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1.
AAPS PharmSciTech ; 21(2): 50, 2020 Jan 03.
Artigo em Inglês | MEDLINE | ID: mdl-31900727

RESUMO

Commercial development of nanosuspensions for oral drug delivery generally involves a drying step which aims to generate a stable product that rapidly releases the nanocrystals once rehydrated and can be easily processed into a final dosage form (e.g., filled into hard capsule). Cryopelletisation is a freeze drying technique allowing the production of lyophilised micrometric spheres with good flowability. In the current work, the possibility to formulate redispersible ketoconazole nanocrystal-based cryopellets able to withstand intensive handling was investigated. Cryopellets were generated by first freezing regular droplets of nanosuspension using liquid nitrogen followed by water removal by sublimation in a standard freeze dryer. Low-friable cryopellets (< 1%) were produced by embedding the nanocrystals in a stabilizing hydroxypropyl cellulose SSL grade matrix, thus proving that these structures can withstand intensive handling. A threshold quantity of hydroxypropyl cellulose SSL grade (5/20 hydroxypropyl cellulose SSL grade-to-drug mass ratio) was required in combination with D-α-tocopherol polyethylene glycol 1000 succinate (vitamin E TPGS) to successfully recover the nanocrystals over storage. A further addition of micronised crospovidone has shown a positive effect on the dissolution performance of cryopellets. Altogether, this study demonstrated that the design of cryopellets combining the strengths of freeze-dried powders (porous internal structure, low residual humidity) and pellets (free-flowing units, mechanical resistance during handling) can potentially improve the nanocrystal's redispersibility compared with other drying techniques while facilitating the downstream processing.


Assuntos
Antifúngicos/química , Cetoconazol/química , Nanopartículas/química , Liofilização/métodos , Solubilidade
2.
Carbohydr Polym ; 228: 115350, 2020 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-31635721

RESUMO

The digestibility properties and structural changes of retrograded starch (RS3) induced by ultrasonic treatment (UT) were investigated. The digestion profiles showed that UT increased the slowly digestible starch (SDS) or resistant starch (RS) of RS3 as an effective green process, corresponding to a change in hydrolysis kinetic parameters (equilibrium starch hydrolysis percentage and kinetic constant). SEM analysis showed that ultrasound led to breakage of RS3 particles followed by cracking, reorientation and crystallization. Differences in amylose content, granule size, and ζ-potential were found for native RS3 and ultrasound-treated RS3 (UT-RS3). UT decreased the relative crystallinity and gelatinization enthalpy but enhanced short-range order of RS3 based on the results of XRD, DSC, and FT-IR, respectively. Surprisingly, diffractive peaks at 13°and 20° (V-type crystalline structure) and a new exothermic peak were also observed for UT-RS3. The outcome was believed to open new pathways for regulating the digestibility properties of RS3 by UT and development of low glycemic response food.


Assuntos
Manipulação de Alimentos/métodos , Plantas Comestíveis/química , Amido/química , Amido/metabolismo , Amilose/química , Digestão , Liofilização/métodos , Hidrólise , Cinética , Ondas Ultrassônicas , Zea mays/metabolismo
3.
AAPS PharmSciTech ; 20(8): 328, 2019 Oct 31.
Artigo em Inglês | MEDLINE | ID: mdl-31673810

RESUMO

This work presents a new user-friendly lyophilization simulation and process optimization tool, freely available under the name LyoPRONTO. This tool comprises freezing and primary drying calculators, a design-space generator, and a primary drying optimizer. The freezing calculator performs 0D lumped capacitance modeling to predict the product temperature variation with time which shows reasonably good agreement with experimental measurements. The primary drying calculator performs 1D heat and mass transfer analysis in a vial and predicts the drying time with an average deviation of 3% from experiments. The calculator is also extended to generate a design space over a range of chamber pressures and shelf temperatures to predict the most optimal setpoints for operation. This optimal setpoint varies with time due to the continuously varying product resistance and is taken into account by the optimizer which provides varying chamber pressure and shelf temperature profiles as a function of time to minimize the primary drying time and thereby, the operational cost. The optimization results in 62% faster primary drying for 5% mannitol and 50% faster primary drying for 5% sucrose solutions when compared with typical cycle conditions. This optimization paves the way for the design of the next generation of lyophilizers which when coupled with accurate sensor networks and control systems can result in self-driving freeze dryers.


