Agro-Morphological Traits
The 100-berry weight, berry shape, berry skin color, ripening time, growth habit, presence of thorns, and leaf area of 10 genotypes are shown in
Table 1. Statistically significant differences among genotypes for 100-berry weight (
p < 0.05) were determined. The genotypes exhibited 100-berry weight in a range between 13.85 g (S-4) and 27.87 g (S-8) indicating a twofold difference among S-4 and S-8 genotypes. Most of the genotypes (six genotypes) revealed oblong fruit shape, followed by elliptical (two genotypes) and oblate berry shape (two genotypes) (
Table 1).
Four genotypes had light-orange berry skin color, three genotypes had yellow berry skin color, two genotypes had orange berry skin color, and only one genotype had dark-orange berry skin color. The ripening time of the 10 genotypes occurred from 23 September (S-1) to 7 October (S-6). Regarding the growth habit, trees and shrubs were represented equally, with five genotypes each. Seven genotypes presented only few thorns per plant, while the remaining genotypes showed medium presence of thorns. Leaf area of the genotypes ranged from 2.56 cm2 (S-1) to 4.22 cm2 (S-6) with statistically significant differences (p < 0.05) among the analyzed genotypes.
The results indicate that sea buckthorn genotypes were quite variable in most of the agro-morphological characteristics. In fact, previous studies conducted in different countries also showed great diversity in most of the agro-morphological traits in sea buckthorn. For example, Sezen et al. [
38] used 30 female seed-propagated sea buckthorn genotypes to determine their landscape and horticulture value and reported quite variable 100-berry weight that ranged between 15 and 26 g. Zheng et al. [
39] used a large number of sea buckthorn cultivars belonging to ssp.
mongolica and found 100-berry weight between 37 and 74 g. Zheng et al. [
40] reported 100-berry weight between 9 and 20 g among ssp.
sinensis genotypes. In India, 100-berry weight of sea buckthorn genotypes was found to be between 11.53 g and 18.87 g [
41]. In China, it was reported to vary between 18.5 and 19.5 g among genotypes belonging to
H. rhamnoides and between 19.0 and 20.5 g among genotypes belonging to
H. salicifolia [
42]. In another study conducted in different valleys in India, diverse 100-berry weight was documented. For example, in Mana valley, they reported average 100-berry weight of 21.25 g, whereas, in Niti valley, the 100-berry weight amounted to 16.73 g [
43]. Li et al. [
16] used 78 diverse sea buckthorn accessions and reported 100-berry weight between 10.73 g (
H. rhamnoides ssp.
sinensis) and 47.69 g (
H. rhamnoides ssp.
mongolica).
Yadav et al. [
41] used a number of wild-grown sea buckthorn genotypes in India and found that fruit shape varied from round to ovate, while berry skin color ranged from greenish-yellow to yellow-orange in
H. salicifolia accessions. Sezen et al. [
38] found that the majority of
H. rhamnoides genotypes in Coruh valley in Turkey had oblong berry shape while yellow, light-yellow, dark-yellow, yellow-orange, orange, and dark-orange peel colors were present among wild-grown sea buckthorn genotypes. India is one of the richest countries in terms of variability of sea buckthorn’s gene pool, and Dhyani et al. [
43] found oval, elliptical, round oval/elliptical, and ovate berry shape with diverse berry skin color including orange yellow, orange, reddish yellow, red orange, and orange. Li et al. [
16] used 78 diverse sea buckthorn accessions and reported oblong, ovate, and elliptical berry shape.
Sezen et al. [
38] reported early or medium ripening characteristics in sea buckthorn genotypes in Turkey. Singh and Singh [
44] also found great variations for this morphological trait in native
H. salicifolia and
H. rhamnoides female plants in Himachal Pradesh. Sezen et al. [
38] also reported that most of the sea buckthorn genotypes had a bush growth habit, but the tree growth habit was also evident. This was also strongly supported in a study conducted in India, which revealed that most of the sea buckthorn genotypes had a bush growth habit [
41].
Previously, Sezen et al. [
38] found that most of the sea buckthorn genotypes had few or medium thorns, and the leaf area of these genotypes was between 1.59 and 4.26 cm
2, indicating great variability. In India, leaf area was found to be between 2.28 and 9.35 cm
2 among sea buckthorns belonging to different species [
41]. Sabir et al. [
20] also reported a quite variable number of thorns and leaf sizes among sea buckthorn genotypes grown in Pakistan. The agro-morphological characteristics varied among studies conducted on different continents. These differences could be connected to diverse origins, species, different parts of the fruit analyzed, climatic and growing conditions, etc.
