Lipid composition of the Amazonian ‘Mountain Sacha Inchis’ including Plukenetia carolis-vegae Bussmann, Paniagua & C.Téllez

Fatty acid profile

Plukenetia volubilis

The fatty acid composition of P. volubilis is the most well studied in the genus, and the results from the two P. volubilis accessions from Ecuador and Peru in the current study are similar to previous results. The most abundant fatty acid in the seed oil of P. volubilis from Ecuador and Peru, respectively, is α-linolenic acid (C18:3 n-3, ω-3, ALA; 51.5 ± 3.3 and 46.6 ± 1.2%), followed by linoleic acid (C18:2 n-6, ω-6, LA; 32.5 ± 3.9 and 36.5 ± 0.8%), oleic acid (C18:1, OA; 8.5 ± 1,2 and 8.3 ± 0,4%) and smaller amounts (< 5%) of palmitic (C16:0), stearic (C18:0), eicosanoic (C20:0), and eicosenoic acids (C20:1; Fig. 2). Earlier studies have found approx. 35–51% ALA, 33–41% LA, and 8–11% OA in the seed oil of P. volubilis^{5,6,7,8,9,10,11,12}. The absolute values vary between studies, but comparisons of different accessions or cultivars of P. volubilis within single studies also demonstrate a large amount of variability⁵. The seed oil of the Ecuadorian accession in the current study contains slightly more ALA and slightly less LA than previously observed in P. volubilis, however, as we observed small morphological differences between the accessions from Peru and Ecuador, we hypothesise that this may be attributed to genetic differences, although growing conditions including elevation and temperature may also have had an effect^30,31.

Regardless of the compositional variation, the amount of ALA in the seed oil of P. volubilis is high, and only a few vegetable oils are comparable. Linseed (Linum usitatissimum L.) and chia (Salvia hispanica L.) are known for their high ALA content, and according to Ciftci et al.³², linseed oil contains 58.2 ± 0.64% ALA, 15.3 ± 1.01% LA, and 18.1 ± 0.45% OA, while chia oil contains 59.76 ± 0.13% ALA, 20.37 ± 0.19% LA, and 10.53 ± 0.17% OA. In comparison, the common cooking oils olive (Olea europaea L.) and sunflower (Helianthus annuus L.) contain < 1% ALA, while rapeseed oil (Brassica napus L.) contains approx. 10% ALA^33,34,35.

Plukenetia huayllabambana

Plukenetia huayllabambana is one of the more recently described species of Plukenetia² and have very large seeds; estimated as 6627 mm³ per seed compared with the approx. 997 mm³ of P. volubilis⁴. According to a recently published revised classification of the Plukenetia genus, P. huayllabambana is a putative hybrid between P. volubilis and the newly described P. sylvestris, a large-seeded species of the high elevation species complex sister to P. volubilis³. Our analysis shows that seed oil from P. huayllabambana has a significantly higher content of ALA (56.6 ± 0.2%) than P. volubilis from both Ecuador and Peru (51.5 ± 3.3 and 46.6 ± 1.2%, respectively), while the content of LA is also significantly lower (26.8 ± 0.1%, Fig. 2). This content corresponds well with the values previously reported for P. huayllabambana, which range from 51.3 to 58.2% ALA and from 25.8 to 29.3% LA; an ALA content generally exceeding that of P. volubilis^12,15,17.

Plukenetia carolis-vegae

The oil composition of P. carolis-vegae was analysed for the first time in the current study. Plukenetia carolis-vegae has been hypothesised to be a cultivated and fully or semi-domesticated species derived from wild, naturally occurring populations of P. sylvestris. Further, P. carolis-vegae is a part of the high elevation ‘Mountain Sacha Inchi’ species complex sister to P. volubilis also consisting of P. huayllabambana and P. sylvestris³. The oil composition of P. carolis-vegae is of particular interest since the seeds are the largest in the species complex; approx. 7069 mm³ per seed⁴.