Assuntos
Química Farmacêutica/métodos , Manitol/síntese química , Sacarose/síntese química , Dessecação/métodos , Liofilização/métodos , Congelamento , Temperatura Alta , Temperatura Ambiente
4.
AAPS PharmSciTech ; 20(8): 327, 2019 Oct 28.
Artigo em Inglês | MEDLINE | ID: mdl-31659590

RESUMO

Freeze-dried immunoglobulin G (IgG) incorporating trehalose and human serum albumin (HSA) was statistically evaluated regarding the existence of synergism between additives on the stability profile. The levels of HSA (X1) and trehalose (X2) were independent variables. Aggregation following the process (Y1), after 2 and 3 months at 40°C (Y2) and (Y3), respectively, along with the rate constant of aggregation (Y4) were dependent variables. Aggregation and beta-sheet conformation were quantified through size-exclusion chromatography (SEC-HPLC) and Fourier transform infrared spectroscopy (FTIR). Central composite design (CCD) suggested the best formulation. The integrity and thermodynamic stability of optimized formulation were investigated through sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and differential scanning calorimetry (DSC). The calculated responses were Y1, 0-0.90%; Y2, 0.4-4.3%; Y3, 2.10-13.46%; and Y4, 0.16-0.69 1/month. The optimized formulation had 10 mg IgG, 86 mg trehalose, and 1 mg HSA with observed responses of Y1, 0.01%; Y2, 0.51%; Y3, 3.08%; and Y4, 0.33 1/month. The models were statistically well-fitted. The optimized formulation was amorphous during freeze-drying (FD), and no fragmentation was observed. Trehalose and HSA demonstrated statistical synergism. CCD was successfully employed to recommend the best ratio of stabilizers and achieve the maximum stabilization of IgG as a model freeze-dried antibody.


Assuntos
Desenho de Drogas , Imunoglobulina G/química , Albumina Sérica Humana/síntese química , Trealose/síntese química , Varredura Diferencial de Calorimetria/métodos , Combinação de Medicamentos , Estabilidade de Medicamentos , Liofilização/métodos , Humanos , Imunoglobulina G/administração & dosagem , Albumina Sérica Humana/administração & dosagem , Espectroscopia de Infravermelho com Transformada de Fourier/métodos , Trealose/administração & dosagem
5.
Biomed Res Int ; 2019: 7679319, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31531367

RESUMO

Aims: To assess the bone dimensional changes after extraction and alveolar ridge preservation (ARP) using primary coverage (closed flap technique, CFT) or healing by secondary intention (open flap technique, OFT). Materials and Methods: Ten patients (split mouth design) were planned for extraction and ARP. All sites received ARP with freeze-dried bone allograft (FDBA) and nonresorbable membrane after extraction. Clinical standardized measurements were used to assess the dimensional alterations of the alveolar ridge. Results: All patients completed the study, and a total of 20 sites were randomized to CFT or OFT group. Center height (mean difference of 8.1 mm, SD =1.9 CFT, and 7.5 mm, SD= 1.8 OFT) and buccal height (mean difference of 0.8 mm, SD =1.0 CFT, and 0.3 mm, SD= 1.1 OFT) were significantly different within the same group. However, there was no statistically significant difference between groups. In the OFT group, the keratinized tissue width was higher and the pain VAS scores at 24 hours were lower compared with the CFT (p = 0.004 and p = 0.006, respectively). Conclusions: Leaving the flap open did not have any effects on the dimensional changes of bone height or width. However, there was a wider band of keratinized tissue and less pain with the CFT compared with the OFT. The study protocol was registered at ClinicalTrials.gov, Identifier NCT03136913.


