of silks, to effectively position a functionally-diverse, renewable class of silk materials within the rapidly expanding field of smart biomaterials. Study of the process of building macroscopic silk fibers provides insight into the initial steps in the broader picture of silk assembly, yielding biomaterials with greatly improved attributes in the assembled state over those of protein precursors alone. Self-organization processes in silk proteins enable their aggregation into highly organized architectures through simple, physical association processes. In this work, a model is developed for the process of aqueous behavior and aggregation, and subsequent two-dimensional behavior of natural silk sericin, to enable formation of a range of distinct, complex architectures. This model is then translated to an engineered system of fibroin microparticles, demonstrating the role of similar phenomena in creating autonomously-organized structures, providing key insight into future “bottom up” assembly strategies. The aqueous behavior of the water-soluble silk sericin protein was then exploited to create biocomposites capable of enhanced response and biocompatibility, through a novel protein-template strategy. In this work, sericin was added to the biocompatible and biodegradable poly(amino acid), poly(aspartic acid), to improve its pH-dependent swelling response. This work demonstrated the production of a range of porous scaffolds capable providing meaningful response to environmental stimuli, with application in tissue engineering scaffolds and biosensing technologies. Finally, to expand the capabilities of silk proteins beyond process-driven parameters to directly fabricate engineered architectures, a method for silk photopatterning was explored, enabling the direct fabrication of biologically-relevant structures at the micro and nanoscales. Using a facile bioconjugation strategy, native silk proteins could be transformed into proteins with a photoactive capacity. The well-established platform of photolithography could then be incorporated into fabrication strategies to produce a range of architectures capable of addressing spatially-directed material requirements in cell culture and further applications in the use of non-toxic, renewable biological materials.

Cocoon cooking wastewaters (CW) and silk degumming wastewaters (SDW) of silk processing industry were treated by membrane processes for sericin recovery. CW contains only sericin while SDW contains both sericin and soap. Sericin in CW had four molecular weight (MW) fractions / 175-200 kDa (Sericin-1), 70-90 kDa (Sericin-2), 30-40 kDa (Sericin-3) and 10-25 kDa (Sericin-4). Two alternative process trains were developed for CW / 1. centrifugation + microfiltration + nanofiltration + precipitation, 2. centrifugation + microfiltration + nanofiltration + dialysis + precipitation. In the first process, a sericin/silkworm protein mixture was obtained with a sericin content of 39-46%. In the second one, however, a pure sericin product was obtained. The sericin recovery efficiency of the developed process train was found as 76%. Severe flux declines of 70-75% were observed in NF stage in both process trains. However, cleaning with 0.5 M NaOH and 190-200 mg/L free chlorine restored the fluxes by 83-127%. The MW of sericin in SDW was 110-120 kDa. The soap and sericin were separated in the pre-treatment stage consisting of centrifugation (pH 3.5, 10 min) and gravity settling (4 oC, 24 h). The ultrafiltration membrane with molecular weight cut-off of 5 kDa achieved 59% sericin recovery at pH 3.5, accompanied by severe flux decline of 88%. Furthermore, clean water flux was restored by only 31% via chemical cleaning.