Table 2 shows the results obtained from the juice analyses, including juice yield, vitamin C content, titratable acidity, SSC, and protein and lipid content of the 10 analyzed genotypes. For all researched parameters, statistically significant differences were evident at the 0.05 level.
As shown in
Table 2, genotypes S-2, S-3, S-10, S-1, and S-5 had higher values of fruit juice yield, i.e., 57.15%, 54.42%, 54.10%, 52.25%, and 50.40%, respectively. The lowest fruit juice yield was obtained from genotype S-4, which amounted to 44.87%. Sezen et al. [
38] reported quite variable berry juice yield among 30 seed-propagated female sea buckthorn genotypes grown in Coruh valley in Turkey, ranging between 37.00% and 53.60%, indicating similarity with our study. Zheng et al. [
39] used a large number of sea buckthorn cultivars belonging to ssp.
mongolica and found juice yield between 50.5% and 59.4%. Zheng et al. [
40] reported juice yield between 38.9% and 62.7% among ssp.
sinensis genotypes. Previous researches showed higher fruit juice yield (60–80%) among sea buckthorn genotypes and cultivars [
20,
41,
43,
45,
46] compared to our study. The observed differences could be the result of plant material used (different species, genotypes, accessions, etc.), growing locality, agronomic practices applied, etc.
The vitamin C content of the 10 sea buckthorn genotypes in this study was in the range of 37.45 mg/100 g (S-5) to 62.85 mg/100 g (S-8), indicating nearly twofold differences between these two genotypes. Sezen et al. [
38] found lower vitamin C content (between 19 and 34 mg per 100 g) among sea buckthorn genotypes. Yao et al. [
47] studied vitamin C concentrations of 71
Hippophae rhamnoides genotypes and found quite variable vitamin C content ranging from 28 to 201 mg/100 g. Jalakas et al. [
48] reported vitamin C content to vary between 49 and 65 mg per 100 g among sea buckthorn cultivars. The great variations in the vitamin C content of sea buckthorn genotypes are characteristic for this unique plant species. This trait could also be affected by the genotype, geographical origin, level of maturity of the berries, growing conditions, etc. [
40].
The soluble solid content (SSC) and titratable acidity content of the genotypes in this study varied from 12.56% (S-2) to 14.67% (S-4) and 3.14% (S-6) to 4.73% (S-9), respectively. Genotypes greatly differed from each other for SSC and titratable acidity content at the 0.05 level (
Table 2). Sezen et al. [
38] reported SSC and titratable acidity among sea buckthorn (
H. rhamnoides) genotypes to be 10.65–14.60% and 2.75–5.02%, respectively which is in accordance with our results. Sea buckthorn berries are considered highly acidic fruits. In Finland, the SSC content of sea buckthorn was reported to be between 7.4% and 12.6% [
29]. Kuhkheil et al. [
49] found that the content of vitamin C and SSC were the main variables in chemical constituents for the effective detection of original wild populations of sea buckthorn (
Hippophae rhamnoides L.) in central Alborz Mountains in Iran. They reported the lowest SSC of 8.60% and average SSC content between 17% and 20% in different years by using a large number of populations. Zheng et al. [
39] used a large number of sea buckthorn cultivars belonging to ssp.
mongolica and found SSC and acidity to be 7.40–9.00% and 2.90–4.99%, respectively. Zheng et al. [
40] found SSC to be between 7.3% and 21.8% among ssp.
sinensis genotypes.
The protein and lipid content of sea buckthorn genotypes is given in
Table 2. The genotypes exhibited statistically significant differences for both analyzed parameters at the 0.05 level. Protein and lipid content was 0.60% (S-4) to 0.80% (S-8) and 5.02% (S-2) to 6.17% (S-6), respectively. These results indicate that sea buckthorn berries are a rich source of proteins and lipids (
Table 2). Criste et al. [
25] analyzed four varieties of sea buckthorn (
H. rhamnoides) in Romania and found protein content ranging between 0.72% in the Carmen variety and 0.86% in the SF-6 variety. These results agree with our findings on protein content. One of the most important properties of sea buckthorn berries is its lipid content in the mesocarp section, as well as in the seeds [
49,
50,
51]. The lipid content of whole berries can vary considerably with the variety and other factors. Criste et al. [
25] investigated four varieties of sea buckthorn (
H. rhamnoides) in Romania and reported lipid content of berries (pulp) ranging between 4.61% and 5.71%. The lipid content of the mesocarp (pulp) of sea buckthorn berries is mainly determined by the used genotype, as well as by the environmental conditions. Previous studies also reflected a genotypic effect on the lipid content of fresh berries of
Hippophae spp.; the reported lipid percentage ranged from 1.4% in ssp.
sinensis from China up to 13.7% in ssp.
turkestanica from the Western Pamirs [
52]. The specific sugar content of genotypes is given in
Table 3. The genotypes contained mainly glucose (0.14–0.71%) and fructose (0.10–0.59%), while a few genotypes contained negligible sucrose content (
Table 3).