Assuntos
Processo Alveolar/cirurgia , Aumento do Rebordo Alveolar/métodos , Preservação Biológica/métodos , Retalhos Cirúrgicos/fisiologia , Idoso , Transplante Ósseo/métodos , Feminino , Liofilização/métodos , Humanos , Masculino , Membranas Artificiais , Pessoa de Meia-Idade , Projetos Piloto , Extração Dentária/métodos
6.
AAPS PharmSciTech ; 20(7): 278, 2019 Aug 08.
Artigo em Inglês | MEDLINE | ID: mdl-31396732

RESUMO

The study mainly aimed to improve the aqueous solubility of Balofloxacin (BLFX) by preparing the inclusion complexes (ICs) of BLFX with cyclodextrins (CDs). In this study, ICs in solid state were obtained by using beta-CD (ß-CD), 2-hydroxypropyl-ß-CD (HP-ß-CD), 2, 6-dimethyl-ß-CD (DM-ß-CD) through a freeze-drying technique. The formation of ICs was confirmed through Fourier-transform infrared spectroscopy, differential scanning calorimetry, powder X-ray diffraction, nuclear magnetic resonance, and scanning electron microscopy. Results demonstrated that the water solubility and dissolution rates of three ICs were distinctly improved than that of parent BLFX. Bacteriostatic experiment manifested that the antibacterial effect of BLFX was not inhibited after encapsulation in CDs. The damage of BLFX to kidney and liver cells was reduced. Consequently, successful preparation of the ICs of BLFX with CDs provided possibility for devising new dosage form of BLFX, which held great promise for further applications in clinical fields.


Assuntos
Fluoroquinolonas/química , beta-Ciclodextrinas/química , Varredura Diferencial de Calorimetria , Composição de Medicamentos , Liofilização/métodos , Espectroscopia de Ressonância Magnética , Microscopia Eletrônica de Varredura , Pós , Espectroscopia de Infravermelho com Transformada de Fourier/métodos , Difração de Raios X
7.
Molecules ; 24(15)2019 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-31374946

RESUMO

Arthrospira platensis (spirulina) is considered a source of natural molecules with nutritional and health benefits. As the different storage forms can affect the quantity and quality of bioactive ingredients, the aim of the present work was to evaluate the effects of freezing, oven-drying and freeze-drying on chemical composition of spirulina biomass. Total proteins, photosynthetic pigments and antioxidants, were analyzed and compared to respective quantities in fresh biomass. The frozen sample exhibited the highest content of phycocyanin-C, phenols, and ascorbic acid, also respect to the fresh biomass. The highest total flavonoid amount was in the freeze-dried biomass. HPLC-DAD analysis of phenolic acids revealed the presence of the isoflavone genistein, known for its therapeutic role, in all the spirulina samples. The phosphomolybdenum method (TAC) and DPPH scavenging activity were applied to determine the antioxidant activity of different samples. The highest DPPH scavenging activity was detected in fresh and freeze-dried biomass and it was positively related to carotenoid content. A positive correlation indicated that carotenoids, chlorophyll, ascorbic acid and all phenolic compounds were the major contributors to the TAC activity in spirulina biomass. The results highlighted a different functional value of spirulina biomass, depending on the processing methods used for its storage.


Assuntos
Antioxidantes/química , Liofilização/métodos , Fotossíntese , Spirulina/química , Ácido Ascórbico/química , Carotenoides/química , Clorofila/química , Dessecação/métodos , Flavonoides/química , Fenóis/química , Ficocianina/química , Pigmentação
8.
Int J Pharm ; 570: 118631, 2019 Oct 30.
Artigo em Inglês | MEDLINE | ID: mdl-31442499

