Bioactivity-guided isolation of inositol as acetylcholinesterase inhibitory from endemic Campanula baskilensis Behcet: In vitro bioactivity, PCA analysis, and silico supporting studies

Sarmad Marah; Yaşar İpek; Tevfik Ozen; İbrahim Demirtas; Lütfi Behçet

doi:10.18016/ksutarimdoga.vi.1632935

EN TR

Bioactivity-guided isolation of inositol as acetylcholinesterase inhibitory from endemic Campanula baskilensis Behcet: In vitro bioactivity, PCA analysis, and silico supporting studies

Abstract

This study aimed to identify the bioactivity-guided molecule in the fractions of Campanula baskilensis leaf methanol: chloroform extract. The bioactivity of leaf fractions was investigated to assess and isolate bioactive molecule/molecules and structural configurations. Fractionation and isolation processes were done using advanced column chromatography techniques. In-vitro bioactivity tests were applied, including enzyme inhibition, antibacterial, and DNA protection activities. The isolated compound was characterized using the NMR technique. In silico analyses were investigated using molecular docking, molecular dynamics, and final-state free energy calculations. 14 different fractions were obtained (F1-F14) through the fractionation. F12 has the highest AChE inhibition (IC50; 6.97±2.90 μg/mL), F6 has significant inhibition against carbonic anhydrase and -amylase (IC50; 5.61±0.01 and 18.82±1.48 μg/mL). F12 and F11 have the highest antibacterial activity against E. coli (15.40±1.10 and 13.00±0.80 mm). F12 and F5 fractions have the highest protection activity in plasmid DNA, and F6 has the highest deoxyribose protection activity. Many fractions have high and varied bioactivity due to the bioactive compound components, as in F12. Principal component analysis showed that F12 positively correlated with the high inhibition activity for several bacteria and enzymes and high DNA protection. Therefore, further fractionation was applied using Sephadex LH-20 with ethyl acetate:methanol: hexane (5:5:1) to F12. Inositol was isolated according to results from the obtained fraction; the molecule characterization was clarified using the H-NMR and C13-NMR spectra. Molecular docking results showed binding between inositol and AChE. Further, molecular dynamics results showed the stability of inositol-AChE within 100 nanoseconds, and the energy calculations (gmx-MMPBSA) showed the strength of this interaction.

Keywords

Endemik Campanula baskilensis Behçet bitkisinden asetilkolinesteraz inhibitörü olarak inositolün biyoaktivite rehberliğinde izolasyonu: İn vitro biyoaktivite, PCA analizi ve İn silico destekleyici çalışmalar

Öz

Bu çalışmada Campanula baskilensis yaprak metanal:kloroform ekstraktının fraksiyonlarında biyoaktivite-yönlendirmeli moleküle ulaşılması amaçlanmıştır. Biyoaktif molekül/moleküllere ve yapısal konfigürasyona ulaşmak ve izole etmek için yaprak fraksiyonlarının biyoaktivitesi araştırıldı. Fraksiyonlama ve izolasyon işlemleri gelişmiş kolon kromatografisi tekniği kullanılarak gerçekleştirildi. Fraksiyon örnekleri için enzim inhibisyonu, antibakteriyel ve DNA koruma aktiviteler dahil olmak üzere in vitro biyoaktivite testleri uygulandı. İzole edilen bileşik NMR tekniği kullanılarak karakterize edildi. İn silico analizlerle moleküler doking, moleküler dinamikler ve son durum serbest enerji hesaplamalarını araştırıldı. Fraksiyonlama işlemi sonucunda 14 farklı fraksiyon elde edildi (F1-F14). F12 AChE karşı (IC50; 6.97±2.90 μg/mL), F6 karbonik anhidraz ve -amilaza karşı (IC50; 5.61±0.01 ve 18.82±1.48 μg/mL) yüksek inhibisyon aktivite gösterdi. F12 ve F11 E. coli'ye karşı en yüksek antibakteriyel aktivite gösterdi (15.40±1.10 ve 13.00±0.80 mm). F12 ve F5 fraksiyonları plazmit DNA'sında en yüksek koruma aktivitesine sahiptir ve F6, deoksiribozu korumak için en yüksek aktiviteye sahiptir. Birçok fraksiyonun, F12'de olduğu gibi, yüksek biyoaktif bileşikler içerdiğinden yüksek ve çeşitli bir biyoaktiviteye sahip olduğu gösterilmiştir. F12'nin test edilen çeşitli bakteri ve enzimler için yüksek inhibisyon aktivitesinin yanı sıra yüksek DNA koruması ile ana bileşen analizi pozitif korelasyon gösterdiği. Bu nedenle F12'ye Sephadex LH-20 ve etil asetat:metanol:hekzan (5:5:1) kullanılarak daha ileri fraksiyonlama ilerletildi. F12'den inositol izole edildi ve karakterizasyonu H-NMR ve C13-NMR spektrumları kullanılarak açıklandı. Moleküler doking sonuçlarına göre inositolün AChE enzimine bağlanabileceğini gösterdi. Ayrıca, moleküler dinamik sonuçları inositol-AChE'nin 100 nanosaniye içinde kararlı olduğunu gösterdi ve enerji hesaplamaları (gmx-MMPBSA) bu etkileşimin gücünü gösterdi.

Anahtar Kelimeler

Supporting Institution

Ondokuz Mayis University

Project Number

PYO.FEN.1904.20.003 ve PYO.FEN.1904.19.006

Ethical Statement

yok

Thanks

The financial support of this study was provided by the Scientific Research Project Commission Fund of Ondokuz Mayis University, Turkey (Samsun), under the project numbers of PYO.FEN.1904.20.003 and PYO.FEN.1904.19.006.

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Details

Primary Language

English

Subjects

Pharmaceutical Botany

Journal Section

Research Article

Authors

Sarmad Marah
0000-0001-6765-4605
Iraq

Yaşar İpek
0000-0002-1041-267X
Türkiye

Tevfik Ozen ^*
0000-0003-0133-5630
Türkiye

İbrahim Demirtas
0000-0001-8946-647X
Türkiye

Lütfi Behçet
0000-0001-8334-7816
Türkiye

Early Pub Date

May 1, 2025

Publication Date

June 9, 2025

Submission Date

February 5, 2025

Acceptance Date

April 10, 2025

Published in Issue

Year 2025 Volume: 28 Number: 3

DOI

https://doi.org/10.18016/ksutarimdoga.vi.1632935

IZ

https://izlik.org/JA96ZP26FL

Cite

RIS / Bibtex

APA

Marah, S., İpek, Y., Ozen, T., Demirtas, İ., & Behçet, L. (2025). Bioactivity-guided isolation of inositol as acetylcholinesterase inhibitory from endemic Campanula baskilensis Behcet: In vitro bioactivity, PCA analysis, and silico supporting studies. Kahramanmaraş Sütçü İmam Üniversitesi Tarım Ve Doğa Dergisi, 28(3), 717-735. https://doi.org/10.18016/ksutarimdoga.vi.1632935

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