Phytochemical composition and antimicrobial potential of Bertoni extract and its topical spray formulation against animal skin pathogens.

BACKGROUND AND AIM

The rise of antimicrobial resistance in veterinary medicine is a significant concern, particularly for pathogens responsible for skin infections. Although Bertoni (stevia) has demonstrated effective antimicrobial properties, there is limited research on its efficacy against animal skin pathogens. This study aimed to identify natural compounds in stevia extract, develop a topical spray formulation, and assess its effectiveness against six common bacterial and fungal pathogens associated with animal skin infections.

MATERIALS AND METHODS

The aerial parts of stevia plants were extracted using hexane in a Soxhlet apparatus. Total phenolic and flavonoid contents were quantified using colorimetric assays. The volatile oil content was analyzed using gas chromatography-mass spectrometry (GC-MS). The antimicrobial activity of stevia extract against , , , , , and was evaluated using broth microdilution and time-kill tests. Environmental scanning electron microscopy (E-SEM) and leakage studies were conducted to assess the extract's impact on microbial morphology and cell membrane integrity. The antimicrobial efficacy and stability of a topical spray formulation containing stevia extract were evaluated using time-kill and freeze-thaw testing.

RESULTS

The stevia extract yield was 3.59% of the dry plant weight with 259.96 ± 23.66 mg gallic acid equivalent (GAE)/g extract of total phenolics and 247.41 ± 19.92 mg quercetin equivalent (QE)/g extract of total flavonoids. GC-MS analysis identified major volatile components, including N-acetyl-14, 15, 16-trinorlabd-8(17)-en-13-amine (37.70% of peak area), phytol (11.02% of peak area), (-)-spathulenol (9.46% of peak area), n-hexadecanoic acid (8.01% of peak area), and (diphenylphosphinoyloxymethyl) dimethylsilane (7.59% of peak area). The minimum inhibitory concentration of the extract against the tested microorganisms ranged from 0.25 to 128.00 mg/mL. Time-kill kinetics exhibited time- and concentration-dependent germicidal effects. E-SEM and cell leakage analyses indicated that stevia extract compromised microbial cell membrane integrity. A spray formulation containing 10% w/w stevia extract displayed excellent eradication efficacy, achieving a 99.9999% reduction of and a 99.999% reduction of and dermatophytes, with good stability after six freeze-thaw cycles.

CONCLUSION

Stevia extract is an effective antimicrobial against , , , , and . Future research will investigate the pharmaceutical properties and toxicity profiles of purified compounds and determine appropriate dosages and clinical efficacy.