RESUMEN
High concentration protein solutions are generally produced by spin column concentration (SCC) during early development and by tangential flow filtration (TFF) during later stages, when greater quantities of protein become available. This is based on the assumption that the protein generated by the SCC process would be fairly similar to the TFF process material. In this study, we report the case of high concentration solutions of an Fc fusion protein produced by the two processes using the same upstream drug substance (DS) with very different storage stability. The TFF and SCC batches were characterized for aggregation, viscosity, and hydrodynamic radius before and after storage at different temperatures (5°C, 25⯰C, and 40⯰C). Aggregation and viscosity of the solutions processed by TFF were higher than those processed by SCC upon storage at 25⯰C and 40⯰C for three months. Differential scanning fluorimetry (DSF) revealed differences in initial protein conformation. Upon exposure to shear stress, protein solutions showed conformational instability and increased aggregation upon storage at 35⯰C. In addition, protein solution showed higher aggregation upon shearing under mixed (downstream purification process and final formulation) buffer conditions - which are more likely to be encountered during the TFF, but not SCC, process. These results were further confirmed in an independent experiment by Fourier transform-infrared (FT-IR) spectroscopy and aggregation analysis. Taken together, these data indicate that shearing the protein in intermediate, unstable buffer conditions can lead to conformational perturbation during TFF processing, which led to higher rate of aggregation and viscosity upon storage. This study highlights the importance of testing shear stress sensitivity in the transitional buffer states of the TFF process early in development to de-risk process related product instability.
Asunto(s)
Proteínas Recombinantes de Fusión/química , Tampones (Química) , Estabilidad de Medicamentos , Almacenaje de Medicamentos , Fragmentos Fc de Inmunoglobulinas/química , Conformación Proteica , Temperatura , ViscosidadRESUMEN
The Biophorum Development Group (BPDG) is an industry-wide consortium enabling networking and sharing of best practices for the development of biopharmaceuticals. To gain a better understanding of current industry approaches for establishing biopharmaceutical drug product (DP) robustness, the BPDG-Formulation Point Share group conducted an intercompany collaboration exercise, which included a bench-marking survey and extensive group discussions around the scope, design, and execution of robustness studies. The results of this industry collaboration revealed several key common themes: (1) overall DP robustness is defined by both the formulation and the manufacturing process robustness; (2) robustness integrates the principles of quality by design (QbD); (3) DP robustness is an important factor in setting critical quality attribute control strategies and commercial specifications; (4) most companies employ robustness studies, along with prior knowledge, risk assessments, and statistics, to develop the DP design space; (5) studies are tailored to commercial development needs and the practices of each company. Three case studies further illustrate how a robustness study design for a biopharmaceutical DP balances experimental complexity, statistical power, scientific understanding, and risk assessment to provide the desired product and process knowledge. The BPDG-Formulation Point Share discusses identified industry challenges with regard to biopharmaceutical DP robustness and presents some recommendations for best practices.
Asunto(s)
Industria Farmacéutica/métodos , Preparaciones Farmacéuticas/química , Biofarmacia/métodos , Química Farmacéutica/métodos , Ensayos Clínicos como Asunto , Diseño de Fármacos , Humanos , Colaboración Intersectorial , Medición de Riesgo , Tecnología Farmacéutica/métodosRESUMEN
BMS-779788 contains a reactive tertiary hydroxyl attached to a weakly basic imidazole ring. Propensity of the carbinol toward dehydration to yield the corresponding alkene, BMS-779788-ALK, was evaluated. Elevated levels of BMS-779788-ALK were observed in excipient compatibility samples. Stability studies revealed that BMS-779788 degrades to BMS-779788-ALK in capsules and tablets prepared by both dry and wet granulation processes. An acid-catalyzed dehydration mechanism, in which the heterocyclic core contributes resonance stability to the cationic intermediate via charge transfer to the imidazole ring, was proposed. Therefore, neutralization via a buffered (pH 7.0) granulating solution was used to mitigate dehydration. Solution studies revealed degradation of BMS-779788 to BMS-779788-ALK over the pH range of 1-7.5. Reversibility was confirmed by initiating reactions with BMS-779788-ALK over the same pH range. Accordingly, a simple reversible scheme can be used to describe reactions initiated with either BMS-779788 or BMS-779788-ALK. To eliminate potential for charge delocalization across the heterocycle and probe the degradation mechanism, the imidazole ring of BMS-779788 was methylated (BMS-779788-Me). The propensity for acid-catalyzed dehydration was then evaluated. The acid stability of BMS-779788-Me confirmed that the heterocyclic core contributes to reactivity liability of the tertiary hydroxyl.
