Formulation and evaluation of anti-MRSA nanoemulsion loaded with Achyrocline satureioides: a new sustainable strategy for the bovine mastitis

Abstract

Methicillin-resistant Staphylococcus aureus (MRSA) causes mastitis in dairy cattle with serious economic and public health significance. This study developed nanoemulsions of Linum usitatissimun oil loaded with Achyrocline satureioides (macela) extract and investigated their in vitro antimicrobial activity against MRSA. Macela-nanoemulsions (NE-ML) were prepared using high-pressure homogenization (HPH) with different proportions of flaxseed oil, Tween 80 and crude extract. Four majoritarian flavonoids were identified in the macela extract: 3-O methylquercetin, achyrobichalcone, quercetin and luteolin (187.3 ± 0.1, 155.4 ± 11.6, 76.3 ± 0.1 and 30.4 ± 0.0 μg ml−1, respectively). NE-ML nanoemulsions were successfully obtained by the HPH method and showed a milky aspect with yellowish color. The mean particle size was around 200 nm with monodisperse distribution (PdI < 0.2), remaining stable for 160 days at room temperature. When analyzed on a LUMiSizer high-end dispersion analyzer, low values were found (≤0.5), indicating high stability index, mainly for NE-ML1:5 (0.2). The encapsulation efficiency of macela-nanoemulsions was greater than 94%, considering the four chemical compounds from extract. Minimum inhibitory concentration (MIC) against planktonic bacteria, inhibition of biofilm formation (MBIC), and eradication of MRSA biofilms (MBEC) were determined through in vitro tests on microplates. The MIC of NE-ML against planktonic MRSA showed values ranging from 1.2 to 10% (v/v), while blank-nanoemulsions (NE-B, without macela extract) showed values ranging from 6 to 50% (v/v). MBIC and MBEC of NE-ML were 25 and 80% (v/v), respectively. MBIC showed a mass reduction greater than 64%, and MBEC showed a mass reduction greater than 73%. Macela-nanoemulsions (NE-ML), mainly NE-ML1:5, showed high antimicrobial activity and appeared to represent a new alternative of sustainable antimicrobial product for the control of MRSA. Since this innovative nanoemulsion can impact animal health, future research should include in vitro and in vivo studies to evaluate intramammary therapy and control of MRSA infections in organic and agroecological milk production systems.