RESUMO
Biodegradable polyesters are widely employed in the development of controlled release systems for peptide drugs. However, one of the challenges in developing a polyester-based delivery system for peptides is the acylation reaction between peptides and polymers. Peptide acylation is an important factor that affects formulation stability and can occur during storage, in vitro release, and after drug administration. This review focuses on the mechanisms and parameters that influence the rate of peptide acylation within polyesters. Furthermore, it discusses reported strategies to minimize the acylation reaction.
Assuntos
Poliésteres , Ácido Poliglicólico , Copolímero de Ácido Poliláctico e Ácido Poliglicólico , Ácido Láctico , Peptídeos , AcilaçãoRESUMO
There are many benefits to drug delivery from drug-carrier nanostructure systems. It might be developed to carefully control drug release rates or to deliver a precise amount of a therapeutic substance to particular body areas. Self-assembling is the process by which molecules and nanoparticles spontaneously organize into organized clusters. For instance, proteins and peptides can interact with one another to create highly organized supramolecular structures with various properties, such as helical ribbons and fibrous scaffolds. Another advantage of self-assembly is that it may be effective with a variety of materials, including metals, oxides, inorganic salts, polymers, semiconductors, and even organic semiconductors. Fullerene, graphene, and carbon nanotubes (CNTs), three of the most fundamental classes of three-dimensionally self-assembling nanostructured carbon-based materials, are essential for the development of modern nanotechnologies. Self-assembled nanomaterials are used in a variety of fields, including nanotechnology, imaging, and biosensors. This review study begins with a summary of various major 3D nanomaterials, including graphene oxide, CNTs, and nanodiamond, as well as 3D self-assembled polyfunctionalized nanostructures and adaptable nanocarriers for drug delivery.
Assuntos
Portadores de Fármacos , Nanotubos de Carbono , Humanos , Portadores de Fármacos/química , Sistemas de Liberação de Medicamentos , Nanoestruturas/química , Nanotecnologia/métodosRESUMO
Acylation of biopharmaceuticals such as peptides has been identified as a major obstacle for the successful development of PLGA controlled release formulations. The purpose of this study was to develop a method to inhibit peptide acylation in poly(d,l-lactide-co-glycolide) (PLGA) formulations by reversibly and temporarily blocking the amine groups of a model peptide (octreotide) with a self-immolative protecting group (SIP), O-4-nitrophenyl-O'-4-acetoxybenzyl carbonate. The octreotide with two self-immolative protecting groups (OctdiSIP) on the N-terminus and lysine side chain was synthesized by reaction of the peptide with O-4-nitrophenyl-O'-4-acetoxybenzyl carbonate, purified by preparative RP-HPLC and characterized by mass spectrometry. Degradation studies of OctdiSIP in aqueous solutions of different pH values showed that protected octreotide was stable at low pH (pH 5) whereas the protecting group was eliminated at physiological pH, especially in the presence of an esterase, to generate native octreotide. OctdiSIP encapsulated in PLGA microspheres, prepared using a double emulsion solvent evaporation method, showed substantial inhibition of acylation as compared to the unprotected octreotide: 52.5% of unprotected octreotide was acylated after 50 days incubation of microspheres in PBS pH 7.4 at 37 °C, whereas OctdiSIP showed only 5.0% acylation in the same time frame. In conclusion, the incorporation of self-immolative protection groups provides a viable approach for inhibition of acylation of peptides in PLGA delivery systems.
Assuntos
Aminas/química , Ácido Láctico/química , Octreotida/química , Ácido Poliglicólico/química , Acilação , Cromatografia Líquida de Alta Pressão , Cromatografia de Fase Reversa , Hidrólise , Espectrometria de Massas , Microesferas , Copolímero de Ácido Poliláctico e Ácido PoliglicólicoRESUMO
Free radical polymerization is often used to prepare protein and peptide-loaded hydrogels for the design of controlled release systems and molecular imprinting materials. Peroxodisulfates (ammonium peroxodisulfates (APS) or potassium peroxodisulfates (KPS)) with N,N,N,N-tetramethylethylenediamine (TEMED) are frequently used as initiator and catalyst. However, exposure to these free radical polymerization reagents may lead to modification of the protein and peptide. In this work, we show the modification of lysine residues by ammonium peroxodisulfate (APS)/TEMED of the immunostimulant thymopentin (TP5). Parallel studies on a decapeptide and a library of 15 dipeptides were performed to reveal the mechanism of modification. LC-MS of APS/TEMED-exposed TP5 revealed a major reaction product with an increased mass (+12 Da) with respect to TP5. LC-MS(2) and LC-MS(3) were performed to obtain structural information on the modified peptide and localize the actual modification site. Interpretation of the obtained data demonstrates the formation of a methylene bridge between the lysine and arginine residue in the presence of TEMED, while replacing TEMED with a sodium bisulfite catalyst did not show this modification. Studies with the other peptides showed that the TEMED radical can induce methyleneation on peptides when lysine is next to arginine, proline, cysteine, aspargine, glutamine, histidine, tyrosine, tryptophan, and aspartic acid residues. Stability of peptides and protein needs to be considered when using APS/TEMED in in situ polymerization systems. The use of an alternative catalyst such as sodium bisulfite may preserve the chemical integrity of peptides during in situ polymerization.
