Chemical Constituents and Pharmacological Activity of Agarwood and Aquilaria Plants

Agarwood, a highly precious non-timber fragrant wood of Aquilaria spp. (Thymelaeaceae), has been widely used in traditional medicine, religious rites, and cultural activities. Due to the inflated demanding and depleted natural resources, the yields of agarwood collected from the wild are shrinking, and the price is constantly rising, which restricts agarwood scientific research and wide application. With the sustainable planting and management of agarwood applied, and especially the artificial-inducing methods being used in China and Southeast Asian countries, agarwood yields are increasing, and the price is becoming more reasonable. Under this condition, illuminating the scientific nature of traditional agarwood application and developing new products and drugs from agarwood have become vitally important. Recently, the phytochemical investigations have achieved fruitful results, and more than 300 compounds have been isolated, including numerous new compounds that might be the characteristic constituents with physiological action. However, no one has focused on the new compounds and presented a summary until now. Alongside phytochemical advances, bioactivity screening and pharmacological investigation have also made a certain progress. Therefore, this review discussed the new compounds isolated after 2010, and summarized the pharmacological progress on agarwood and Aquilaria plants.


Flavonoids
Flavonoids consist of a large group of polyphenolic compounds with a benzo-γ-pyrone structure, which is ubiquitously present in plants; there is no exception for the genus Aquilaria    Figure 10. Structures of flavonoids from Aquilaria malaccensis.

Neural Activity
Agarwood has been traditionally used as a medicine for tranquilizing and reducing excitement in China, Southeast Asia, and the Middle East for centuries. Modern pharmacological studies have demonstrated that agarwood has an active effect on the nervous system [48,49]. Okugawa et al. [50] determined that a benzene extract of A. malaccensis agarwood reduced spontaneous motility, prolonged hexobarbiturate-induced sleeping time, and decreased rectal temperature, whereas petroleum ether, chloroform, or water extracts did not have that effect. A further bio-guided isolation of a benzene extract found that jinkoh-eremol and agarospirol were the main active constituents [51,52]. Takemoto et al. [53] reported that agarwood essential oil sedated mice through vapor inhalation, in which the main volatile constituents were benzylacetone, α-gurjunene, and (+)-calarene. As benzylacetone had a sedative effect, a number of derivatives were synthesized and assessed for a sedative effect. The results demonstrated that benzylacetone-like compounds had sedative activities, and their intensities varied depending on the functional group in the carbon chain, the substituent in the benzene ring, and their combinations [54]. Our recent studies showed that both the ethanol extract and essential oil of agarwood, induced by the whole-tree agarwood inducing technique in A. sinensis trees, had a sedative-hypnotic effect, where its potential mechanism is related to regulating the gene expression of GABA A receptors and potentiating the GABA A receptor function [55,56]. Agarofuran, derived from agarwood essential oil, was reported to have anxiolytic and anti-depression activity in mice [49]. To explore a potential drug for treating anxiety and depression, a series of agarofuran-like derivatives were synthesized and the activity screened, among which, buagafuran was an effective compound for anti-anxiety and anti-depression, with low toxicity and a high safety coefficient [49,57]. The potential mechanism might be through modulating central neurotransmitters, such as dopamine [58]. A metabolic study showed that buagafuran could be transformed to hydroxy metabolite and carbonyl one in a human liver microsome, where carbonyl metabolite was the main one [59]. Until now, phase II clinical trials are being conducted on buagafuran. Furthermore, many other activity screening results have also shown that compounds from agarwood have an effect on neural activity. Compound 7 (10 µg/mL) showed neural protective activity against both glutamate-induced and corticosterone-induced neurotoxicity in PC12 pheochromocytoma and human U251 glioma cells [16]. Compounds 118 and 119 exhibited potent anti-depressant activity in vitro by inhibiting [ 3 H]-5-HT reuptake in rat synaptosomes [37]. Compound 120 demonstrated remarkable antidepressant activity in vitro, by inhibiting norepinephrine reuptake in rat brain synaptosomes [38]. Simultaneously, seventeen new 2-(2-phenylethyl)chromones, including compounds 22, 27-29, 31-33, 68, 69, 78-80, 82-86, and eleven new terpenoids, such as 103-105 and 110-117, had acetylcholinesterase inhibitive effect [8,20,21,26,28,[32][33][34][35]. Above all, neural activity of agarwood is one of the most studied aspects with many active compounds and a promising drug candidate found, which will sustain it as a research hotspot in the future.

