# Blunt-Nosed Viper (Macrovipera lebetinus) Venom: Proteomic Composition, Antioxidant Properties, and Anti-Alzheimer Assessment

## TL;DR

This study explores the venom of the blunt-nosed viper, analyzing its protein composition and antioxidant and anti-Alzheimer properties.

## Contribution

The study identifies nonprotein components in viper venom that may contribute to its biological activities, expanding beyond traditional protein-based analysis.

## Key findings

- Methanol and ethyl acetate fractions of the venom showed significant antioxidant and enzyme inhibitory properties.
- Nineteen unique proteins from eight families were identified in the crude venom and its fractions.
- The ethyl acetate fraction exhibited strong acetylcholinesterase inhibition comparable to the drug donepezil.

## Abstract

This study examines the antioxidant and enzyme (acetylcholinesterase,
butyrylcholinesterase) inhibitory properties of crude and different
solvent fractions of Macrovipera lebetinus obtusa venom and investigates the proteomic composition of its fractions
(ethyl acetate, methanol, and water) employing liquid chromatography-tandem
mass spectrometry (LC-MS/MS) analysis. The results revealed many peptide
and protein families, suggesting considerable biological potential.
Nineteen unique protein identifications belonging to eight families
were achieved in the crude venom, along with additional identifications
in particular fractions. Although major bioactive molecules in snake
venoms are proteins and peptides, which dissolve readily in water-based
solvents and can be separated by chromatography, here we also wondered
if nonprotein components with different polarities and solubilization
characteristics may also contribute to their activities. To this end,
sequential fractionation with solvents of increasing polarity was
performed. Utilizing techniques such as metal chelation, reducing
power assessment, 2,2′-azinobis-3-ethylbenzothiazoline-6-sulfonic
acid (ABTS), and free radical scavenging capabilities of 2,2-diphenyl-1-picrylhydrazyl
(DPPH), it was determined that the methanol and ethyl acetate fractions
demonstrated antioxidant activity, whereas other fractions displayed
minimal effects. The methanol fraction of M. lebetinus
obtusa venom exhibited significant metal chelation
(60.77 ± 0.10%) and showed substantial antioxidant activity (reducing
power) with an EC50 of 281.30 μg/mL. The ethyl acetate
fraction also showed a reducing effect with an EC50 of
263.20 μg/mL. None of the fractions and crude venom showed antioxidant
activity in the DPPH free radical scavenging assay and the ABTS·+ radical cation decolorization test. The ethyl acetate
fraction exhibited significant acetylcholinesterase (AChE) inhibitory
activity (75.39 ± 0.30%) comparable to donepezil (98.60 ±
0.10%); however, it displayed reduced butyrylcholinesterase (BuChE)
inhibitory activity (20.65 ± 4.50%). The results highlight the
potential of M. lebetinus obtusa venom
fractions as sources of bioactive chemicals and the possible contribution
of nonpolypeptide content to its activities.