RESUMO

The applicability of DCCs in a continuous freeze-drying concept based on spin freezing and infrared heating was evaluated. Maximum applicable filling volume was evaluated. Secondly the mechanistic model for the determination of the optimal dynamic infrared heater temperature during primary drying of regular vials during continuous freeze-drying was adapted and validated for DCCs. Finally, since spin frozen DCCs may be more prone to choked flow due to the small neck opening and the large product surface area, it was evaluated if the choked flow constraints in the model could be increased to improve the efficiency of the drying process. The experiments revealed that the maximum allowable filling volume for spin freezing at the current experimental setup was 0.8 ml which is 80% of the maximum filling volume. Applying the mechanistic model for the determination of the optimal dynamic infrared heater temperature during primary drying of the studied DCCs and experimentally verifying this determined infrared heater temperature trajectory resulted in an elegant freeze-dried product without visual signs of collapse. The experimentally determined primary drying time agreed with the one calculated based on the mechanistic model. Choked flow did not occur during the continuous freeze-drying of DCCs containing 3% sucrose or 3% mannitol.


Assuntos
Preparações Farmacêuticas/química , Composição de Medicamentos/métodos , Liofilização/métodos , Congelamento , Manitol/química , Sacarose/química , Temperatura Ambiente
9.
Meat Sci ; 158: 107910, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31419600

RESUMO

The aim of this study was to compare visible-near-infrared spectroscopy (VIS/NIRS) models developed from fresh or freeze-dried samples for predicting the fatty acid (FA) composition of beef samples. The hypothesis tested is that the removal of water from samples could improve the VIS/NIRS model performance. A total of 454 beef samples obtained from different bovine muscles were used. No significant differences were found in the performance of VIS/NIRS models developed from fresh or freeze-dried samples for predicting both major individual FAs and families of FAs and for some FAs (16:0, 18:0, 18:1 n-9, 18:2 n-6, 20:4 n-6, 22:5 n-3, 22:6 n-3, saturated, mono-unsaturated FA, and total n-3 long chain poly-unsaturated FAs (PUFA)). In contrast, the standard error of predictions for total PUFAs, total n-3 PUFAs, total conjugated linoleic acid, 20:5 n-3, and 18:3 n-3 were improved (by 21% on average; P < .05) in freeze-dried samples compared with fresh samples.


Assuntos
Ácidos Graxos/análise , Carne Vermelha/análise , Animais , Bovinos , Liofilização/métodos , Espectroscopia de Luz Próxima ao Infravermelho/métodos
10.
J Agric Food Chem ; 67(33): 9203-9209, 2019 Aug 21.
Artigo em Inglês | MEDLINE | ID: mdl-31369261

RESUMO

With the monitoring of hundreds of pesticides in food and feed, the comminution step is equally crucial as any other to achieve valid results. However, sample processing is often underestimated in its importance and practical difficulty to produce consistent test portions for analysis. The scientific literature is rife with descriptions of microextraction methods, but ironically, sample comminution is often ignored or dismissed as being prosaic, despite it being the foundation upon which the viability of such techniques relies. Cryogenic sample processing using dry ice (-78 °C) is generally accepted in practice, but studies have not shown it to yield representative test portions of <1 g. Remarkably, liquid nitrogen has rarely been used as a cryogenic agent in pesticide residue analysis, presumably as a result of access, cost, and safety concerns. However, real-world implementation of blending unfrozen bulk food portions with liquid nitrogen (-196 °C) using common food processing devices has demonstrated this approach to be safe, simple, fast, and cost-effective and yield high-quality results for various commodities, including increased stability of labile or volatile analytes. For example, analysis of dithiocarbamates as carbon disulfide has shown a significant increase of thiram recoveries (up to 95%) using liquid nitrogen during sample comminution. This perspective is intended to allay concerns among working laboratories about the practical use of liquid nitrogen for improved sample processing in the routine monitoring of pesticide residues in foods and feeds, which also gives promise for feasible test sample size reduction in high-throughput miniaturized methods.


