**5. Isoflavones**

Soybean is considered to be the most abundant natural source of isoflavones in the human and animal diet [184]. Isoflavones are the main components of flavonoids and the most common form of phytoestrogens, i.e. non-steroidal compounds with oestrogen-like biological properties [185]. They are considered nutraceuticals as it is claimed they have potential benefits in preventing the development of cardiovascular diseases and cancers, menopausal symptoms and osteoporosis, as well as antifungal and antioxidant properties [186–188]. Their role in plants is to encourage infection and nodulation by nitrogen-fixing bacteria [189] and to help in abiotic and biotic stress resistance [190]. The most studied isoflavones are genistein, daidzein and glycitein [191]. As a quantitative trait controlled by many minor genes [192], isoflavone content in soybean seed largely depends on the environment [185, 193–195], to the extent that even the smallest changes in the microclimate can cause significant changes in the isoflavone contents. For example, it is reported that lower temperatures during the seed fill period increase the isoflavone content, whereas environments which were warmer and drier resulted in lower isoflavone contents [196–199]. In 76 environments during 12 years, Carrera and Dardanelli [200] found a 91% decrease in total isoflavone content with mean temperatures during the seed development rising from 14.1 to 26.7°C. Nevertheless, Morrison et al. [201] evaluated 14 cultivars across 12 years at one location determining that high mean temperature during seed development did not result in isoflavone concentration decrease which might be due to the fact that only one location provides a somewhat narrower range of temperature variation [200]. The negative impact of water deficit on soybean isoflavone concentration was confirmed in many earlier studies [192, 198, 202, 203] as well. Furthermore, synthesis of isoflavones in soybean seed depends on the geographical origin, seed size, seed colour, maturity, disease tolerance and resistance to insects as well [185, 187, 204–206]. Nevertheless, isoflavone broad-sense heritability was estimated to range from moderate [207] to high [192], indicating that genotype effects were high enough to enable efficient improvement of isoflavone contents [194, 208]. The predominance of genotype effects over environmental was confirmed by Gutierrez-Gonzalez et al. [192], Murphy et al. [199] and Hoeck et al. [206]. Successful high-isoflavone cultivar development necessitates investigating the relationships between isoflavones and other important traits. For example, total isoflavone (TI) content was reported to be in a positive [204, 205, 209, 210] and negative [211] correlation with seed yield, but the lack of interrelationship has also been reported [199, 212], which suggests that the development of high-yield, high-isoflavone cultivars would be possible. The relationship between isoflavone content and each oil and protein content was reported to be negative [194, 199, 205, 209, 211], but no correlation between isoflavones and proteins was reported as well [212]. Conflicting reports on the relationships between total seed isoflavone content and other traits necessitate the need for studies including a wide range of genetic material grown across a range of environments to provide additional insight into the associations [199].

Because of the many abiotic and biotic factors influencing the isoflavone content, it is expected to vary in wide ranges. Wang and Murphy [213] found TI content to vary from 1176 to 3309 μg/g within a single cultivar of soybean. Gutierrez-Gonzales et al. [192] reported of extremely wide content range (12.4–317.4 mg 100−<sup>1</sup> g DM) for TI determined in the population of RILs in the USA. Murphy et al. [199] determined 160–370 mg 100−<sup>1</sup> g range for TI content in RIL population tested in 2-year, multilocation trials in Canada, while Adie et al. [194] determined 14.97–39.85 mg 100−<sup>1</sup> g content range for 10 soybean lines tested during 1-year trial on eight locations in Indonesia. In Europe, Cvejić et al. [208] found TI content in 20 F1 soybean progenies ranged from 156 to 366 mg 100−<sup>1</sup> g, while Bursać et al. [214] determined the TI content range of 211 to 524 mg 100 g−<sup>1</sup> for different seed coat-coloured offsprings derived from the single cross between commercial variety and germplasm collection genotype with black seed coat. Matoša Kočar et al. [195] investigated 22 MG 00 to II soybean genotypes from the Agricultural Institute Osijek (Croatia) during 3 years (2010–2012) and determined average TI content varied from 124.06 to 286.2 mg 100−<sup>1</sup> g, while average daidzein, glycitein and genistein contents were 49.27 mg 100−<sup>1</sup> g, 20.90 mg 100−<sup>1</sup> g and 94.41 mg 100−1 g DM, respectively. Although breeding of isoflavone content at the Agricultural Institute Osijek is at its beginnings, some shifts in the right direction have been made which can be seen from the higher average isoflavone contents in elite breeding lines than in standard cultivars of the same MG (**Table 4**). In Matoša Kočar et al. [195] study, genistein was the most abundant isoflavone component, followed by daidzein and glycitein. The same order of abundance was determined in the research by Lozovaya et al. [198] for two French and three US cultivars, whereas Sumardi et al. [215] researched 34 black soybeans in Indonesia and found that daidzein content was higher in 31 genotypes. In the research done by Cvejić et al. [208], Bursać et al. [214] and Tepavčević et al. [216], total daidzein was the highest followed by total genistein and total glycitein. Gutierrez-Gonzalez et al. [192] found that glycitein was the most abundant isoflavone, followed by genistein and daidzein. Although all three isoflavones are considered to have health benefits, the order of abundance is important because genistein is reported to have approximately 10 times higher biological activity compared to daidzein and glycitein [217], so genotypes with high genistein content should be favoured in selection processes aiming to create soybean genotypes suitable for food and dietetic supplement industries.

