Rosemary essential oil enhances culture establishment and inhibits contamination and enzymatic browning: Applications for in vitro propagation of Aloe vera L.
Aloe vera L. is used largely in the pharmaceutical, cosmetic, and agro-food industries. The in vitro technique was proposed as an alternative method for large-scale propagation and/or conservation breeding programmes in response to the increasing demand for Aloe vera. Unfortunately, a major problem in the application of plant tissue culture is the browning of cultured tissues and microbial contamination. This work investigates for the first time the efficacy of using Essential Oils (EOs) to control enzymatic browning and contamination of cultures from Aloe vera. Murashige and Skoog (MS) (1962) medium supplemented with 1 mg/L β-indole butyric acid (IBA) was used as a basic medium for A. vera for in vitro culture. The effect of the addition of charcoal (at 0.5 g/L), thyme (Thymus vulgaris L.) and rosemary (Rosmarinus officinalis L.) essential oils (at concentrations of 0.1 and 0.2%) to the basic medium was assessed. While thyme Essential Oil (EO) inhibited explant growth, rosemary EO induced the highest explant survival percentage, the lowest browning diffusion and contamination percentage. Moreover, a second experiment was conducted to determine the optimal rosemary EO concentration (0.025, 0.05, 0.075, and 0.1%). Explant survival percentage was 100% after four weeks of culture with rosemary EO concentrations of 0.05, 0.075, and 0.1% with no signs of browning. The lowest infection percentage (10%) was observed for media containing 0.075 and 0.1% of rosemary EO. The highest average number of leaves per explant was 3.71 with 0.1% rosemary EO and the greatest leaf length was 3.18 cm with 0.05%. The analytical techniques used to detect rosemary EO compounds were GC-FID (gas chromatography - flame ionization detection) and GC–MS (gas chromatography-mass spectrometry). The main constituents found were 1,8-cineole (53.92%), α-pinene (15.86%), β-pinene (10.79%), and camphor (7.99%). Additionally, the antioxidant activity of the rosemary EO was determined by the DPPH (Free radical-scavenging activity) (24.66 µg mL−1) and FRAP (Ferric reducing antioxidant power) (35.16 mmol Fe2+L − 1) tests. The chemical composition together with the antioxidant effect and the most widely recognized anti-infectious properties of the used rosemary EO could explain its role in the inhibition of both enzymatic browning and microbial growth in culture media. These results could be a scope for further screening of EOs to be used for the improvement of in vitro culture of A. vera and other industrial crops.