Assuntos
Etilenodiaminas/química , Peptídeos/química , Polimerização , Sequência de Aminoácidos , Sulfato de Amônio/química , Radicais Livres/química , Peso Molecular , Biblioteca de Peptídeos , Estabilidade ProteicaRESUMO
PURPOSE: Polyesters with hydrophilic domains, i.e., poly(D,L-lactic-co-glycolic-co-hydroxymethyl glycolic acid) (PLGHMGA) and a multiblock copolymer of poly(ε-caprolactone)-PEG-poly(ε-caprolactone) and poly(L-lactide) ((PC-PEG-PC)-(PL)) are expected to cause less acylation of encapsulated peptides than fully hydrophobic matrices. Our purpose is to assess the extent and sites of acylation of octreotide loaded in microspheres using tandem mass spectrometry analysis. METHODS: Octreotide loaded microspheres were prepared by a double emulsion solvent evaporation technique. Release profiles of octreotide from hydrophilic microspheres were compared with that of PLGA microspheres. To scrutinize the structural information and localize the actual modification site(s) of octreotide, liquid chromatography ion-trap mass spectrometry (LC-ITMS) was performed on the acylated adducts. RESULTS: Hydrophilic microspheres showed less acylated adducts in comparison with PLGA microspheres. LC-MS/MS showed that besides the N-terminus and primary amine of lysine, the primary hydroxyl of the end group of octreotide was also subjected to acylation. Nucleophilic attack of the peptide can also occur to the carbamate bond presented in (PC-PEG-PC)-(PL) since 1,4-butanediisocyanate was used as the chain extender. CONCLUSIONS: Hydrophilic polyesters are promising systems for controlled release of peptide because substantially less acylation occurs in microspheres based on these polymers. LC-ITMS provided detailed structural information of octreotide modifications via mass analysis of ion fragments.
Assuntos
Octreotida/química , Poliésteres/química , Acilação , Cromatografia Líquida/métodos , Ácido Láctico/química , Microesferas , Ácido Poliglicólico/química , Copolímero de Ácido Poliláctico e Ácido Poliglicólico , Espectrometria de Massas em Tandem/métodosRESUMO
ß-propiolactone (BPL) is an alkylating agent used for inactivation of biological samples such as vaccines. Due to its known carcinogenic properties, complete hydrolysis of BPL is essential, and the detection of trace amounts is crucial. In this study a novel High-Performance Liquid Chromatography-Ultraviolet (HPLC-UV) method was developed. Rhodamine B hydrazide (RBH) was synthesized and utilized as a derivatizing reagent to react with BPL. The reaction was optimized in a weak acidic solution, resulting in a high yield. The separation of the RBH-derivatized BPL was achieved on a C8 column and detected by a UV detector at a wavelength of 560 nm. The method's validation demonstrated a high linearity (r2 > 0.99) over a concentration range of 0.5-50 µg/mL, with detection and quantification limits of 0.17 µg/mL and 0.5 µg/mL, respectively. The average recovery of samples was 85.20 % with a relative standard deviation (RSD) of 1.75 %. This method was successfully applied for BPL residue analysis in inactivated COVID-19 vaccines. This novel derivatization method offers a promising solution for monitoring BPL residues in the vaccine production process for quality control purposes and compliance with regulatory standards.
