Chemical Composition and Acetylcholinesterase Activity of the Essential Oil of Anisophyllea disticha

Published in Khimiya Prirodnykh Soedinenii, No. 2, March–April, 2021, pp. 315–316.

Anisophyllea disticha (Jack) Baill. (Anisophylleaceae) is locally known as 'raja berangkat' in Malaysia, distributed mainly in Sumatra, Lingga Island, Peninsular Malaysia, and Borneo. It grows in lowland forest, freshwater swamp forest, and on granitic sands. In Malaysia, an infusion of the plant is used to treat diarrhea and dysentery, while a decoction of the leaves is used to treat jaundice. In addition, the roots is boiled with onion and black cumin (Nigella sativa) to relieve body aches and tiredness. In Sumatra, the decoction of the roots is used during confinement, while the astringent leaves are used to treat cuts and wounds [[1]]. The phytochemical analysis of the genus Anisophyllea led to the isolation of triterpenoids and phenolic compounds that showed antifungal, antibacterial, anti-leishmanial, and cytotoxicity activities [[2]–[4]]. The literature search did not reveal any report on the essential oil composition of the genus Anisophyllea.

The leaves of Anisophyllea disticha were collected from Gambang, Pahang in September 2019, and identified by Shamsul Khamis from UKM. A voucher specimen (SK15/19) was deposited at the Herbarium of UKM. The fresh leaves (500 g) was subjected to hydrodistillation in a Clevenger-type apparatus for 5 h. The essential oil obtained was dried over anhydrous magnesium sulfate and stored at 4–6°C. The oil yield was 0.20% based on the fresh weight.

Analysis of the chemical composition of the essential oil was performed using gas chromatography (GC) and gas chromatography-mass spectrometry (GC-MS). GC analysis was performed on Agilent Technologies 7890B and Agilent 7890B FID systems equipped with an HP-5MS capillary column (30 m × 0.25 mm × 0.25 μm film thickness). Helium was used as a carrier gas at a flow rate of 0.7 mL/min. Injector and detector temperatures were set at 250 and 280°C, respectively. The oven temperature was kept at 50°C, then gradually raised to 280°C at 5°C/min and finally held isothermally for 15 min. Diluted samples (1:100 in diethyl ether) of 1.0 μL were injected manually (split ratio 50:1). In addition, GC-MS chromatograms were recorded using Agilent Technologies 7890A and Agilent 5975 GC-MSD systems equipped with an HP-5MS capillary column. Helium was used as carrier gas at a flow rate of 1 mL/min. The injector temperature was 250°C. The oven temperature was programmed from 50°C (5 min hold) to 250°C at 10°C/min and finally held isothermally for 15 min. For GC-MS detection, an electron ionization system with ionization energy of 70 eV was used. A scan rate of 0.5 s (cycle time: 0.2 s) was applied, covering a mass range from 50–400 amu.

For identification of essential oil components, co-injection with the standards was used, together with correspondence of retention indices and mass spectra with respect to those occurring in Adams, NIST 08, and FFNSC2 libraries [[5]]. Semi-quantification of essential oil components was made by peak area normalization considering the same response factor for all volatile components.

The identified essential oil components with their percentages are listed in Table 1. Thirty components, accounting for 98.3% of the total composition, were fully characterized and grouped into two classes, sesquiterpene hydrocarbons and oxygenated sesquiterpenes. The essential oil was characterized by sesquiterpene hydrocarbons, which constituted 16 compounds, accounting for 67.5% of the total composition. Meanwhile, oxygenated sesquiterpenes were present in substantial amounts, which comprised of fourteen components, accounting for 30.8% of the total composition. The major components were germacrene D (25.3%), δ-cadinene (15.2%), caryophyllene oxide (7.0%), and spathulenol (6.4%).

Table 1. Chemical Composition of the Essential Oil from Anisophyllea districhta

aLinear retention index experimentally determined using homologous series of C6-C30 alkanes; brelative percentage values are means of three determinations ± SD.

Germacrene D is one of the most common plant volatiles considered to be a biogenetic precursor of many sesquiterpenes such as cadinane, muurolane, and amorphane derivatives. This metabolite is involved in plant-insect interaction acting as a pheromone on receptor neurons [[6]]. It was also shown to be an important deterrent and insecticidal agent against different parasites such as mosquitos, aphids, and ticks [[7]].

AChE inhibitory activity of the essential oil was measured by a slight modification of the previous spectrophotometric method [[8]]. Electric eel AChE was used, while acetylthiocholine iodide was employed as substrates of the reaction. DTNB acid was used for the measurement of the anticholinesterase activity. Galantamine was used as a reference. The essential oil indicated moderate AChE (I%: 62.5% ± 0.3) and inhibitory activity at 1.000 μg/mL concentration compared to galantamine, which gave 95.9% ± 0.2 inhibition. In previous reports, AChE inhibition can be explained by the high content of monoterpenes [[9]]. This study shows that the essential oils had not detected the presence of monoterpenes, hence contributing to moderate AChE inhibition.

Recent research shows that the genus Anisophyllea has been extensively studied from the medicinal perspective. The genus Anisophyllea shows food exploitation opportunities [[10]]. The past decade has seen an increasingly rapid interest in two species of the same genus, A. laurina and A. quangensis, which reveal a substantial level of oil production [[11]]. Meanwhile, A. boehmii kernels have been reported to be a rich source of good quality oil [[12]]. In conclusion, these results shed light on the phytochemistry of this unexplored species of the Flora of Malaysia.