We found statistically significant differences in fructose and glucose content among the 10 sea buckthorn genotypes. Previously, Yang [
53] studied specific sugars in sea buckthorn berries and reported glucose and fructose as the main sugars in berries for all three major subspecies (
H. rhamnoides ssp.
sinensis, ssp.
rhamnoides, and ssp.
mongolica). Criste et al. [
25] reported glucose and fructose as the main specific sugars in four sea buckthorn cultivars belonging to
H. rhamnoides in Romania and reported fructose and glucose content of 0.18–1.10% and 0.17–0.46%, respectively. They also found that only one cultivar contained a negligible amount of sucrose, determining at the same time higher glucose than fructose content, similarly to our findings. Glucose content in our research was also similar to the findings of Yang [
53] in all samples of
H. rhamnoides. In another study, Yang et al. [
54] found fructose and glucose to range from 0.6% in ssp.
rhamnoides to 24.2% in berries of ssp.
sinensis. Zheng et al. [
39,
40] indicated glucose and fructose as the main sugars in sea buckthorn berries, suggesting that both sugars levels are affected by the cultivars.
In
Table 4, the total phenolic content, total anthocyanins, and antioxidant capacity in berries from the 10 sea buckthorn genotypes are shown. As indicated in
Table 4, the differences in all analyzed parameters among genotypes were statistically significant (
p < 0.05).
A high genotypic variation in terms of total phenolic content was observed (412–622 mg GAE/100 g FW). The highest total phenolic content was observed in genotype S-8 (622 mg GAE/100 g FW), followed by the S-5 genotype (587 mg GAE/100 g FW), while the lowest value was recorded in genotype S-4 (412 mg GAE/100 g FW;
Table 4).
Total phenolic content was previously reported to be quite variable among sea buckthorn cultivars and genotypes grown in different agro-climatic conditions in the world. Saeidi et al. [
55] reported 247 mg GAE/100 g FW of total phenolic content in wild-grown
H. rhamnoides ssp.
rhamnoides, indicating lower values than those recorded in our samples. However, Rop et al. [
56] and Crieste et al. [
25] found total phenolic content in four sea buckthorn cultivars grown in Czech Republic and Romania to range from 862–1417 mg GAE/100 g and 1012–1866 mg GAE/100 g FW, values higher than those recorded in our study. Bittová et al. [
57] also reported higher total phenolic content (i.e., between 1070 and 1730 mg GAE/100 g) in berries of sea buckthorn cultivars. All of the abovementioned studies revealed significant differences existing among sea buckthorn cultivars in terms of total phenolic content. Di Mauro et al. [
58] reported that the polyphenolic profile in olive is cultivar-dependent. The local sea buckthorn genotypes were characterized by markedly higher contents of total polyphenols compared to blackberry (262 mg/100 g), blueberry (300 mg/100 g), raspberry (322 mg/100 g), strawberry (323 mg/100 g), and blackcurrant (434 mg/100 g) [
59]. The World Health Organization (WHO) recommendation to increase consumption of fruit, vegetables, and fiber is a key lifestyle change that could help to reduce the risk of noncommunicable diseases (NCDs) [
60]. Although deficiencies in polyphenol intake do not result in specific deficiency diseases, adequate intake of polyphenols could confer health benefits, especially with regard to chronic diseases. Tea, cocoa, fruits, and berries, as well as vegetables, are rich in polyphenols [
61].
Genotype S-6 with dark-orange color had the highest total anthocyanin content in berries (38.7 mg/L), while yellow berry genotypes such as S-4, S-8, and S-10 genotypes had the lowest total anthocyanin content, i.e., 11.2 mg/L, 11.4 mg/L, and 9.3 mg/L, respectively (
Table 4). The differences in total anthocyanins among genotypes were found to be statistically significant at
p < 0.05 (
Table 4). Tiitinen et al. [
29] previously studied a number of sea buckthorn genotypes in Finland and reported the total anthocyanin content of sea buckthorn berries to range from 7 to 38 mg/L, demonstrating similarities with our study. Sezen et al. [
38] also found that relatively dark-orange sea buckthorn berries contain more anthocyanin than yellow and light-yellow sea buckthorn berries. Sabir et al. [
20] studied sea buckthorn genotypes in Pakistan and reported anthocyanin content ranging between 0.5 and 25 mg/L, which in accordance with our results.
