Lipozomal juglon formülasyonu geliştirilmesi ve domates patojenleri üzerine antibakteriyel aktivitesinin belirlenmesi
Küçük Resim Yok
Tarih
2024
Yazarlar
Dergi Başlığı
Dergi ISSN
Cilt Başlığı
Yayıncı
Bursa Teknik Üniversitesi, Lisansüstü Eğitim Enstitüsü
Erişim Hakkı
info:eu-repo/semantics/embargoedAccess
Özet
Bitki temelli doğal antibakteriyel ajanlar, ilaca dirençli suşları kontrol etmede etkin rol oynamaktadır. Bu aktiflerden biri olan juglon, bitki hastalıklarının mücadelesinde kullanılmak üzere, güçlü antimikrobiyal özelliklere ve geniş uygulama alanlarına sahip bir molekül olarak öne çıkmaktadır. Suda çözünmemesi, kolay oksidasyonu ve zayıf biyoyararlanımı gibi dezavantajlarının bulunması ise endüstride kullanımını sınırlamaktadır. Juglonun dağıtımı için lipozom gibi nano-taşıyıcıların kullanılması, bu kısıtlamaların üstesinden gelebilecek bir stratejidir. Hidrofobik maddelerin lipozomal enkapsülasyonunda karşılaşılan problemlerin üstesinden gelmek için araştırmacılar siklodekstrinleri lipozomal ilaç formülasyonlarına dahil ederek özelliklerini iyileştirmektedir. Bu tez kapsamında; üretilen juglon/β-siklodekstrin inklüzyon kompleksinin, literatürde ilk kez lipozomlara enkapsülasyonunun çalışılması amaçlanmıştır. Kompleksin inklüzyon oranı, yükleme kapasitesi ve reaksiyon verimi incelenmiş, formülasyonun enkapsülasyon etkinliği, boyut, polidispersite indeksi (PDI), ve zeta potansiyeli belirlenmiştir. Formülasyonların fizikokimyasal karakterizasyonu için Fourier dönüşümlü kızılötesi spektroskopisi (FT-IR), morfolojik karakterizasyonu için taramalı elektron mikroskobu (SEM) kullanılmıştır. Juglon salımı diyaliz yöntemi ile in vitro incelenmiştir. Pseudomonas syringae pv. tomato (SH-1), Xanthomonas euvesicatoria (SH-2) ve Clavibacter michiganensis subsp. michiganensis (SH-3) bakteri suşları üzerinde agar disk difüzyon ve mikrodilüsyon yöntemleri kullanılarak antibakteriyel aktiviteleri in vitro incelenmiştir. Antibakteriyel aktivitenin görüldüğü konsantrasyon belirlendiğinde bu konsantrasyon baz alınarak uygulanan formülasyonların domates tohum çimlenmesi üzerine etkisi 14 gün boyunca gözlemlenmiştir. Hazırlanan inklüzyon komplekslerinin inklüzyon oranı %79,3, yükleme kapasitesi %11,8 ve reaksiyon verimi %89,4 olarak, juglon/β-siklodekstrin lipozomal formülasyonunun enkapsülasyon etkinliği %72,88, boyutu 106,1 ± 0,5 nm, PDI değeri 0,247 ± 0,012 ve zeta potansiyeli -49,2 ± 1,36 mV olarak ölçülmüştür. FTIR sonuçları juglonun çoğunlukla -CD kavitesinde olduğunu ve lipozomal enkapsülasyonun da başarılı olduğunu göstermektedir. SEM sonuçları yüzeyleri düz, düzgün bir boyut dağılımda, yuvarlak partiküllerin oluşturulduğunu doğrulamıştır. Salım çalışması lipozomlardan juglon salımının bifazik salım profilinde olduğunu ve kümülatif salımın 72 saatte %46,73 olduğunu göstermiştir. Agar disk difüzyon testine göre J/-CD/L formülasyonu Pst üzerine 9 mm, Xeu üzerine 10,6 mm ve Cmm üzerine 8 mm inhibisyon zonu göstermiştir. J/β-CD/L'nin SH-1, SH-2 ve SH-3 üzerindeki MİK değerleri sırasıyla 68,93 µg/mL, 34,47 µg/mL ve 68,93 µg/mL olarak, MBK değerleri ise 68,93 µg/mL, 68,93 µg/mL ve 137,87µg/mL olarak hesaplanmıştır. J/β-CD/L'nin kontrollü salım yapması nedeniyle test edilen her üç patojen bakteri türü için de serbest juglona göre antibakteriyel aktivitesinin 2 kat fazla olduğu belirlenmiştir. Tohum çimlenme testine göre 14. günün sonunda en çok çimlenen set 73,75 µg/ml serbest juglon içeren; 0,625 mg/mL J/β-CD/L konsantrasyonunda formülasyon ile muamele edilmiş tohumlar olmuş ve çimlenme yüzdesi de %86,67 olarak belirlenmiştir. Sonuç olarak; tez kapsamında geliştirilen juglon/β-CD'in lipozomal enkapsülasyonu hidrofobik bir molekül olan juglonun çözünürlüğünü arttırmış ve maddenin biyoyararlanımını iyileştirmiştir.