The seed oil of P. carolis-vegae was found to contain 57.4 ± 0.0% ALA (Fig. 2), which was the highest measured value in the study and was significantly different from the values of ALA measured in the seed oil of both P. volubilis cultivars and the hybrid P. volubilis × P. carolis-vegae. It was also higher than the value measured for P. huayllabambana, albeit not significantly. Conversely, the value of LA was the lowest measured, 25.2 ± 0.0% of the seed oil, which was significantly lower than the values measured in the oil of the P. volubilis cultivars and the hybrid, and also lower, though not significantly, than the value measured for P. huayllabambana seed oil. The very high ALA content of P. carolis-vegae may be caused by genetic factors or may be related to altitude and temperature; the material of P. carolis-vegae was collected at the highest altitude of the ‘Mountain Sacha Inchis’ in this study (1610 m), and Cai et al. (2012) observed that the ALA content of P. volubilis generally increased with higher altitude and decreasing temperatures³⁰. However, studies of the oil composition of P. carolis-vegae cultivated at lower altitudes need to be conducted to assess whether the high ALA content is due to environment, genetics or both.

The amount of OA in P. carolis-vegae seed oil was 10.5 ± 0.0%, which was significantly higher than in both P. volubilis cultivars, P. huayllabambana, and the hybrid P. volubilis × P. carolis-vegae. The content of eicosenoic and palmitic acid in P. carolis-vegae seed oil (0.6 ± 0.0 and 4.9 ± 0.0%, respectively) was found to be mostly similar to the content in the seed oil of the other analysed species. In contrast, the content of eicosanoic and stearic acid (0.3 ± 0.0 and 1.0 ± 0.1%, respectively) was significantly lower than in both the P. volubilis cultivars and P. huayllabambana but similar to the levels in the hybrid. Overall, the most striking difference between the oils was the very high ALA content of P. carolis-vegae seed oil, a property which may be promising for further cultivation and domestication of the species.

Plukenetia volubilis × Plukenetia carolis-vegae

The ALA content of the P. volubilis × P. carolis-vegae hybrid seed oil is 46.8%, which is similar to that of the two P. volubilis cultivars, but lower than that of P. huayllabambana and P. carolis-vegae. Conversely, the LA content of the oil is 39.9%, which is significantly higher than all other samples except P. volubilis from Peru (Fig. 2). The OA lipid fraction is 7.1%, which is similar to that of the two P. volubilis cultivars and P. huayllabambana, but significantly different from that of P. carolis-vegae. Since the P. volubilis × P. carolis-vegae hybrid is a cross between P. volubilis and P. carolis-vegae, it is to be expected that fatty acid composition of the oil will be in between the composition of these two. While the values are not significantly different from those of P. volubilis from Peru, they are not very similar to the values of P. carolis-vegae, although the fruit morphology of the hybrid shows similarity with that of P. carolis-vegae. This dissimilarity may be due to either the genetic composition of this specific cross or may be an effect of environmental conditions. The hybrid was cultivated in a nursery at a lower altitude than the collection altitudes of either of the parents, and if altitude and temperature is indeed an essential driver of conversion from ALA to LA³⁰, this might have influenced the seed oil composition of the hybrid.

Triacylglycerol (TAG) profile

The distribution of fatty acids in the TAG molecules varies between species and cultivars and is responsible for the chemical, physical, and biological properties of oils and fats³⁶. In P. volubilis from Ecuador, P. volubilis from Peru, and P. huayllabambana, 15 different TAGs were identified, containing six different fatty acids (Table 1).

The most abundant TAG in the P. volubilis cultivars from Ecuador and Peru, and in P. huayllabambana was LLnLn, constituting 35.0 ± 2.2, 28.8 ± 2.5, and 35.6 ± 0.9%, respectively. However, a comparably very high amount of LnLnLn was found in P. huayllabambana; 23.5 ± 0. 8%. Following LLnLn, the predominant components in the two P. volubilis cultivars from Ecuador and Peru were LnLL (21.5 ± 0.3 and 22.1 ± 1.8%, respectively) and LnLnLn (13.5 ± 1.3 and 12.5 ± 2.3%, respectively), while in P. huayllabambana they were LnLnLn (23.5 ± 0.8%) and LnLL (14.1 ± 0.8%). In all samples, TAGs composed of polyunsaturated Ln (ALA) and L (LA) constituted more than two-thirds of the total TAG molecules (72.8% in P, volubilis from Ecuador, 67.2 in P. volubilis from Peru, and 74.1% in P. huayllabambana). Moreover, most of the identified TAGs (88. 5–95.9%) contained at least one residue of ALA.

