Roque-Borda Cesar Augusto, Silva Hanyeny Raiely Leite, Crusca Junior Edson, Serafim Jéssica Aparecida, Meneguin Andréia Bagliotti, Chorilli Marlus, Macedo Wagner Costa, Teixeira Silvio Rainho, Guastalli Elisabete Aparecida Lopes, Soares Nilce Maria, Blair Jessica M A, Pikramenou Zoe, Vicente Eduardo Festozo
São Paulo State University (Unesp), School of Agricultural and Veterinarian Sciences, Jaboticabal, São Paulo CEP 14884-900, Brazil.
São Paulo State University (Unesp), Institute of Chemistry, Araraquara, São Paulo CEP 14800-900, Brazil.
Int J Biol Macromol. 2021 Jul 31;183:1236-1247. doi: 10.1016/j.ijbiomac.2021.05.011. Epub 2021 May 14.
Microencapsulation is a potential biotechnological tool, which can overcome antimicrobial peptides (AMP) instabilities and reduce toxic side effects. Thus, this study evaluates the antibacterial activities of the Ctx(Ile)-Ha AMP against multidrug-resistant (MDR) and non-resistant bacteria and develop and characterize peptide-loaded microparticles coated with the enteric polymers hydroxypropylmethylcellulose acetate succinate (HPMCAS) and hydroxypropylmethylcellulose phthalate (HPMCP). Ctx(Ile)-Ha was obtained by solid phase peptide synthesis (SPPS) method, purified and characterized by HPLC and Mass Spectrometry. The peptide exhibited potent antibiotic activities against Salmonella enteritidis, Salmonella typhimurium, Pseudomonas aeruginosa (MDR), Acinetobacter baumannii (MDR), and Staphylococcus aureus (MDR). Ctx(Ile)-Ha microencapsulation was performed by ionic gelation with high efficiency, maintaining the physical-chemical stability. Ctx(Ile)-Ha coated-microparticles were characterized by DSC, TGA, FTIR-Raman, XRD and SEM. Hemolytic activity assay demonstrated that hemolysis was decreased up to 95% compared to single molecule. In addition, in vitro release control profile simulating different portions of gastrointestinal tract was performed and showed the microcapsules' ability to protect the peptide and release it in the intestine, aiming pathogen's location, mainly by Salmonella sp. Therefore, use of microencapsulated Ctx(Ile)-Ha can be allowed as an antimicrobial controller in monogastric animal production as an oral feed additive (antimicrobial controller), being a valuable option for molecules with low therapeutic indexes or high hemolytic rates.
微囊化是一种潜在的生物技术工具，它可以克服抗菌肽（AMP）的不稳定性并减少毒副作用。因此，本研究评估了Ctx（Ile）-Ha AMP对多重耐药（MDR）菌和非耐药菌的抗菌活性，并制备和表征了负载该肽的微粒，这些微粒用肠溶聚合物醋酸琥珀酸羟丙甲纤维素（HPMCAS）和邻苯二甲酸羟丙甲纤维素（HPMCP）包衣。Ctx（Ile）-Ha通过固相肽合成（SPPS）方法获得，经高效液相色谱（HPLC）和质谱法纯化及表征。该肽对肠炎沙门氏菌、鼠伤寒沙门氏菌、铜绿假单胞菌（MDR）、鲍曼不动杆菌（MDR）和金黄色葡萄球菌（MDR）表现出强大的抗菌活性。通过离子凝胶法高效进行Ctx（Ile）-Ha的微囊化，保持其物理化学稳定性。通过差示扫描量热法（DSC）、热重分析法（TGA）、傅里叶变换红外-拉曼光谱法（FTIR-Raman）、X射线衍射法（XRD）和扫描电子显微镜法（SEM）对Ctx（Ile）-Ha包衣微粒进行表征。溶血活性测定表明，与单分子相比，溶血率降低了95%。此外，进行了模拟胃肠道不同部位的体外释放控制曲线研究，结果表明微胶囊能够保护该肽并在肠道中释放，主要针对病原菌的位置，尤其是沙门氏菌属。因此，微囊化的Ctx（Ile）-Ha作为一种口服饲料添加剂（抗菌控制剂），可被允许用于单胃动物生产中的抗菌控制，对于治疗指数低或溶血率高分子来说是一个有价值的选择。