Plant-based natural antibacterial agents play an effective role in controlling drug-resistant strains. One of these actives, juglone, stands out as a molecule with strong antimicrobial properties and wide application areas for the control of plant diseases. Its disadvantages, such as insolubility in water, easy oxidation and poor bioavailability, limit its use in industry. The use of nanocarriers such as liposomes for the delivery of juglone is a strategy that can overcome these limitations. To overcome the problems encountered in liposomal encapsulation of hydrophobic substances, researchers are incorporating cyclodextrins into liposomal drug formulations to improve their properties. In this thesis; it was aimed to study the encapsulation of the juglon/β-cyclodextrin inclusion complex produced for the first time in the literature. The inclusion ratio, loading capacity and reaction efficiency of the complex were investigated, and the encapsulation efficiency, size, polydispersity index (PDI), and zeta potential of the formulation were determined. Fourier transform infrared spectroscopy (FT-IR) was used for physicochemical characterization, and scanning electron microscopy (SEM) was used for morphological characterization.Juglone release was studied in vitro by dialysis method. Antibacterial activities of Pseudomonas syringae pv. tomato (SH-1), Xanthomonas euvesicatoria (SH-2) and Clavibacter michiganensis subsp. michiganensis (SH-3) were investigated in vitro using agar disk diffusion and microdilution methods. When the concentration at which antibacterial activity was observed was determined, the effect of the formulations applied based on this concentration on tomato seed germination was observed for 14 days. The inclusion rate, loading capacity and reaction efficiency of the prepared inclusion complexes were 79.3%, 11.8% and 89.4%, respectively. The encapsulation efficiency of the juglone/β-cyclodextrin liposomal formulation was 72.88%, the size was 106.1 ± 0.5 nm, the PDI value was 0.247 ± 0.012, and the zeta potential was -49.2 ± 1.36 mV. FTIR results show that the juglone is mostly in the β-CD cavity and liposomal encapsulation is also successful. SEM results confirm the formation of round particles with smooth surfaces and a uniform size distribution. The release study showed that the release of juglone from liposomes had a biphasic release profile and the cumulative release was 46.73% at 72 hours. According to the agar disk diffusion test, the J/β-CD/L formulation showed an inhibition zone of 9 mm on Pst, 10.6 mm on Xeu and 8 mm on Cmm. The MIC values of J/β-CD/L molecule on SH-1, SH-2 and SH-3 were 68.93 µg/mL, 34.47 µg/mL and 68.93 µg/mL, respectively, and the MBC values were 68.93 µg/mL, 68.93 µg/mL and 137.87 µg/mL, respectively. The controlled release of J/β-CD/L resulted in a determined antibacterial activity that was found to be 2 times higher than that of free juglone for all three pathogenic bacterial species tested. According to the seed germination test, the most germinated set at the end of the 14th day was the seeds treated with the formulation containing 73.75 µg/ml free juglone at a concentration of 0.625 mg/mL J/β-CD/L and the germination percentage was 86.67%. In summary, the liposomal encapsulation of J/β-CD/L, developed as part of the thesis, resulted in enhanced solubility of juglone, a hydrophobic molecule, and contributed to improved bioavailability of the substance.