Asunto(s)
Imidazoles/química , Sulfonas/química , Cápsulas , Deshidratación , Composición de Medicamentos , Estabilidad de Medicamentos , Excipientes/química , Concentración de Iones de Hidrógeno , Cinética , Solubilidad , ComprimidosRESUMEN
A sensitive and effective method has been developed for the rapid determination of polysorbate-80 content in therapeutic monoclonal antibody (mAb) products. The method is based on the detection of the fluorescence emission of 4,4'-dianilino-1,1'-binaphthyl-5,5'-disulfonic acid dipotassium salt (bis-ANS) enhanced by the presence of polysorbate-80. The developed method includes two approaches. One requires removal of the mAb from solution prior to analysis, while the other requires only simple sample dilution. The limits of detection and quantitation, calculated from the calibration curve generated in the absence of mAb-A, were 1.5 and 4.7 parts per million, respectively. Given the comparable linear range and linearity of the linear line between the solutions, with or without mAb, the limit of detection and quantitation is assumed to be similar. The dilution method is not only fast and simple in terms of sample preparation, but it is also particularly useful for analyzing the level of polysorbate-80 contained in highly concentrated mAb products. However, given that this method does require availability of polysorbate-80-free materials of mAb for preparation of calibration standards, the protein removal method may be useful for the cases where appropriate protein materials for standard preparation are limited or unavailable.
RESUMEN
As of October 31, 2014, the Sierra Leone Ministry of Health and Sanitation had reported 3,854 laboratory-confirmed cases of Ebola virus disease (Ebola) since the outbreak began in May 2014; 199 (5.2%) of these cases were among health care workers. Ebola infection prevention and control (IPC) measures are essential to interrupt Ebola virus transmission and protect the health workforce, a population that is disproportionately affected by Ebola because of its increased risk of exposure yet is essential to patient care required for outbreak control and maintenance of the country's health system at large. To rapidly identify existing IPC resources and high priority outbreak response needs, an assessment by CDC Ebola Response Team members was conducted in six of the 14 districts in Sierra Leone, consisting of health facility observations and structured interviews with key informants in facilities and government district health management offices. Health system gaps were identified in all six districts, including shortages or absence of trained health care staff, personal protective equipment (PPE), safe patient transport, and standardized IPC protocols. Based on rapid assessment findings and key stakeholder input, priority IPC actions were recommended. Progress has since been made in developing standard operating procedures, increasing laboratory and Ebola treatment capacity and training the health workforce. However, further system strengthening is needed. In particular, a successful Ebola outbreak response in Sierra Leone will require an increase in coordinated and comprehensive district-level IPC support to prevent ongoing Ebola virus transmission in household, patient transport, and health facility settings.
Asunto(s)
Brotes de Enfermedades/prevención & control , Fiebre Hemorrágica Ebola/prevención & control , Evaluación de Necesidades , Fiebre Hemorrágica Ebola/epidemiología , Humanos , Sierra Leona/epidemiología , Factores de TiempoRESUMEN
A high-throughput thermal-scanning method to rank-order formulation conditions for therapeutic proteins is described. Apparent transition temperatures for unfolding and aggregation of four different proteins are determined using the dyes SYPRO Orange and thioflavin T (ThT) under a variety of buffer conditions. The results indicate that the ThT-based thermal scanning method offers several advantages over the previously described SYPRO Orange-based thermal scanning and allows rapid rank ordering of solution conditions relevant toward long-term storage of therapeutic molecules. The method is also amenable to high protein concentration and does not require sample dilution or extensive preparation. Additionally, this parallel use of SYPRO Orange and ThT can be readily applied to the screening of mutants for their unfolding and aggregation propensities.