These results correspond well with the TAG composition in P. volubilis oil measured by Fanali et al.³⁷, who identified LLnLn as the most abundant TAG, and found that > 80% of TAGs contained ALA. The predominant TAGs after LLnLn in P. volubilis were LnLL and LnLnLn³⁷. Similarly, Chasquibol et al.¹⁵ found LLnLn to be the most prevalent TAG in both P. volubilis and P. huayllabambana.

Genetic control of the fatty acid composition

Across all the examined species and cultivars, the SFA content is relatively low, ranging from 5.7% in P. carolis-vegae × P. volubilis to 8.4% in P. huayllabambana. Similarly, the MUFA content ranges from 7.6% in P. carolis-vegae × P. volubilis to 11.1% in P. carolis-vegae (Fig. 3). The remainder of the fatty acids is PUFA, comprised of ALA and LA, in total 84% of the seed oil in P. volubilis from Ecuador, 83.1% in P. volubilis from Peru, 83.4% in P. huayllabambana, 82.6% in P. carolis-vegae, and 86.7% in P. carolis-vegae × P. volubilis. Comparatively, linseed and chia oil contain approx. 74 and 80% PUFA, respectively³². The total PUFA content observed in the current study is largely similar across the species and cultivars. However, the relative fractions of ALA and LA vary considerably (Fig. 3), with P. carolis-vegae seed oil containing the highest amount of ALA (57.4%) and the P. carolis-vegae × P. volubilis hybrid containing the lowest (46.8%) although it has the highest total amount of PUFA. The differences in the composition of the PUFA fraction might be a result of genetic differences between the species and cultivars in combination with environmental factors.

The common pathway of PUFA biosynthesis in plants is initiated in the plastid with the formation of acyl-chains by the fatty acid synthase (FAS) complex, generating C16:0 and C18:0 fatty acids. Desaturation ensues by the action of a stearoyl-acyl carrier protein desaturase (SAD) to form OA (C18:1), which is further desaturated to LA (C18:2) by fatty acid desaturase-2 (FAD2) in the endoplasmic reticulum, and the third double bond is introduced at the ω-3 position of LA by fatty acid desaturase-3 (FAD3), also in the endoplasmic reticulum^38,39. Until recently, the molecular mechanisms underlying the synthesis of the very high PUFA content in P. volubilis had not been elucidated, although a few studies had been published^31,40,41. However, in a study by Yang et al.³⁹ two FAD genes named PvFAD2 and PvFAD3 were isolated from P. volubilis and demonstrated to catalyse the synthesis of LA and ALA, respectively, although the authors point out that their results do not fully explain the massive accumulation of PUFA in P. volubilis seeds. Nevertheless, the differences observed between the species and cultivars included in the current study may at least in part be caused by differences in the expression of PvFAD2 and PvFAD3. If so, P. carolis-vegae may have a relatively high expression of PvFAD3, leading to the synthesis of a very high amount of ALA in the seed oil, while, conversely, P. volubilis from Peru may have a lower expression of PvFAD3, yielding a lower amount of ALA in the seed oil. Furthermore, it can be speculated that the similar amounts of PUFA, but varying compositions of the PUFA fraction, observed in almost all the species and cultivars of the study could be an effect of LA being used as a substrate for PvFAD3 to produce ALA, reducing the amount of LA while increasing the amount of ALA.

Intriguingly, P. carolis-vegae × P. volubilis has the highest PUFA fraction of all the studied species and cultivars of Plukenetia, although the ALA fraction is comparably small. This characteristic could be caused by a higher expression of genes early in the biosynthetic pathway of ALA synthesis, e.g. SAD, in combination with a lower expression of PvFAD3 compared to the other samples. However, it is also possible that a lower growing temperature would have induced activity of PvFAD3 and led to a higher accumulation of ALA relatively to LA; Yang et al.³⁹ found that the activity of PvFAD3 was sensitive to temperature when expressed in yeast (Saccharomyces cerevisiae) cells, with low temperature (20 °C) significantly increasing biosynthesis of ALA. Accordingly, an analysis of the seed oil of all the ‘Mountain Sacha Inchis’, including the new species, P. sylvestris, and P. volubilis, grown at 20 and 30 °C might be useful in better understanding the very high accumulation of PUFA, primarily ALA in the large-seeded species of Plukenetia.

Source: Ecology - nature.com