Chemical variability of volatile concentrate from two Ipomoea L. species within a seasonal gradient

Abstract In the present study, are extracted volatile concentrate from Ipomoea asarifolia Poir. and Ipomoea setifera (Desr.) Roem. & Schult. in five-month seasonal gradient. The flowers were subjected to simultaneous distillation − extraction (SDE). The chemical composition of the volatile concentrate was determined by gas chromatography (CG/MS) and (CG-FID). Principal Component Analysis (PCA) and Hierarchical Clustering Analysis (HCA) were performed with the chemical constituents. It was observed that the chemical composition of I. asarifolia varied more with seasonality in relation to the species I. setifera. Furthermore, there is a possibility that germacrene D and α-copaene, the main components of the variation volatile of I. asarifolia and with higher concentrations in the rainy months, have ecological importance, attracting specific pollinators for the rainy season. This is the first study to report the chemical composition of the volatile compounds of I. asarifolia and I. setifera along a seasonal gradient. Graphical Abstract

The Ipomoea L. genus is the most abundant and richest of the family (Austin and Bianchini 1998;Wilkin 1999) with pantropical distribution and about 800 species (Wood et al. 2020). Ipomoea flowers are often showy and colorful, but with little or no perceptible odor to the human sense of smell (Austin and Cavalcante 1982;Staples et al. 2012).
Some ruderal species of Ipomoea with a long flowering period are frequent in Northern Brazil: Ipomoea asarifolia (Desr.) Roem. & Schult. which blooms from March to December, and Ipomoea setifera Poir., which blooms all year round (Austin and Cavalcante 1982). Both species are possibly melitophilous, pollinated by bees (Schlising 1970;Kiill and Ranga 2003). Ipomoea asarifolia has been confirmed to occur in South and Central America, tropical Africa, East Asia and India (Khaled et al. 2017;Wiersema 2019). This species of Ipomoea causes several cases of poisoning in livestock, especially during the dry season (Austin and Bianchini 1998;Costa et al. 2011;Carvalho et al. 2014;Souza da Silva J unior et al. 2021).
Ipomoea setifera is more frequent in tropical America, place of its probable origin, occurring in forest edges, roadsides, and also being considered a grass weed (Simão-Bianchini 2002;Hassler et al. 2021). The variation in the aromatic profile directly impacts the pollination dynamics and is linked to the physiological response of each species. Thus, the present study describes the influence of the transition from the dry to the rainy season on the aromatic profile of the flowers of ruderal of Ipomoea species, in an urban region within the Amazon Forest.
The relative results of the Pearson correlation (Table S2) for the classes of compounds in relation to the climatological data indicate that the strongest linear correlations were between the volatiles of I. asarifolia versus the averages of precipitation and insolation.
The alkanes from I. asarifolia showed a directly proportional correlation with the variation in insolation (r ¼ 0.936 and p-value of 0.019) and inversely proportional to the variation in precipitation (r ¼ À0.837 and p-value of 0.077). The sesquiterpenes showed an inverse behavior: correlation inversely proportional to the AWI variation (r ¼ À0.849 and p-value of 0.069) and directly proportional to the AWR variation (r ¼ 0.27 and p-value of 0.023).
The main volatile among the alkanes is n-pentadecane, the majority compound in I. asarifolia samples from August to October. The concentration of n-pentadecane decreases along the gradient as mean insolation decreases and precipitation increases. This contradicts studies carried out with other species, which indicate an increase in the production of this constituent in rainier seasons (Assad et al. 1997;Syamsudin et al. 2019). On the other hand, I. setifera showed low correlations in all analyzes performed, with the highest correlation index between its sesquiterpenes and temperature variation (r ¼ 0.313 and p-value of 0.608). These results may indicate that the aromatic profiles of the two species of Ipomoea have different responses to the change in precipitation and insolation of the seasonal gradient.
In this study, temperature and humidity parameters remained constant during the seasonal gradient, but considerable variation was observed in precipitation and insolation values, key factors to delineate the transition from summer to winter in the Amazon Region (Harris et al. 2004;Souza et al. 2009). Previous studies have shown the influence of rainfall variation on volatile production and composition (Gaspar et al. 2021), including in the volatile floral production (Vallat et al. 2005;Fernandes et al. 2017;Syamsudin et al. 2019). However, no studies were found reporting the influence of seasonality on the chemical composition of the volatile compounds of Convolvulaceae species.
The HCA and PCA multivariate analysis were applied to the chemical compounds identified in the volatile concentrate of I. setifera and I. asarifolia, collected during the seasonal gradient from August to December 2019 ( Figures S1 and S2). The HCA ( Figure S1) indicated the formation of four distinct groups: group I formed by samples ISAUG, ISSEP, ISDEC and ISOCT (52.12%); group II formed only by the ISNOV sample (similarity of 26.41%); group III formed by the IAAUG sample (25.21% similarity); and the fourth and last group, formed by the samples IASEP, IANOV, IADEC and IAOCT (similarity between them of 51.68%).