Submitted by Edineia Teixeira (edineia.teixeira@unioeste.br) on 2018-02-23T18:26:15Z No. of bitstreams: 2 Jose_santos2017.pdf: 2362954 bytes, checksum: 3cd8da60eb3efb9e2ed240ef51fa9a23 (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) / Made available in DSpace on 2018-02-23T18:26:15Z (GMT). No. of bitstreams: 2 Jose_santos2017.pdf: 2362954 bytes, checksum: 3cd8da60eb3efb9e2ed240ef51fa9a23 (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Previous issue date: 2017-08-01 / Conselho Nacional de Pesquisa e Desenvolvimento Científico e Tecnológico - CNPq / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES / Lung cancer is highly lethal and smoking is an important risk factor. Non-small cell tumor is the most common, less aggressive type of lung carcinoma and treatment involves surgery, chemotherapy, and radiation therapy. However, late diagnosis, due to the absence of signs and symptoms in the early stages of the disease, makes the survival rate low. In this sense, the search for new chemical substances with characteristics of selectivity, effectiveness and low toxicity have been investigated in the treatment of cancer. Studies show that sericin, a protein extracted from silkworm cocoons, exhibits anti-tumor and anticarcinogenic activity in colon and skin cancer cells, stimulating apoptosis and disrupting the cell cycle, and protecting normal cells against oxidative stress and lipid peroxidation . Thus, the therapeutic potential of sericin has raised interest in evaluating its effect on non-small cell lung cancer cell line. The studies were conducted in culture, where the effects of silk protein were evaluated: on cell viability, by neutral red assays and tetrazolium - MTT cytotoxicity; in the apoptotic potential, with annexin-5 and Alexa Fluor® and Propidium Iodide assays; and in cell migration by the Wound Healing model assay. Low doses of sericin were able to increase lysosomal viability, reduce mitochondrial viability, increase apoptosis and cell migration, while high doses of sericin exponentially reduced cell migration and did not alter the rate of apoptosis / necrosis of cancer cells. Sericin is a biomaterial that causes biological effects on the cell line tested, and can be used to increase lysosomal viability, reduce mitochondrial function, increase apoptosis at low doses and inhibit high-dose cell migration. / O câncer de pulmão apresenta alta letalidade e o fumo se constitui como um importante fator de risco. O tumor de células não pequenas é o tipo de carcinoma pulmonar mais comum, menos agressivo e o tratamento envolve cirurgia, quimioterapia e radioterapia. Entretanto, o diagnóstico tardio, em função da ausência de sinais e sintomas nos estágios iniciais da doença, faz com que a taxa de sobrevivência seja baixa. Nesse sentido, a busca de novas substâncias químicas com características de seletividade, de efetividade e de baixa toxicidade tem se refletido em pesquisas que investigam o uso dessas substâncias no tratamento do câncer. Estudos mostram que a sericina, proteína extraída dos casulos do bicho-da-seda, apresenta atividade antitumoral e anticarcinogênica em células cancerosas de cólon e de pele, estimulando a apoptose e interrompendo o ciclo celular, além de proteger células normais contra estresse oxidativo e peroxidação lipídica. Assim, o potencial terapêutico da sericina suscitou o interesse em avaliar seu efeito em linhagem celular de câncer de pulmão do tipo células não pequenas. Os estudos foram conduzidos em cultura, em que foram avaliados os efeitos da proteína da seda: na viabilidade celular, por meio de ensaios pelo vermelho neutro e de citotoxicidade com o tetrazólio - MTT; no potencial apoptótico, com ensaios com anexina-5 e Alexa Fluor® e Iodeto de Propídio; e na migração celular, ensaio com o modelo Wound Healing. Baixas doses de sericina foram capazes de aumentar a viabilidade lisossomal, reduzir a viabilidade mitocondrial, aumentar a apoptose e a migração celular, enquanto altas doses de sericina reduziram exponencialmente a migração celular e não alteraram a taxa de apoptose/necrose das células cancerosas. A sericina é um biomaterial que provoca efeitos biológicos na linhagem celular testada, e pode ser utilizada para aumentar a viabilidade lisossomal, reduzir a função mitocondrial, aumentar a apoptose em baixas doses e inibir a migração celular em doses altas.

Graphene (G) and nanocrystalline diamond (NCD) are carbon allotropes and promising nanomaterials with an excellent combination of their properties, such as high mechanical strength, electrical and thermal conductivity, possibility of functionalization and very high surface area to volume ratio. For these reasons, G and NCD are employed next to electronics in biomedical applications, including implant coating, drug and gene delivery and biosensing. For a fundamental characterization of cell behavior on G and NCD, we studied osteoblast adhesion and proliferation on differently treated G and NCD. Generally, both G and NCD exhibited better properties for osteoblast cultivation than control tissue culture polystyrene. Better cell adhesion but lower cell proliferation were observed on NCD compared to G. The most surprising finding was that hydrophobic G with nanowrinkled topography enhanced cell proliferation extensively, in comparison to hydrophilic and flat G and both NCDs (hydrophobic and hydrophilic) with slightly higher roughness. Promoted cell proliferation enables faster cell colonization of G and NCD substrates, meaning faster new tissue formation which is beneficial in biomedical applications. Furthermore, it was shown that osteoblast adhesion was promoted in the initial absence of fetal bovine...