Gastrointestinal Regulation
Pharmacological studies showed that agarwood and the leaves of A. sinensis trees have a gastrointestinal regulating effect. Our studies demonstrated that the agarwood ethanol extract significantly improved intestinal peristalsis, enhanced gastric emptying, and inhibited gastric ulcer [60]. Li et al. reported that the ethanol extract of agarwood and A. sinensis leaves enhanced intestinal propulsion [61]. Kakina et al. reported that leaves of A. sinensis trees induced laxation via acetylcholine receptors on loperamide-induced constipation in mice [62]. The acetone extract of A. sinensis leaves had a laxative effect without causing diarrhea, in which genkwanin 5-O-β-primeveroside was the active constituent, whereas the methanol extract did not have the laxative effect [63]. The ethanol extract of A. sinensis leaves had a laxative effect without causing diarrhea in a rat model of low-fiber diet-induced constipation [64]. Mangiferin and genkwanin 5-O-primeveroside were the two major bioactive compounds [65]. Additionally, benzylacetone, an active compound from essential oil, had the effect of enhancing appetite [66,67]. Even though agarwood on alleviating abdominal discomfort has been widely used for centuries, the gastrointestinal regulating effect, especially on a specific disease, is not completely clear.

Antibacterial and Antifungal
The original use of agarwood was for anticorrosive deodorization in ancient China, as well as Southeast Asian countries. In Thailand, agarwood has been used for a long time as a traditional treatment for infectious diseases such as diarrhea and skin diseases [68]. Chen et al. [69] found that agarwood essential oil derived from A. sinensis, regardless of whether it originated from artificial or natural agarwood, had inhibitive activities towards Bacillus subtilis and Staphylococcus aureus [69]. Extracts of agarwood (A. crassna), isolated by water distillation, supercritical fluid carbon dioxide, and supercritical fluid carbon dioxide with ethanol as the co-solvent, showed antimicrobial activities against S. aureus and Candida albicans, but were not against Escherichia coli [70]. Sirilak et al. [68] found that an aqueous extract of A. crassna leaves possessed an in vitro antibacterial action against Staphylococcus epidermidis, causing bacterial cells to swell and distort, inhibiting the biofilm formation, and leading to cell wall rupture. An ethyl acetate soluble fraction of ethanol extract from A. crassna exhibited stronger antifungal (Fusarium solani) activity than ethanol extract [10]. Additionally, many other compounds had an antibacterial activity, such as compound 27, exhibiting inhibitory effect against S. aureus [8], compound 105 and 107 against both S. aureus and R. solanacearum, and compound 109 against S. aureus [33]. Even though the antibacterial/antifungal effect of agarwood is definite, the inhibited microbial species are not completely known. Therefore, antibacterial spectrum investigation of agarwood should be carried out.

Analgesic Effect
Wang et al. [74] found that chloroform extracts of agarwood prolonged the pain threshold induced by hot plate, and reduced the times of writhing reactions. Jinkoh-eremol and agarospirol may be the active compounds, and jinkoh-eremol's analgesic effect could be blocked by naloxone (a opioid antagonist), whereas agarosporol was weakly effected by naloxone [51]. At the same time, jinkoh-eremol and agarospirol could inhibit D 2 receptor binding and 5-HT 2A receptor binding [51].

Antiasthma
The antiasthma effect of agarwood has been traditionally used in China, and can be found in the latest Chinese Pharmacopoeia [13]. However, to our knowledge, only one study found that an ethanol extract of agarwood and A. sinensis leaves could inhibit asthma induced by histamine phosphate in guinea pig [75].

Anti-Diabetes
Mei et al. [82] found that the ethanol extracts of both agarwood and A. sinensis leaves alleviated diabetes induced by mesoxyalyurea in mice. The methanol extract of A. sinensis leaves possessed the fast blood glucose activity in rat and glucose uptake transportation by rat adipocytes [83]. Iriflophenone 3-C-β-glucoside decreased the fasting blood glucose levels in streptozocin-induced diabetic mice, and enhanced glucose uptake into adipocytes [84]. Compounds 146-149 isolated from agarwood had an inhibitive effect on α-glucosidase [44].

Others
A methanol extract of A. crassna leaves significantly reduced fever (rectal temperature) induced by baker's yeast at five and six hours after subcutaneous injection in rat [86]. The aqueous extract of A. malaccensis leaves was effective on Trypanosoma evansi with an IC 50 value 36.29 ± 1.32 µg/mL, whereas the ethanol extract was relatively weak (IC 50 = 128.63 ± 6.70 µg/mL) [88]. An ethyl acetate extract of A. crassna showed an anti-ischemic effect by attenuation of P38-MAPK activation [89].

Conclusions
Among the 154 new compounds identified from Aquilaria plants, 2-(2-phenylethyl)-4Hchromen-4-one derivatives and sesquiterpenes account for 57% and 35%, respectively, where most of the new compounds, accounting for 89%, were isolated from A. sinensis. Generally, agarwood originating from different Aquilaria plants share some common compounds, but still have several different compounds [14]. In addition, there are at least 19 species of Aquilaria plants producing agarwood, which means that large quantities of new compounds need to be explored in agarwood and Aquilaria plants. The chemical components of agarwood are diverse and complex, contributing to the diversity of bioactivity and pharmacology, including neural activity, gastrointestinal regulation, antibacterial, anti-inflammation, and cytotoxicity. Based on the specific disease and target, illuminating the active ingredients and compounds of agarwood should be carried out, which may not only contribute to the understanding of the scientific nature of the traditional agarwood application, but also benefit the new drug research and agarwood product development.