We found statistically significant differences (
p < 0.05) among genotypes for antioxidant capacity by using FRAP and TEAC assays. Genotype S-8 showed the highest antioxidant capacity in both methods as 2.93 mmol Trolox equivalent/100 g in the TEAC assay and 1.48 mmol Trolox equivalent/100 g in the FRAP assay (
Table 4). In addition, the genotypes that had the highest total phenolic content also showed the highest antioxidant activity in both assays. The FRAP, TEAC, and TPC (total phenolic content) results showed a close relationship, indicating that antioxidant capacity is attributable to the wide range of polyphenols present in sea buckthorn berry skin and flesh. Chen et al. [
62] also reported that antioxidant activity in sea buckthorn berries followed the same trend as the concentrations of total phenolics. On the basis of these findings, it can be concluded that differences among the phenolic profiles and antioxidant capacities of sea buckthorn berries significantly depend on the genotype because all plants in our study were growing in similar environmental conditions, receiving similar sun exposure and temperature levels. Makovics-Zsohar et al. [
63] investigated six sea buckthorn genotypes in Hungary and revealed a nearly threefold difference between the lowest and highest antioxidant capacities of the tested genotypes. They reported TEAC values that ranged between 1.76 and 3.13 mmol Trolox equivalent/100 g fresh weight and FRAP values that ranged between 0.45 and 1.80 mmol AA equivalent/100 g. They also found that
Hippophae rhamnoides berries possess in vitro antioxidant activity, strongly determined by the genotype as well as by the harvest time. Criste et al. [
25] also reported that the antioxidant capacity of four sea buckthorn genotypes was quite variable among genotypes, and all genotypes had relatively high antioxidant capacity determined by 2.2-diphenyl-1-picrylhdrazyl (DPPH) and TEAC assays.
The fatty acid content of fruits of the 10 sea buckthorn genotypes is given in
Table 5. It is obvious that the genotype strongly influenced fatty acid content, and that there were statistically significant differences among genotypes (
p < 0.05) for individual fatty acids, except stearic acid (
Table 5).
Linoleic acid was the main fatty acid in the pulp of sea buckthorn genotypes and varied among genotypes from 24.11% to 36.37%. This fatty acid was followed by palmitoleic acid (18.13–26.44%) and palmitic acid (15.40–21.20%). The content of linolenic and stearic acid was determined to be lower than that of the abovementioned fatty acids and ranged from 3.88–7.02% and 1.80–3.23%, respectively (
Table 5). Fatty acids in sea buckthorn berries (pulp) are very important from a nutritive point of view because sea buckthorn berries are edible when ripe, and it is clear that sea buckthorn berries are a valuable source of some biologically active compounds, including antioxidants and fatty acids. Saeidi et al. [
55] found that berries of
H. rhamnoides grown in Iran include linoleic (34.2%), palmitoleic (21.37%), palmitic (17.2%), oleic (12%), linolenic (5.37%), and stearic acid (1.67%) as dominant fatty acids. It is well documented that, among fruits, macadamia and sea buckthorn are high in concentration of palmitoleic acid [
64]. Yang and Kallio [
52] also found that sea buckthorn fruit (mesocarp or pulp) exhibits a high content of palmitoleic acid. Regarding fatty acid diversity and content, our results are in accordance with the abovementioned studies. The differences between our results and other studies could be explained by genotype, cultivar used, growing and geographical conditions, and environmental factors [
65].
Major individual phenolic acids are shown in
Table 6. As can be seen, major phenolic acids in pulp of berries belonging to the 10 analyzed sea buckthorn genotypes were gallic acid (5.43–17.12 mg/100 g), followed by quercetin (2.87–11.47 mg/100 g), rutin (2.87–11.47 mg/100 g), quercitrin (2.44–6.57 mg/100 g), luteolin (0.96–5.12 mg/100 g), and kaemferol (0.44–1.29 mg/100 g). Among all sea buckthorn genotypes, significant differences were recorded in phenolic acids at the 0.05 level (
Table 6). Criste et al. [
25] reported that gallic acid was the main phenolic acid in sea buckthorn pulp belonging to four cultivars, with concentrations varying from 6.51 to 19.37 mg/100 g, thus indicating similarities with our findings. They also reported rutin and quercetin to be the major phenolic acids in sea buckthorn berries. Previously, gallic acid, rutin, and quercetin were reported as main phenolic acids in sea buckthorn berries [
64]. Bittova et al. [
57] also reported that the main compounds identified in sea buckthorn berries were gallic acid,
p-coumaric acid, ferulic acid, rutin, and quercitrin