Assuntos
Ração Animal/análise , Contaminação de Alimentos/análise , Liofilização/métodos , Resíduos de Praguicidas/análise , Gelo-Seco , Liofilização/instrumentação , Frutas/química , Nitrogênio/química
11.
Eur J Pharm Biopharm ; 142: 334-343, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31306751

RESUMO

The stability of live-attenuated viruses is very challenging due to thermal sensitivity; therefore, solid form is usually required (often freeze-dried products). Micropellet technology is a lyophilization technology that has the potential to provide greater flexibility in the presentation of a given vaccine particularly in multi-dose format or in combination of different vaccines. As a novel vaccine alternative process, this spray freeze-dried (SFD) micropellet technology was evaluated using as a model a yellow fever virus produced in Vero cells (vYF). Screening of excipients was performed in order to optimize physico-chemical properties of the micropellets. Sugar/polymer-based formulations induced high glass transition temperature (Tg), adequate breaking force and attrition resistance of the SFD micropellets. These mechanical parameters and their stability are of considerable importance for the storage, the transport but also the filling process of the SFD micropellets. By adding excipients required to best preserve virus infectivity, an optimal sugar/polymer-based formulation was selected to build micropellets containing vYF. Monodisperse and dried micropellets with a diameter of about 530 µm were obtained, exhibiting similar potency to conventional freeze-dried product in terms of vYF infectious titer when both solid forms were kept under refrigerated conditions (2-8 °C). Comparable kinetics of degradation were observed for vYF formulated in micropellets or as conventional freeze-dried product during an accelerated stability study using incubations at 25 °C and 37 °C over several weeks. The results from this investigation demonstrate the ability to formulate live-attenuated viruses in micropellets. Pharmaceutical applications of this novel vaccine solid form are discussed.


Assuntos
Vacina contra Febre Amarela/química , Animais , Química Farmacêutica/métodos , Estabilidade de Medicamentos , Excipientes/química , Liofilização/métodos , Vacinas Atenuadas/química , Células Vero
12.
AAPS PharmSciTech ; 20(7): 263, 2019 Jul 23.
Artigo em Inglês | MEDLINE | ID: mdl-31338714

RESUMO

Modeling of the lyophilization process, based on the steady-state heat and mass transfer, is a useful tool in understanding and optimizing of the process, developing an operating design space following the quality-by-design principle, and justifying occasional process deviations during routine manufacturing. The steady-state model relies on two critical parameters, namely, the vial heat transfer coefficient, Kv, and the cake resistance, Rp. The classical gravimetric method used to measure Kv is tedious, time- and resource-consuming, and can be challenging and costly for commercial scale dryers. This study proposes a new approach to extract both Kv and Rp directly from an experimental run (e.g., temperature and Pirani profiles). The new methodology is demonstrated using 5% w/v mannitol model system. The values of Kv obtained using this method are comparable to those measured using the classic gravimetric method. Application of the proposed approach to process scale-up and technology transfer is illustrated using a case study. The new approach makes the steady-state model a simple and reliable tool for model parameterization, thus maximizes its capability and is particularly beneficial for transfer products from lab/pilot to commercial manufacturing.


Assuntos
Liofilização/métodos , Transferência de Tecnologia , Tecnologia Farmacêutica/métodos , Temperatura Ambiente
13.
Acta Pharm ; 69(1): 17-32, 2019 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-31259719

RESUMO

Ibuprofen, a weakly acidic non-steroidal anti-inflammatory drug having poor aqueous solubility, is a challenging drug for the development of pharmaceutical formulations, resulting in numerous research attempts focusing on improvement of its solubility and consequently bioavailability. Most studies have been done for solid dosage forms, with very little attention paid to parenterals. Hence, the main purpose of the present study was to enhance ibuprofen solubility as a result of formulation composition and the freeze drying process. Moreover, the purpose was to prepare a freeze dried dosage form with improved ibuprofen solubility that could, after simple reconstitution with water for injection, result in an isotonic parenteral solution. Solubility of ibuprofen was modified by various excipients suitable for parenteral application. Drug interactions with selected excipients in the final product/lyophilisate were studied by a combined use of XRPD, DSC, Raman and ss-NMR. Analyses of lyophilized samples showed solubility enhancement of ibuprofen and in situ formation of an ibuprofen salt with the alkaline excipients used.