Asunto(s)
Química Farmacéutica/métodos , Ensayos Analíticos de Alto Rendimiento/métodos , Proteínas/química , Algoritmos , Anticuerpos Monoclonales/administración & dosificación , Anticuerpos Monoclonales/química , Benzotiazoles , Tampones (Química) , Quimotripsinógeno , Péptidos/administración & dosificación , Péptidos/química , Conformación Proteica , Estructura Secundaria de Proteína , Reacción en Cadena en Tiempo Real de la Polimerasa , Solubilidad , Espectrometría de Fluorescencia , Temperatura , TiazolesRESUMEN
BACKGROUND: School smoking bans give officials the authority to provide a smoke-free environment, but enacting policies within the school walls is just one step in comprehensive tobacco prevention among students. It is necessary to investigate factors beyond the school campus and into the neighborhoods that surround schools. The purpose of this study was to explore the relationship between the density of tobacco retailers and the illegal tobacco sales rate within school neighborhoods and smoking behaviors among students. METHODS: This study utilized secondary data from the baseline of the Youth Tobacco Access Project. Data were collected from 10,662 students attending 21 middle schools and 19 high schools, in addition to 512 tobacco retailers, all within 24 towns in Illinois during 2002. A random-effects regression analysis was performed to assess the relationship between the density of tobacco retailers and illegal tobacco sales rates on current smoking and lifetime smoking prevalence. RESULTS: Schools had a range between 0 and 9 tobacco retailers within their neighborhood with a mean of 2.76 retailers (SD = 2.45). The illegal sales rate varied from 0% to 100%, with a mean of 13%. The density of tobacco retailers was significantly related to the prevalence of ever smoking among students (b = 0.09, t(29) = 2.03, p = .051, OR = 1.10), but not to current smoking (p > .05); the illegal tobacco sales rate was not related to current smoking or lifetime smoking prevalence (p > .05). CONCLUSION: Results indicate that tobacco retailer density may impact smoking experimentation/initiation.
Asunto(s)
Comercio/estadística & datos numéricos , Características de la Residencia , Instituciones Académicas/estadística & datos numéricos , Fumar/epidemiología , Industria del Tabaco/métodos , Adolescente , Conducta del Adolescente , Distribución por Edad , Niño , Intervalos de Confianza , Estudios Transversales , Bases de Datos Factuales , Femenino , Humanos , Masculino , Oportunidad Relativa , Prevalencia , Puerto Rico , Medición de Riesgo , Asunción de Riesgos , Servicios de Salud Escolar , Distribución por Sexo , Fumar/psicología , Prevención del Hábito de Fumar , Factores Socioeconómicos , Estudiantes/estadística & datos numéricos , Productos de Tabaco/estadística & datos numéricosRESUMEN
Saccharides, including sucrose, trehalose, mannitol, and sorbitol, are commonly employed as stabilizers, cryoprotectants, and/or tonicity adjusters in protein formulations. During the thawing of a protein-containing formulated bulk drug substance conducted prior to a drug product (DP) filling operation, a white, crystalline precipitate was observed. In addition, upon thawing, vial breakage was observed for filled DP that had been previously frozen at -40 °C. To investigate the causes of both phenomena, the freeze/thaw behavior of the formulation components was studied. Multiple physical characterization techniques, including differential scanning calorimetry (DSC), electrical resistance measurements, thermomechanical analysis (TMA), and powder X-ray diffraction (PXRD), were utilized to characterize the formulations. The PXRD pattern of precipitate collected from thawed bulk was consistent with that of a mannitol control. An exothermic transition observed by DSC, a sharp increase in electrical resistance detected via resistivity measurements, and the onset of volumetric