First report of phenolic compounds isolated from Inga stipularis DC. (fabaceae) leaves

Abstract Several species of the Inga genus are used by Amazonian indigenous communities to treat injuries, pain and inflammations, which is directly related to the presence of phenolic compounds in these species. Many studies have addressed the phytochemical relevance of this genus, but they are still few considering the large number of species. Therefore, this study aimed to investigate the chemical composition of Inga stipularis leaves in order to find compounds with potential pharmacological application and economic interest. The developed method allowed the isolation and identification of 8 compounds in the ethanol extract of I. stipularis: eucryphin, neoastilbin, astilbin, neoisoastilbin, isoastilbin, quercitrin, engeletin and isoengeletin. Astilbin stands out for having been isolated directly from the fractionation of the extract by SPE with high yield. This study was a pioneer for I. stipularis and revealed the potential of the species as an abundant source of compounds of pharmacological and economic interest. Graphical Abstract


Introduction
Several Amazonian riverine and indigenous communities report the medicinal use of species of the Inga genus in the treatment of injuries, pain, inflammation, diarrhea, rheumatism, among others, that can be directly related to the high content of phenolic compounds that ensure high antioxidant capacity (Lima et al. 2018). Although there are studies about species of the genus Inga that show their phytochemical potential, such investigations are still scarce given the large number of species belonging to the genus. Therefore, this work aimed to carry out the first phytochemical study of I. stipularis leaves, in order to find compounds with potential pharmacological application and economic interest and to contribute to the phytochemical knowledge of the species.

Results and discussion
The method developed by Solid Phase Extraction (SPE) for the fractionation of the ethanol extract of Inga stipularis leaves proved to be effective, generating fractions of low complexity that enabled the isolation of 8 compounds: eucryphin (1), neoastilbin (2), astilbin (3), neoisoastilbin (4), isoastilbin (5), quercitrin (6), engeletin (7) and isoengeletin (8) (Figure 1). Of these, compound 3 was obtained pure directly from the fraction with a yield of 15% (Table S1 and Figure S1, supplementary material), verified after analysis by HPLC and NMR. All the NMR data can be found in the supplementary material.
All these compounds have activity reported as antioxidant and anti-inflammatory, in which they are mentioned as the main responsible for the biological activities in several plants (Carneiro et al. 1993;Xu et al. 2005;Huang et al. 2011;Lu et al. 2014Lu et al. , 2015Trinh et al. 2015;Dong et al. 2017;Ginting et al. 2019;Yuan et al. 2019). Among them, astilbin (3) stands out due to the largest number of studies that indicated potential as an anti-inflammatory, antimicrobial, immunosuppressant, anticancer, analgesic and in the healing of burns and hypoglycemic effect (Sharma et al. 2020).
In Table S2, Figures S2 and S3 (supplementary material) are shown the results of antioxidant activity of I. stipularis leaves ethanolic extract and its isolated compounds. The extract showed activity of 611.78 ± 19.78 mg TEAC g À1 extract by DPPH method and 507.86 ± 5.57 mg TEAC g À1 extract by ABTS þ method. These results reveal a high antioxidant capacity of the extract, considering that it is a complex mixture with about half the activity of the trolox positive control. Such activity can be attributed to the presence of phenolic compounds in the plant, mainly the major compound astilbin which showed activity of 1.51 ± 0.08 mM TEAC by DPPH method and 1.04 ± 0.04 mM TEAC by ABTS þ method, more active than trolox positive control, demonstrating high antioxidant capacity. The presence of astilbin in large quantities and others phenolic compounds in I. stipularis leaves ethanolic extract and its high antioxidant capabilities reveals the potential of this species as a source for several pharmacological applications and adds value to the Amazon flora.

Experimental section
All the experimental procedures carried out in this study are described in the supplementary material of this article.

Conclusions
All of these are phenolic substances with high antioxidant capacity, which indicates that the leaves of the species are a source of substances with possible application in the treatment of diseases related to oxidative stress, such as astilbin, which stands out for having been isolated in large quantities directly from the fractionation of the extract. This study was a pioneer for I. stipularis and provides data that may guide more detailed studies aimed at obtaining economically interesting bioactive compounds for pharmaco-logical application, in addition to contributing to the chemical knowledge of the species.