Assuntos
Ibuprofeno/química , Solubilidade/efeitos dos fármacos , Anti-Inflamatórios não Esteroides/química , Disponibilidade Biológica , Varredura Diferencial de Calorimetria/métodos , Química Farmacêutica/métodos , Cristalização/métodos , Formas de Dosagem , Excipientes/química , Liofilização/métodos , Pós/química , Água/química , Difração de Raios X/métodos
14.
Eur J Pharm Biopharm ; 142: 216-221, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31233863

RESUMO

The specific surface area (SSA) of freeze-dried biologics (FD) is usually measured via a Brunauer-Emmett-Teller (BET) analysis of volumetric nitrogen adsorption isotherms. However, this technique has accuracy limitations for materials <0.5 m2/g, requires dry samples, must be measured at 77 K and has slow sample preparation times (drying/degassing). Inverse gas chromatography (IGC) is chromatographic characterization technique which can be used to analyse the SSA (down to ≈0.1 m2/g) of various solid-state materials including powders using organic molecules such as octane at ambient temperatures/pressure for a range of relative humidities. This study presents a comprehensive comparison between the N2 BET adsorption versus octane BET adsorption using IGC methods for determining the SSA's for a range of freeze dried biological materials. These include IgG 5% w/w, an influenza antigen standard, sucrose 5% w/w and trehalose 5% w/w. IGC provided comparable SSA values to the N2 BET method with better reproducibility (lower RSDs %). Large variations in average SSA within manufactured FD batches were observed for both IGC and volumetric determinations. IGC was also used to measure the change in SSA with increasing humidity, with FD cakes shrinking and losing their structural integrity with increasing moisture content. Such information highlights the importance of moisture content in determining the physical stability of FD cakes as exemplified by their SSA. In conclusion, IGC is a suitable alternative method for determining the SSA of freeze-dried biological materials which are generally strongly dependent on their moisture content.


Assuntos
Produtos Biológicos/química , Adsorção , Química Farmacêutica/métodos , Cromatografia Gasosa/métodos , Dessecação/métodos , Excipientes/química , Liofilização/métodos , Umidade , Pós/química , Reprodutibilidade dos Testes , Sacarose/química , Temperatura Ambiente , Trealose/química
15.
Prep Biochem Biotechnol ; 49(9): 846-857, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31244369

RESUMO

Saccharomyces boulardii (S. boulardii) is widely adopted in the diarrhea treatment for humans or livestock, so guaranteeing the survival rate of S. boulardii is the critical issue during freeze-drying process. In this study, the survival rate of S. boulardii with composite cryoprotectants during freeze-drying procedure and the subsequent storage were investigated. With the aid of response surface method, the composite cryoprotectants were comprehensively optimized to be lactose of 21.24%, trehalose of 22.00%, and sodium glutamate of 4.00%, contributing to the supreme survival rate of S. boulardii of 64.22 ± 1.35% with the viable cell number of 9.5 ± 0.07 × 109 CFU/g, which was very close to the expected rate of 65.55% with a number of 9.6 × 109 CFU/g. The accelerated storage test demonstrated that the inactivation rate constant of the freeze-dried S. boulardii powder was k-18 = 8.04 × 10-6. In addition, the freeze-dried goat milk powder results exhibited that the inactivation rate constants were k4 = 4.48 × 10-4 and k25 = 9.72 × 10-3 under 4 and 25 °C, respectively. This work provides a composite cryoprotectant formulation that has a good protective effect for the probiotic S. boulardii during freeze-drying process, possessing the potential application prospect in food, medicine, and even feed industry.


Assuntos
Liofilização/métodos , Saccharomyces boulardii/citologia , Crioprotetores/química , Ácido Glutâmico/química , Lactose/química , Viabilidade Microbiana , Probióticos/análise , Trealose/química
16.
Eur J Pharm Biopharm ; 142: 265-279, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31252071

RESUMO

This paper provides an organic overview of the most interesting continuous freeze-drying concepts that have been proposed over the years. Attention has mainly been focused on the field of pharmaceuticals, but some background has also been given on the food industry. This work aims at providing a solid starting point for future research on continuous manufacturing for the freeze-drying of pharmaceuticals.