expansion of the frozen matrix evident in the TMA curve offer evidence that the frozen mannitol solution undergoes transitions at or near the vial breakage temperature (-22 to -23 °C) observed during warming. In addition, osmolality measurements taken from fractionated aliquots of frozen samples indicated that non-uniform concentration gradients contributed to precipitation of mannitol observed at larger scales. Small-scale laboratory experiments (i.e., 10-125 mL) failed to adequately predict behavior at larger scale (i.e., in 1 L and 2 L bottles). Upon linking the detrimental behavior to the freeze/thaw properties of the tonicity adjustor, mannitol, alternative saccharide excipients, including sorbitol, sucrose, and trehalose, were evaluated at isotonic concentrations over a temperature range of -80 to 25 °C using physical-chemical techniques and visual observation. Neither precipitation nor vial breakage was observed for the alternate saccharides. Recommendations for saccharide selection are provided based on storage conditions and scale considerations for liquid biopharmaceutical formulations. LAY ABSTRACT: Saccharides, including sucrose, trehalose, mannitol, and sorbitol, are commonly employed as stabilizers, cryoprotectants and/or tonicity adjusters in protein formulations. During thawing of formulated bulk drug substance, a white, crystalline precipitate was observed. In addition, upon thawing, vial breakage was observed for filled drug product that had been previously frozen at -40 °C. To investigate the causes of both phenomena, multiple physical characterization techniques were utilized to characterize the formulations. The powder X-ray diffraction pattern of precipitate collected from thawed bulk was consistent with that of a mannitol control. Upon linking the detrimental behavior to the freeze/thaw properties of the tonicity adjustor, mannitol, alternative saccharide excipients, including sorbitol, sucrose, and trehalose, were evaluated at isotonic concentrations over a temperature range of -80 to 25 °C using physico-chemical techniques and visual observation. Neither precipitation nor vial breakage was observed for the alternate saccharides. Recommendations for saccharide selection are given based on storage conditions and scale considerations for liquid biopharmaceutical formulations.
Asunto(s)
Excipientes , Liofilización , Rastreo Diferencial de Calorimetría , Química Farmacéutica , Excipientes/química , Congelación , Manitol/química , Sacarosa/químicaRESUMEN
Poly(vinyl alcohol) (PVA)-based formulations are used for pharmaceutical tablet coating with numerous advantages. Our objective is to study the stability of PVA-based coating films in the presence of acidic additives, alkaline additives, and various common impurities typically found in tablet formulations. Opadry® II 85F was used as the model PVA-based coating formulation. The additives and impurities were incorporated into the polymer suspension prior to film casting. Control and test films were analyzed before and after exposure to 40°C/75% relative humidity. Tests included film disintegration, size-exclusion chromatography, thermal analysis, and microscopy. Under stressed conditions, acidic additives (hydrochloric acid (HCl) and ammonium bisulfate (NH(4)HSO(4))) negatively impacted Opadry® II 85F film disintegration while NaOH, formaldehyde, and peroxide did not. Absence of PVA species from the disintegration media corresponded to an increase in crystallinity of PVA for reacted films containing HCl. Films with NH(4)HSO(4) exhibited slower rate of reactivity and less elevation in melting temperature with no clear change in melting enthalpy. Acidic additives posed greater risk of compromise in disintegration of PVA-based coatings than alkaline or common impurities. The mechanism of acid-induced reactivity due to the presence of acidic salts (HCl vs. NH(4)HSO(4)) may be different.