Assuntos
Vacinas contra COVID-19 , Limite de Detecção , Propiolactona , Rodaminas , Cromatografia Líquida de Alta Pressão/métodos , Propiolactona/química , Rodaminas/química , Reprodutibilidade dos Testes , Vacinas contra COVID-19/química , Vacinas de Produtos Inativados/química , Vacinas de Produtos Inativados/análise , Modelos Lineares , SARS-CoV-2/química , Humanos , Hidrazinas/química , Hidrazinas/análiseRESUMO
Due to the lack of pharmacopeia guidelines for injectable microspheres based on poly (D, L-lactide-co-glycolide) (PLGA), an internal method validation is a critical prerequisite for quality assurance. One of the essential issues of developing peptide-based drugs loaded PLGA microspheres is the precise determination of the amount of peptide drug entrapped in the microspheres. The aim of this study is the development and optimization of a method for measuring the drug content loading of PLGA microspheres using exenatide as a model peptide drug. Exenatide-loaded PLGA microspheres were prepared by a double emulsion solvent evaporation method. The extraction method to determine exenatide content in microspheres was optimized using Design of Experiments (DoE) approach. After the initial screening of six factors, using Fractional Factorial design (FFD), four of them, including type of organic solvent, buffer/organic solvent ratio (v/v), shaking time and pH, exhibited significant effects on the response, namely the exenatide loading, and a Box-Behnken design (BBD) was subsequently applied to obtain its optimum level. The optimum level for organic solvent volume, buffer/organic solvent ratio, shaking time, and pH were 4 ml, 1, 5.6 hrs, and pH 6, respectively. The exenatide content in microspheres under these conditions was 6.4 ± 0.0 (%w/w), whereas a value of 6.1% was predicted by the derived equation. This excellent agreement between the actual and the predicted value demonstrates that the fitted model can thus be used to determine the exenatide content.
Assuntos
Ácido Láctico , Ácido Poliglicólico , Exenatida , Copolímero de Ácido Poliláctico e Ácido Poliglicólico , Ácido Poliglicólico/química , Ácido Láctico/química , Poliglactina 910/química , Microesferas , Peptídeos/química , Solventes , Tamanho da PartículaRESUMO
Wound healing is one of the most complex biological processes. Studies show that Matrixyl (MTI), known as a cosmetic peptide, can lead to a faster healing process. The contribution of MTI to collagen formation during wound healing also depends on its mode of delivery and its release over time. Here, we investigate two modes of MTI-delivery system, the influence of MTI patch for wound healing application in comparison with MTI cream. In this study, animals were randomly divided into seven groups and studied for 21 days: patches containing two different concentrations of MTI (P-MTI-0.1 mg and P-MTI-1 mg), a cream containing MTI (C-MTI-1 mg), a patch (P-MTI-0), a cream with no MTI (C-MTI-0), a positive control (Comfeel), and a negative control (sham) group. To study the wound healing process, the change in collagen density, angiogenesis, epitheliogenesis, histopathology, immunohistochemical analysis, and wound area through imaging was monitored and measured. The macroscopic results showed that wound healing was improved from 63.5 up to 81.81% in treatment groups compared to that in the negative control group (P < 0.05 and P < 0.001). In addition, C-MTI-1 and P-MTI-1 had a larger impact on wound healing compared to that in the positive control group (Comfeel, P < 0.05). In hematoxylin and eosin (H&E) staining analysis, the rejuvenation of skin appendage was visible in both groups of cream and patches with MTI. According to the obtained results, the re-epithelialization had a higher range for the patch with MTI in comparison with cream containing MTI and positive control.
RESUMO
Acylation of peptides is a well-known but unwanted phenomenon in polyester matrices such as poly(d,l-lactic-co-glycolic acid) (PLGA) microspheres used as controlled release formulations. Acylation normally occurs on lysine residues and the N-terminus of the peptide. The purpose of the present work was to assess other possible acylation sites on peptides. Goserelin was used as a model peptide that lacks lysine and a free N-terminus, but contains other nucleophilic residues, i.e. serine, tyrosine and arginine, which potentially can be acylated. Goserelin loaded PLGA microspheres were prepared by a double emulsion solvent evaporation technique. Liquid chromatography ion-trap mass spectrometry (LC-ITMS) was used for determining and monitoring acylation of released goserelin. It is demonstrated that arginine is subjected to acylation with glycolic acid and lactic acid units of PLGA, which was followed by loss of NH3 from the guanidine group to obtain 2-oxazolin-4-one and 5-methyl-2-oxazolin-4-one residues with masses that are 41 and 55Da higher, respectively, than the native goserelin. There was no evidence for acylation of serine and tyrosine in goserelin. Our results demonstrate that beside lysine also acylation of arginine can occur in peptides and proteins that are loaded and released from PLGA matrixes.