Plant-based natural antibacterial agents play an effective role in controlling drug-resistant strains. One of these actives, juglone, stands out as a molecule with strong antimicrobial properties and wide application areas for the control of plant diseases. Its disadvantages, such as insolubility in water, easy oxidation and poor bioavailability, limit its use in industry. The use of nanocarriers such as liposomes for the delivery of juglone is a strategy that can overcome these limitations. To overcome the problems encountered in liposomal encapsulation of hydrophobic substances, researchers are incorporating cyclodextrins into liposomal drug formulations to improve their properties. In this thesis; it was aimed to study the encapsulation of the juglon/β-cyclodextrin inclusion complex produced for the first time in the literature. The inclusion ratio, loading capacity and reaction efficiency of the complex were investigated, and the encapsulation efficiency, size, polydispersity index (PDI), and zeta potential of the formulation were determined. Fourier transform infrared spectroscopy (FT-IR) was used for physicochemical characterization, and scanning electron microscopy (SEM) was used for morphological characterization.Juglone release was studied in vitro by dialysis method. Antibacterial activities of Pseudomonas syringae pv. tomato (SH-1), Xanthomonas euvesicatoria (SH-2) and Clavibacter michiganensis subsp. michiganensis (SH-3) were investigated in vitro using agar disk diffusion and microdilution methods. When the concentration at which antibacterial activity was observed was determined, the effect of the formulations applied based on this concentration on tomato seed germination was observed for 14 days. The inclusion rate, loading capacity and reaction efficiency of the prepared inclusion complexes were 79.3%, 11.8% and 89.4%, respectively. The encapsulation efficiency of the juglone/β-cyclodextrin liposomal formulation was 72.88%, the size was 106.1 ± 0.5 nm, the PDI value was 0.247 ± 0.012, and the zeta potential was -49.2 ± 1.36 mV. FTIR results show that the juglone is mostly in the β-CD cavity and liposomal encapsulation is also successful. SEM results confirm the formation of round particles with smooth surfaces and a uniform size distribution. The release study showed that the release of juglone from liposomes had a biphasic release profile and the cumulative release was 46.73% at 72 hours. According to the agar disk diffusion test, the J/β-CD/L formulation showed an inhibition zone of 9 mm on Pst, 10.6 mm on Xeu and 8 mm on Cmm. The MIC values of J/β-CD/L molecule on SH-1, SH-2 and SH-3 were 68.93 µg/mL, 34.47 µg/mL and 68.93 µg/mL, respectively, and the MBC values were 68.93 µg/mL, 68.93 µg/mL and 137.87 µg/mL, respectively. The controlled release of J/β-CD/L resulted in a determined antibacterial activity that was found to be 2 times higher than that of free juglone for all three pathogenic bacterial species tested. According to the seed germination test, the most germinated set at the end of the 14th day was the seeds treated with the formulation containing 73.75 µg/ml free juglone at a concentration of 0.625 mg/mL J/β-CD/L and the germination percentage was 86.67%. In summary, the liposomal encapsulation of J/β-CD/L, developed as part of the thesis, resulted in enhanced solubility of juglone, a hydrophobic molecule, and contributed to improved bioavailability of the substance.
Açıklama
01.02.2025 tarihine kadar kullanımı yazar tarafından kısıtlanmıştır.
Anahtar Kelimeler
Biyomühendislik, Bioengineering