Asunto(s)
Química Farmacéutica/métodos , Excipientes/química , Alcohol Polivinílico/química , Química Farmacéutica/normas , Cristalización/normas , Análisis Diferencial Térmico/normas , Estabilidad de Medicamentos , Excipientes/normas , Alcohol Polivinílico/normas , SolubilidadRESUMEN
PEGylated poly(carboxylic acid)s, PEG-b-PCAs, were evaluated as additives for solubilized oral formulations of weakly acidic compounds. Micelles of poly(ethylene glycol)-block-poly(acrylic acid), PEG-b-PAA, and poly(ethylene glycol)-block-poly(methacrylic acid), PEG-b-PMAA, were prepared. Fluorescence spectroscopy and dynamic light scattering revealed that both polymers assemble into nanoscopic structures (< 200 nm) in acidic media and exhibit pH-sensitive colloidal phase behavior. Using a solvent evaporation technique, the block copolymers and corresponding PCA homopolymers were incorporated into PEG3350-based solid dispersions. The kinetic solubility profile of a BMS compound, BMS-A (Seq ~ 12.5 µg/mL at pH 1.1) in 0.1 N HCl was monitored as a function of polymer composition. While BMS-A precipitated rapidly in 0.1 N HCl in the absence of PEG-b-PCAs, a supersaturated level of ca. 400 µg/mL was maintained for variable lengths of time in the presence of PEG-b-PCAs. Although the kinetic solubility of BMS-A was also enhanced in the presence of the PCA homopolymers, the relative magnitude and duration of supersaturation as a function of polymer composition suggests that micellar solubilization, rather than specific interaction, contributes to enhanced solubility of BMS-A in 0.1 N HCl. Under acidic conditions, pH-responsive PEG-b-PCAs may offer the kinetic supersaturation necessary to minimize precipitation of compounds which have limited solubility in acidic milieu.
Asunto(s)
Ácidos Carboxílicos/química , Química Farmacéutica/métodos , Micelas , Polietilenglicoles/química , Tensoactivos/química , Ácidos Carboxílicos/farmacocinética , Coloides , Concentración de Iones de Hidrógeno , Polietilenglicoles/farmacocinética , Polímeros/química , Polímeros/farmacocinética , Solubilidad , Tensoactivos/farmacocinética , Difracción de Rayos X/métodosRESUMEN
This study evaluated the effects of tobacco PUP (Purchase, Use and Possession) laws on tobacco use patterns among students in twenty-four towns, which were randomly assigned into an experimental and a control group. The experimental group involved both PUP law enforcement and reducing minors' access to commercial sources of tobacco, and the condition for the control group involved only efforts to reduce minors' access to commercial sources of tobacco. The present study found that adolescents in the control group had a significantly greater increase in the percentage of youth who smoked 20 or more cigarettes per day when compared to the experimental group.
Asunto(s)
Aplicación de la Ley , Prevención del Hábito de Fumar , Adolescente , Comercio/legislación & jurisprudencia , Humanos , Illinois/epidemiología , Fumar/epidemiologíaRESUMEN
OBJECTIVE: To determine whether youths who have smoked cigarettes in the past 30 days perceive themselves as smokers. METHODS: Sensitivity and specificity for 3 classifications were analyzed and compared to youths' perceptions of smoking status. RESULTS: The common criterion of having smoked cigarettes in the past 30 days reflected youths' perceptions of their smoking status with modest accuracy although adding a second criterion of having also smoked 100 or more cigarettes in a lifetime more accurately reflects youths' perceptions of their smoking status. CONCLUSIONS: Youths frequently determine smoking status based on behavioral criteria that differ from the standard criterion of 30-day point prevalence.
Asunto(s)
Percepción , Fumar/epidemiología , Adolescente , Niño , Femenino , Humanos , Masculino , Sensibilidad y Especificidad , Encuestas y Cuestionarios , Estados Unidos/epidemiologíaRESUMEN
BACKGROUND: The school setting is frequently used both to educate youth about risks involved in tobacco use and to implement tobacco prevention and cessation programs. Given that school-based programs have resulted in limited success, it is necessary to identify other setting-level intervention strategies. School tobacco policies represent a type of universal intervention that might have some promise for preventing or reducing tobacco use. METHODS: Hierarchical linear modeling was used to assess whether school tobacco policies were related to observations of tobacco use and current smoking among 16,561 seventh through twelfth graders attending 40 middle and high schools in Illinois. RESULTS: Results indicated that the enforcement of school tobacco policies, but not the comprehensiveness of those policies, was associated with fewer observations of tobacco use by minors on school grounds as well as lower rates of current smoking among students. CONCLUSIONS: The school setting is a key system to impact youth tobacco use. Findings underscore the need to train school personnel to enforce school tobacco policy.
