Essential oil extraction from fresh needles of Pinus pumila (Pall.) Regel using a solvent-free microwave-assisted methodology and an evaluation of acetylcholinesterase inhibition activity in vitro compared to that of its main components

A solvent-free microwave-assisted method and the screw extrusion of raw materials are combined and successfully applied to obtain the essential oil of fresh needles of Pinus pumila (Pall.) Regel. Single-factor screening and the Box-Behnken design method were applied to optimize the extraction of essential oils, and the yield of essential oil under the optimal conditions was 12.00 ± 0.60 mL/kg, which was highly consistent with the results predicted by the Box-Behnken design model. The process developed offers advantages in terms of the yield of essential oil and a reduced extraction time (31 min in comparison to 4 h for conventional hydrodistillation). The microstructures, chemical structures and thermal stability of needle samples before and after extraction were also investigated. The main components found in the essential oil extracted from the fresh needles of P. pumila were (+)-3-carene (22.75 %), δ-cadinene (12.12 %), terpinolene (12.11 %) and β-bisabolene (9.12 %). Furthermore, the semi-inhibition concentrations (IC50) of acetylcholinesterase inhibition activity of the essential oil, terpinolene, (−)-trans-caryophyllene and (+)-3-carene were determined to be 0.76 ± 0.04, 0.55 ± 0.03, 0.45 ± 0.02 and 0.64 ± 0.03 mg/mL, respectively. In general, P. pumila needle essential oil showed notable inhibitory ability to acetylcholinesterase, and the components terpinolene, (−)-trans-caryophyllene, and (+)-3-carene play major roles in the process of essential oils inhibiting acetylcholinesterase. Additionally, the acetylcholinesterase inhibition activities of all the tested samples were mostly related to the corresponding concentrations. The acetylcholinesterase inhibition activity of kojic acid was also determined in parallel experiments as a positive control.