Assuntos
Liofilização/métodos , Tecnologia Farmacêutica/métodos , Indústria Alimentícia/métodos , Humanos
17.
Acta Biomater ; 94: 330-339, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31176843

RESUMO

Human amniotic membrane (AM) has intrinsic anti-inflammatory, anti-fibrotic and antimicrobial properties. Tissue preservation methods have helped to overcome the short shelf life of fresh AM allowing "on demand" use of AM grafts. Cryopreserved AM that retains all native tissue components, including viable cells, has clinical benefits in treating chronic wounds. However, cryopreservation requires ultra-low temperature storage, limiting the use of cryopreserved products. To overcome this limitation, a new lyopreservation method has been developed for ambient storage of living tissues. The goal of this study was to investigate the viability and functionality of AM cells following lyopreservation. Fresh AM and devitalized lyopreserved AM (DLAM) served as positive and negative controls, respectively. Using live/dead staining, we confirmed the presence of living cells in viable lyopreserved AM (VLAM) and showed that these cells persisted up to 21 days in culture medium. The functionality of cells in VLAM was assessed by their differentiation potential and anti-fibrotic activity in vitro. With osteogenic induction, cells in VLAM deposited calcium within the membrane, a marker of osteogenic cells, in a time-dependent manner. The migration of human lung fibrotic fibroblasts in a scratch wound assay was reduced significantly in the presence of VLAM-derived conditioned medium. Quantitative PCR analyses indicated that VLAM reduced the expression of pro-fibrotic factors such as type I collagen and increased the expression of anti-fibrotic factors such as hepatocyte growth factor and anti-fibrotic microRNA in fibrotic fibroblasts. Taken together, these results demonstrate that endogenous cells in VLAM remain viable and functional post-lyophilization. STATEMENT OF SIGNIFICANCE: This study, for the first time, provides direct evidence showing that tissue viability and functional cells can be preserved by lyophilization. Similar to fresh amniotic membrane (AM), viable lyopreserved AM (VLAM) retains viable cells for extended periods of time. More importantly, these cells are functional and maintain their osteogenic differentiation potential and anti-fibrotic activity. Our results confirmed that the novel lyophilization method preserves tissue viability.


Assuntos
Âmnio/fisiologia , Fibroblastos/fisiologia , Fibrose/prevenção & controle , Liofilização/métodos , Diferenciação Celular , Linhagem Celular , Membrana Celular/metabolismo , Sobrevivência Celular , Criopreservação/métodos , Meios de Cultivo Condicionados/metabolismo , Fibrose/metabolismo , Regulação da Expressão Gênica , Fator de Crescimento de Hepatócito/metabolismo , Humanos , MicroRNAs/metabolismo , Osteogênese , Sobrevivência de Tecidos
18.
Eur J Pharm Sci ; 136: 104954, 2019 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-31189083

RESUMO

Clopidogrel bisulfate (CB) is a golden antiplatelet treatment, yet its benefits are limited by its low bioavailability (<50%) caused by poor intestinal solubility and absorption. The present study aims to improve CB intestinal solubility and absorption through developing a novel stable dry CB procubosomes tablets ready to disintegrate and re-disperse upon dilution in the GIT forming in situ CB cubosome nanoparticles while simultaneously overcome the poor stability of conventional cubosome dispersion at room temperature. Glyceryl monooleate based CB cubosome dispersion was prepared using Poloxamer 407 as surfactant, freeze dried using different stabilizing excipients (dextrose, mannitol and avicel) then compressed into procubosome tablets. The effect of excipient's physicochemical properties on the flowability, in vitro dissolution and stability at accelerated conditions (40 ±â€¯2 °C/75 ±â€¯5% RH) were evaluated. The prepared procubosomes exhibited an excipient type dependent dissolution profile where Avicel based procubosome tablet CF2 showed the highest in vitro dissolution profile among other excipients used during the freeze drying process. Upon transition to intestinal pH of 6.8 to mimic the drug absorption site, CF2 procubosome Avicel tablet, was able to preserve the enhanced CB release profile (99.6 ±â€¯6.92%) compared to commercial Plavix® where, CB dissolved % dropped dramatically to 79.1 ±â€¯2.45%. After storage for six months, CF2 retained the fresh tablet drug content of 98.5 ±â€¯5.82% and dissolution properties. Moreover, following oral administration in rabbits, CF2 showed higher relative bioavailability (153%) compared to commercial Plavix® with significant higher Cmax,shorter tmax, as well as enhanced antiplatelet activity.