Asunto(s)
Conducta del Adolescente , Política Organizacional , Instituciones Académicas , Fumar/epidemiología , Adolescente , Femenino , Educación en Salud/métodos , Humanos , Illinois/epidemiología , Modelos Lineales , Masculino , Servicios de Salud Escolar , Cese del Hábito de Fumar , Prevención del Hábito de FumarAsunto(s)
Contaminación del Aire Interior/prevención & control , Comercio/legislación & jurisprudencia , Promoción de la Salud/legislación & jurisprudencia , Evaluación de Resultado en la Atención de Salud , Prevención del Hábito de Fumar , Fumar/legislación & jurisprudencia , Adolescente , Contaminación del Aire Interior/legislación & jurisprudencia , Humanos , Illinois , Aplicación de la LeyRESUMEN
This study examined the impact of tobacco possession laws on public smoking among youth. There were two intervention sites: a fast food restaurant and a shopping mall. Two control sites were also monitored for public smoking among youth. Preliminary findings suggest that when police issued tickets to minors for violating tobacco possession laws, the number of youth smoking in public declined in both towns, with a more dramatic decrease occurring at the fast food site. In contrast, public smoking among youth in the control sites was not affected. The significance of reducing number of youth smoking in public through tobacco possession laws is discussed.
Asunto(s)
Salud Pública , Cese del Hábito de Fumar/métodos , Prevención del Hábito de Fumar , Adolescente , Adulto , HumanosRESUMEN
Amphiphilic block copolymers (ABCs) have been used extensively in pharmaceutical applications ranging from sustained-release technologies to gene delivery. The utility of ABCs for delivery of therapeutic agents results from their unique chemical composition, which is characterized by a hydrophilic block that is chemically tethered to a hydrophobic block. In aqueous solution, polymeric micelles are formed via the association of ABCs into nanoscopic core/shell structures at or above the critical micelle concentration. Upon micellization, the hydrophobic core regions serve as reservoirs for hydrophobic drugs, which may be loaded by chemical, physical, or electrostatic means, depending on the specific functionalities of the core-forming block and the solubilizate. Although the Pluronics, composed of poly(ethylene oxide)-block-poly(propylene oxide)-block-poly(ethylene oxide), are the most widely studied ABC system, copolymers containing poly(L-amino acid) and poly(ester) hydrophobic blocks have also shown great promise in delivery applications. Because each ABC has unique advantages with respect to drug delivery, it may be possible to choose appropriate block copolymers for specific purposes, such as prolonging circulation time, introduction of targeting moieties, and modification of the drug-release profile. ABCs have been used for numerous pharmaceutical applications including drug solubilization/stabilization, alteration of the pharmacokinetic profile of encapsulated substances, and suppression of multidrug resistance. The purpose of this minireview is to provide a concise, yet detailed, introduction to the use of ABCs and polymeric micelles as delivery agents as well as to highlight current and past work in this area.
Asunto(s)
Sistemas de Liberación de Medicamentos/métodos , Polímeros/síntesis química , Tensoactivos/síntesis química , Lactatos/administración & dosificación , Polietilenglicoles/administración & dosificación , Polímeros/administración & dosificación , Tensoactivos/administración & dosificaciónRESUMEN
The core-forming blocks of amphiphilic diblock copolymers based on methoxypoly(ethylene oxide)-block-poly(L-aspartate), PEO-b-p(L-Asp), were derivatized to incorporate stearate side chains. The effects of stearate esterification were assessed in terms of micelle stability and amphotericin B (AmB) encapsulation/release. The level of stearate esterification modulates the relative self-aggregation state of encapsulated AmB as evidenced by absorption spectroscopy. When AmB is physically loaded into polymeric micelles, the onset of hemolytic activity toward bovine erythrocytes is delayed relative to that of the free drug. Furthermore, the extent of esterification (0, 46, or 91%) appears to have profound influence on the time-dependent hemolytic profile of AmB toward bovine erythrocytes. Particularly in the case of highly substituted stearate ester micelles, incomplete and gradual build-up of hemolysis was observed over a period of 24 h. On the basis of the corresponding absorption spectra, we speculate that encapsulated AmB may interact strongly with stearate side chains, resulting in sustained release. In a neutropenic murine model of disseminated candidiasis, kidney colony-forming unit determination revealed dose-dependent efficacy for the polymeric micelle/AmB formulation, which was not significantly different from that of Fungizone at doses of 0.2, 0.3, and 0.6 mg/kg (p = 0.7). Thus, AmB administered via a polymeric micelle formulation retained potent in vivo activity.
