New cycloartane coronalyl acetate and other terpenoids with anti-inflammatory activity from the leaves of Vietnamese Gardenia philastrei

Abstract Phytochemical study on the methanolic extract of the leaves of Vietnamese plant Gardenia philastrei Pierre ex Pit. has led to the isolation of a new cycloartane coronalyl acetate (1) together with six known ones, coronlolide methyl ester (2), sootepin D (3), coronalolide (4), coronalolic acid (5), sootepin G (6) and 23-deoxojessic acid (7). Their structures were elucidated by a combination of 2 D NMR and HR-ESI-MS spectroscopies. These compounds (1-7) were tested for their anti-inflammatory activity. The result showed that six compounds (1-6) inhibit LPS-induced nitric oxide production in RAW264.7 macrophages with their IC50 values ranging from 3.76 − 75.47 µg/mL. This is the first report on the chemical constituents and anti-inflammatory activity of the G. philastrei. Graphical Abstract


Introduction
Gardenia genus of Rubiaceae family consists of about 140 species widely distributed in tropical areas over the world. Several members of this family have high medicinal value and have been used to cure various human diseases for a long time. Extract of the fruits of G. jasminoides expressed strong anti-inflammation activity (Koo et al. 2006), neural protection activity (Zhang et al. 2017); blood glucose stimulation in diabetic type 2 (Chen et al. 2014) and hepatoprotection or anti-depression (Chen et al. 2020). G. erubescens Stapf & Hutch, an important fruit tree in Sudanian savanna of Burkina Faso was used as medicine for treating gastric diseases, sexual asthenia and female infertility (Ou edraogo et al. 2019). G. gummifera, an endemic species in India has been used in Ayurveda remedies for treating nervous disorders, diarrhea. G. gummifera resinous exudation also showed anti-bacterial activity against S.aureus, K. pneumoniae, and E.aerogenes (Tambekar et al. 2009). Because of precious application in medicine, numerous studies on phytochemical constituents of members in the Gardenia genus have been conducted. Amongst these, G. jasminoides was the first species investigated due to its proved benefits in traditional medicine in some Asia countries (Chin 1964). Various secondary metabolites have been purified from this plant such as iridoids, flavonoids, monoterpenoids, triterpenoids, steroids and organic acids (Chen et al. 2020). Potential phytochemicals applied in pharmacy were discovered such as crocins, geniposide, chlorogenic acid and jasminoside A for diabetic treatment (Wu et al. 2009), (Saravanakumar et al. 2021), crocins and crocetin for obesity through lipase inhibition (Lee et al. 2005), enriched iridoids fragment for anti-arthritis medicine (Hu et al. 2019). Since the first study on G. jasminoides, numerous bioactive compounds have been isolated from the genus (Parmar et al. 2000). However, there still has been no report on the constituents of Gardenia philastrei, a Gardenia's member mainly distributed in Vietnam and Cambodia. Herein, we report the isolation, structural elucidation and anti-inflammatory activity of a new cycloartane triterpene derivative (1) together with 6 known compounds: coronlolide methyl ester (2) (Silva et al. 1997), sootepin D (3) (Nuanyai et al. 2009), coronalolide (4) (Silva et al. 1997), coronalolic acid (5) (Silva et al. 1997), sootepin G (Youn et al. 2016) (6) and 23-deoxojessic acid (7) (Banskota et al. 2000) from the leaves of Vietnamese G. philastrei.

Results and discussion
The leaves G. philastrei were extracted in methanol and followed by various separation methods to obtain seven compounds (1-7) as described in the experimental section. The positive ion HR-ESI-MS spectrum of compound 1 showed the quasi-molecular ion [M þ Na] þ peak at m/z 535.3398, corresponding to the molecular formula of C 32 H 48 O 5 with 9 degrees of unsaturation. Analysis of its 1 H NMR spectrum revealed the presence of an aldehyde proton (9.39 ppm), an olefinic proton at 6.49 ppm (bt, J ¼ 6.5 Hz), two exo-methylene protons at 5.10 (s) and 5.14 ppm (s), three quaternary methyl groups at 0.95, 0.98, 1.75 ppm and one acetyl group (2.09 ppm). Especially, one pair of doublets at 0.51 (1H, d, J ¼ 4.5 Hz) and 0.74 (1H, d, J ¼ 4.5 Hz) in its 1 H NMR spectrum were assigned as the C-19 methylene protons of the cyclopropane ring in a cycloartane triterpene (Silva et al. 1997;Nuanyai et al. 2009;Youn et al. 2016). In addition, the 13 C NMR spectrum of compound 1 exhibited 32 carbon signals, including an aldehyde group (195.4 ppm), one carboxylic acid (177.9 ppm), one acetyl group (170.9 ppm). Then, the structure of 1 was deduced from its 2 D NMR spectra. The 1 H-1 H COSY spectrum of compound 1 showed correlations between H-24 and H-23; H-22 and H-23, H-20; H-20 and H-17, H-21 indicating the partial structure [-CH-CH(CH 3 )-CH 2 -CH 2 -CH-] in 1. Furthermore, its HMBC spectrum had long-range correlations between H-26/C-24, C-25, C-27 suggesting the presence of a conjugated aldehyde at C-26. The carboxylic acid was located at C-3 due to HMBC correlation between H-2/C-3. The relative configuration of compound 1 was determined by NOESY spectrum, whose NOE correlations between H-8 and H-18, H-19 indicate that they are all b-face. Consequently, the NMR data of 1 were similar to those of coronalolic acid (5) (Silva et al. 1997), except for the additional signals of the acetyl group which was attached at C-29 due to the HMBC correlation between H-29 and CH 3 CO-. Therefore, compound 1 was characterized as coronalyl acetate as shown in Figure 1.

Conclusions
The new cycloartane triterpenoid named coronalyl acetate (1) together with six known compounds coronlolide methyl ester (2), sootepin D (3), coronalolide (4), coronalolic acid (5), sootepin G (6) and 23-Deoxojessic acid (7) were isolated and structural elucidated from the leaves of Vietnamese medicinal plant Gardenia philastrei. These cycloartanes had good inhibition of NO production in RAW 264.7 cells suggesting the potential application of this plant in the treatment of inflammatory diseases.

Supplemental material
Supplementary material relating to this article is available online: detailed experimental section, MS and NMR spectra and bioassay data.

Disclosure statement
No potential conflict of interest was reported by the authors.