从短芽孢杆菌中分离出的新型 BrSPR-20-P1 抗菌肽 (P1-AMP; NH 2 -VVVNVLVKVLPPPVV-COOH)。 SPR-20被封装在含有不同比例的L -α-磷脂酰胆碱(PC)和胆固醇(CH)的脂质体中。将 P1-AMP 脂质体掺入壳聚糖水凝胶中,以达到 0.02% 的肽浓度。 P1-AMP 已经过抗菌和体外伤口愈合活性测试。研究了脂质体和水凝胶的理化特性,包括体外药物释放、渗透性、细胞毒性、抗菌活性和稳定性研究。 P1-AMP 显示出比阴性对照更高的抗菌和伤口愈合活性。 P1-AMP 在角质形成细胞系中的毒性测试显示,在 1.96–1000 μg mL -1的浓度范围内,细胞活力为 100%。空脂质体的平均粒径范围为324.5±8.6至1823.7±288.2 nm。 P1-AMP脂质体的尺寸范围为378.6±14.0至2363.0±255.6 nm。空白脂质体的zeta电位范围为-40.43±2.51至-60.17±0.93 mV,负载肽的脂质体则降低至-57.33±0.72至-70.33±0.15 mV。 SEM图像显示脂质体为卵形球体,表面光滑。所选择的配方由 PC 与 CH 以 18:1 的比例组成(配方 F3),具有最高的包封效果和较小的粒径,并具有可接受的 zeta 电位。开发的 P1-AMP 脂质体负载水凝胶呈现淡黄色透明外观,粘度为 758.0 ± 149.8 cPs。 P1-AMP 从负载 P1-AMP 的脂质体水凝胶制剂中快速释放。与单独的 P1-AMP 或未掺入脂质体的水凝胶中的 P1-AMP 相比,负载 P1-AMP 的脂质体显示出高渗透性。负载P1-AMP的脂质体水凝胶对金黄色葡萄球菌和耐甲氧西林金黄色葡萄球菌(MRSA)的最低抑制浓度(MIC)为2μgmL -1 ,与P1-AMP相当。分别在 10× 和 5× MIC 孵育 6 小时和 12 小时后完全杀死金黄色葡萄球菌。该制剂不会对测试的角质形成细胞产生细胞毒性,并在研究条件下保持稳定至少 6 个月。
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BrSPR-20-P1 peptide isolated from Brevibacillus sp. developed into liposomal hydrogel as a potential topical antimicrobial agent
A novel BrSPR-20-P1 antimicrobial peptide (P1-AMP; NH2-VVVNVLVKVLPPPVV-COOH) isolated from Brevibacillus sp. SPR-20 was encapsulated in a liposome containing varying proportions of L-α-phosphatidylcholine (PC) and cholesterol (CH). P1-AMP liposomes were incorporated into a chitosan hydrogel to achieve a peptide concentration of 0.02%. P1-AMP has been tested for its antibacterial and in vitro wound healing activities. The physicochemical characteristics of liposomes and hydrogel were investigated, including in vitro drug release, permeability, cell toxicity, antimicrobial activities, and stability studies. P1-AMP showed higher antimicrobial and wound-healing activities than the negative control. A toxicity test of P1-AMP in keratinocyte cell lines revealed cell viability of 100% at a concentration range of 1.96–1000 μg mL−1. The empty liposomes exhibited an average particle size ranging from 324.5 ± 8.6 to 1823.7 ± 288.2 nm. The size range of P1-AMP liposomes was 378.6 ± 14.0 to 2363.0 ± 255.6 nm. The zeta potential of the blank liposome ranged from −40.43 ± 2.51 to −60.17 ± 0.93 mV and it decreased to −57.33 ± 0.72 to −70.33 ± 0.15 mV of the liposome loaded with peptide. SEM images showed liposomes were ovoid spheres with smooth surfaces. The chosen formulation, composed of PC to CH in an 18 : 1 ratio (formulation F3), had the highest entrapment effectiveness with small particle size and possessed an acceptable zeta potential. The developed P1-AMP liposome-loaded hydrogels exhibited a yellowish-clear appearance with a viscosity of 758.0 ± 149.8 cPs. The P1-AMP was rapidly released from the P1-AMP-loaded liposome hydrogel formulation. The P1-AMP-loaded liposome showed high permeability compared to P1-AMP alone or P1-AMP in hydrogel without the incorporation of liposomes. The minimum inhibitory concentration (MIC) against Staphylococcus aureus and methicillin-resistant S. aureus (MRSA) of P1-AMP-loaded liposome hydrogel was 2 μg mL−1, equivalent to P1-AMP. It completely killed S. aureus at 10× and 5× MIC after 6 and 12 h of incubation, respectively. The formulation did not induce cytotoxicity to the tested keratinocyte cell and remained stable for at least 6 months under the studied conditions.