A difference was observed between the volatile concentrate of the two analyzed species and, possibly, a seasonal variation in the aromatic profile of I. asarifolia, with the sample collected in August different from the others. The PCA axis 1 eigenvalue (PC1) explained 47.2% of the data variation, while axis 2 eigenvalue (PC2) explained 23.3% of the variations. The two axes together explain 70.5% of the variation ( Figure  S2). Chemometric analyzes such as this can be used in work on natural products, especially with essential oils and volatile concentrates, to look for possible similarities between the chemical profile of samples of the same species or species of the same genus (Fattahi et al. 2016;Buriani et al. 2017;S.G. Silva et al. 2018;Ayub et al. 2021;Franco et al. 2021).
Group I correspond to all samples of I. setifera, except ISNOV. The compounds with the highest weights in the formation of this group were the oxygenated monoterpene linalol, the alkane n-pentadecane and the sesquiterpenes (E)-caryophyllene, spathulenol, (E,E)-a-farnecene, b-elemene and pogostol. The n-pentadecane was the major constituent in all samples of I. setifera, being one of the main attractive components of orchids pollinated by males of Hymenoptera (Singer 2002;Bohman et al. 2020). The compounds (E)-caryophyllene and b-elemene were described as attractive volatiles for bees, especially when the two compounds are combined (Zhang et al. 2016;Zhang 2018). The presence of these constituents in the volatile concentrate of I. setifera tends to be consistent with the melitophyte pollination syndrome of the species (C.E. . Group II corresponds to the ISNOV sample and was characterized by the high presence of the a-humulene sesquiterpene hydrocarbon. Seasonal variations can affect the production of this volatile (Facanali et al. 2020), however, no studies were found relating a-humulene variations with precipitation variation. Furthermore, group II is formed only by the November sample of I. setifera (ISNOV), isolated from the December and October samples. This implies that the volatile concentrate composition of the species probably did not vary with the seasonal gradient. Humidity and temperature data did not vary along the gradient, while precipitation and insolation data do not justify the formation of this group. Thus, there is the possibility that another environmental factor could be related to the divergence in the composition of the volatile concentrate of the November sample of I. setifera in relation to the other months.
Group III corresponded to the IAAGO sample and was characterized by the presence of n-octadecanol. This sample formed a group distant from the samples of the following months, mainly the samples from the months of November and December. According to INMET data, August was the least rainy month of the seasonal gradient (4.93 mm) and November (10.57 mm) and December (11.13 mm) were the wettest. This result indicates that, possibly, there was a variation in the volatile compounds of I. asarifolia according to the seasonal gradient. However, there are no studies indicating the relationship between seasonality and the concentration of n-octadecanol in plant volatiles. However, there is a record of seasonal variation of this compound in the odor produced by certain birds (Soini et al. 2007).
Group IV corresponded to all samples of I. asarifolia, except IAAGO. The compounds with the highest weights for the formation of this group were the sesquiterpenes bicyclogermacrene, d-elemene, d-cadinene, ß-copaene, a-copaene, germacrene D and the alkanes n-heneicosan, n-tricosane and n-tetracosane.
There are no studies indicating a direct relationship between the production of these components with variations in precipitation or insolation, key factors in the studied seasonal gradient. However, several of these volatiles are related to the attraction of pollinators, which in turn are influenced by changes in rainfall (Lawson and Rands 2019;Mukherjee et al. 2019).
Based on these results, it was observed that I. setifera and I. asarifolia have different chemical profiles. The composition of the volatile concentrate of I. asarifolia may have been influenced by the seasonal gradient, while I. setifera did not show a clear relationship between the chemical composition of its volatile compounds and the variation in rainfall.
The variation in the volatile concentrate of I. asarifolia, more specifically the increase in the concentration of germacrene D, bicyclogermacrene, d-elemene and a-copaene in the rainiest months, may be related to a strategy to attract a new pollinator guild that is more present in this period, as rainy season bees and flies.

Conclusions
In this study, variations in the composition of the volatile concentrate of both species of Ipomoea were observed. However, no relationship between the variation in the aromatic profile of I. setifera and the seasonal gradient was found. However, it was observed that the volatile concentrate of I. asarifolia possibly varied according to the seasonal gradient, with a greater divergence between the sample from the driest month (August) and the samples from the wettest months (November and December). Previous studies have shown that some chemical components identified in the floral aromas of I. setifera and I. asarifolia are related to pollination in other species. There is a possibility that germacrene D, bicyclogermacrene, d-elemene and a-copaene have an ecological role, attracting a specific pollinator guild typical of the rainy season, with these constituents having higher concentrations in the volatile concentrate of I. asarifolia in the rainy months. Further studies are needed to confirm the dynamics of the relationship between the volatile concentrate of these species, the seasonal gradient and how these factors affect the plant-pollinator relationship.