Assuntos
Clopidogrel/química , Clopidogrel/metabolismo , Excipientes/química , Intestinos/efeitos dos fármacos , Administração Oral , Animais , Disponibilidade Biológica , Sistemas de Liberação de Medicamentos/métodos , Liofilização/métodos , Masculino , Nanopartículas/química , Tamanho da Partícula , Poloxâmero/química , Coelhos , Solubilidade , Comprimidos/química , Comprimidos/metabolismo
19.
Methods Mol Biol ; 1992: 63-76, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31148031

RESUMO

In this chapter, conventional techniques are described for the preparation of plant samples at room temperature before examination in the high resolution scanning electron microscopy. Protocols are given on how to collect, to fix, to dehydrate, and to dry plant samples. Subsequently, it is described how to stick them to stubs and cover with a thin conductive layer. These methods are suitable for a wide variety of plant specimens, ranging from microalgae to higher plants.


Assuntos
Microscopia Eletrônica de Varredura/métodos , Plantas/ultraestrutura , Dessecação/métodos , Liofilização/métodos , Temperatura Ambiente , Fixação de Tecidos/métodos
20.
Eur J Pharm Biopharm ; 141: 139-148, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31150807

RESUMO

A novel proliposomal formulation for improved oral delivery of celecoxib (CXB) was developed using a solid dispersion technique. CXB, soy phosphatidylcholine (SPC), sorbitol, and poloxamer 188 were dissolved in a water/ethanol binary solvent system. Subsequent solvent-evaporation and lyophilization steps produced CXB-loaded proliposomes (CXBPLs) with high lipid content (as SPC, ≈20% [w/w]). Powder X-ray diffractometry and differential scanning calorimetry analyses revealed that the physical state of CXB was transformed from crystalline to amorphous after the preparation process. Reconstitution of CXBPLs with gentle shaking by hand generated CXB-loaded liposomes with nano-sized mean diameter, negative zeta potential, vesicular-shaped morphology, and high CXB entrapment efficiency (≈84.7%). CXBPLs exhibited improved dissolution rate and permeability compared with free CXB and Celebrex (a commercial product of CXB). In the pharmacokinetic study performed in rats, the CXBPL-treated group showed a 1.7-fold increase in the bioavailability of CXB compared with the free CXB-treated group (p < 0.05). The histological observation with hematoxylin and eosin staining demonstrated no additional detrimental effect of CXBPLs on the intestinal epithelia of rats compared with that of free CXB. These results suggest that the developed proliposomes provide an efficient and safe way of enhancing the oral bioavailability of poorly water-soluble drugs.


Assuntos
Celecoxib/química , Lipídeos/química , Lipossomos/química , Administração Oral , Animais , Disponibilidade Biológica , Células CACO-2 , Varredura Diferencial de Calorimetria/métodos , Linhagem Celular Tumoral , Química Farmacêutica/métodos , Portadores de Fármacos/química , Sistemas de Liberação de Medicamentos/métodos , Liberação Controlada de Fármacos/efeitos dos fármacos , Liofilização/métodos , Humanos , Masculino , Tamanho da Partícula , Permeabilidade/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley , Solubilidade/efeitos dos fármacos
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