Asunto(s)
Anfotericina B/farmacología , Antifúngicos/farmacología , Hemólisis/efectos de los fármacos , Micelas , Polietilenglicoles/farmacología , Proteínas/farmacología , Anfotericina B/química , Animales , Antifúngicos/química , Ácido Aspártico , Biopolímeros , Candida albicans/efectos de los fármacos , Candida albicans/fisiología , Bovinos , Química Farmacéutica , Femenino , Hemólisis/fisiología , Ratones , Ratones Endogámicos ICR , Polietilenglicoles/química , Proteínas/químicaRESUMEN
We systematically altered the chemical structure of the core-forming poly(L-amino acid) block of an amphiphilic diblock copolymer series based on poly(ethylene oxide)-block-poly(N-hexyl-L-aspartamide), PEO-b-p(N-HA), acyl esters by varying the length of the attached acyl side chain. Drug-loaded micelles were prepared in good yield by a modified solvent evaporation procedure. In addition, the relative aggregation state and hemolytic activity of encapsulated amphotericin B (AmB) were analyzed by absorption spectroscopy. The length of the attached acyl side chain in PEO-b-p(N-HA) acyl ester micelles modulates the relative aggregation state of encapsulated AmB. Furthermore, acyl chain length appears to have a profound influence on the time-dependent hemolytic profile of encapsulated AmB toward bovine erythrocytes. For all acyl conjugate micelle-AmB formulations, the onset of hemolysis is delayed relative to free AmB. Particularly in the case of stearate ester micelles, the incomplete and gradual build-up of hemolysis might reflect the sustained release of drug over a period of 24 h. Based on the corresponding absorption spectrum, we speculate that encapsulated AmB may interact strongly with stearate side chains, resulting in sustained release. Via chemical manipulation of the core-forming region, it may be possible to fine-tune the release of encapsulated AmB from PEO-b-p(N-HA)-acyl ester micelles.
Asunto(s)
Anfotericina B/química , Agregación Celular/efectos de los fármacos , Hemólisis/efectos de los fármacos , Micelas , Poliésteres/química , Polietilenglicoles/química , Anfotericina B/farmacología , Animales , Bovinos , Agregación Celular/fisiología , Hemólisis/fisiología , Poliésteres/farmacología , Polietilenglicoles/farmacologíaRESUMEN
Derivatives of poly(ethylene oxide)-block-poly(beta-benzyl-aspartate), 12:25 have been prepared via aminolysis of the benzyl protecting group with 6-amino-1-hexanol, followed by subsequent acylation with acetic anhydride, hexanoic acid, lauric acid, or stearic acid. A series of amphiphilic diblock copolymers based on poly(ethylene oxide)-block-poly(N-hexyl-aspartamide)acyl conjugates with various acyl chain lengths have been prepared. The extent of esterification was determined by 1H-NMR. Aqueous micelle solutions were prepared by a dialysis method and the polymer series was characterized as a function of the acyl chain length. Transmission electron microscopy and dynamic light scattering revealed micelle-like structures of nanoscopic dimensions (< 100 nm). Environmentally sensitive fluorescent probes were loaded into the micelles in order to study the properties of the hydrophobic microdomain and to determine the critical micelle concentration (CMC). Steady-state fluorescence measurements indicated that the relative apparent core viscosity and polarity are modulated by the relative length of the attached acyl chains, as is the CMC. Increasing the acyl chain length results in a decreased CMC and a more viscous and less polar core region. Carefully chosen chemical moieties can be introduced in order to influence the properties of the poly(L-Asp) blocks of the micelles. As a result, the micellar properties can be altered via chemical modification in order to impact several key properties relevant to drug delivery.