from the cross between high-isoflavone and low-isoflavone cultivars of which 6, 7, 10 and 11 QTLs were associated with daidzein, glycitein, genistein and TI, respectively. Akond et al. [193] identified three QTLs on three different linkage groups, one controlling daidzein content and two controlling glycitein content. Furthermore, QTL epistatic interactions are also thought to contribute to isoflavone variation [205]. Although many QTL have been discovered for isoflavone content, these are mainly minor effect often influenced by the environment, so discovering sufficient loci associated with isoflavone content stable in different environments

**Table 4.** The 3-year (2010–2012) isoflavone content range and mean for different maturity group elite breeding lines

and \*\* indicating significant differences at P ≤ 0.05 and P ≤ 0.01,

Improving Seed Quality of Soybean Suitable for Growing in Europe

http://dx.doi.org/10.5772/intechopen.89922

43

**MG Elite breeding lines Standard cultivars t P**

00 10.65–106.48 65.07 20.34–63.99 45.02 2.11\* P < 0.05 0 11.08–93.39 50.54 19.02–48.53 36.71 3.28\*\* P < 0.01 I 25.03–91.35 51.26 9.81–53.51 36.01 2.70\*\* P < 0.01

00 10.72–40.31 24.26 19.08–27.24 22.77 0.68ns P > 0.05 0 13.09–41.76 21.78 12.97–18.62 15.54 6.46\*\* P < 0.01 I 10.09–47.38 19.91 9.70–18.94 13.41 4.71\*\* P < 0.01

00 43.35–355.31 184.85 57.59–128.23 93.89 4.08\*\* P < 0.01 0 52.32–112.25 80.49 54.82–86.38 67.44 2.86\*\* P < 0.01 I 48.19–142.22 92.24 54.48–104.39 83.80 1.12ns P > 0.05

00 70.49–501.79 273.18 97.22–206.06 161.68 3.50\*\* P < 0.01 0 87.61–212.99 150.05 115.12–148.60 136.25 0.76ns P > 0.05 I 91.34–239.75 160.75 71.14–179.53 129.94 2.34\* P < 0.05

Range Mean Range Mean

**Daidzein (mg 100 g−<sup>1</sup>**

Glycitein (mg 100 g−<sup>1</sup>

Genistein (mg 100 g−<sup>1</sup>

Total isoflavones (mg 100 g−<sup>1</sup>

The means were tested with Student's t-test with \*

respectively, and ns indicating no significant difference.

**)**

)

)

)

created at the agricultural institute Osijek in comparison to standard cultivars.

Although genetic enhancement of soybean seed quality contributes to advances in processing industries and improves the added value properties of final soybean products, soybean as a commodity is still mostly being paid for by weight and not by composition. Considering

that are going to significantly aid in selection is yet to be accomplished [219].

**6. Conclusion**

The large influence the environmental factors have on the isoflavone content emphasises the necessity for molecular research and identification of molecular markers associated with favourable isoflavone profiles. To date, almost 90 QTLs associated with seed isoflavone content are registered in the SoyBase [97]. Akond et al. [193] identified 16 QTLs for seed isoflavones content on 12 different chromosomes (Chr) or linkage groups. Wang et al. [218] identified 33 expression QTLs on 15 soybean chromosomes, with 5 of them overlapping with phenotype QTL. In the overlapping region, 11 candidate genes underlying the accumulation of isoflavones were discovered which could be beneficial for the development of marker-assisted selection to breed soybean cultivars with high-isoflavone contents [218]. Furthermore, Wang et al. [219] identified 23 new isoflavone content QTLs and 34 QTLs in total in 130 RILs derived


The means were tested with Student's t-test with \* and \*\* indicating significant differences at P ≤ 0.05 and P ≤ 0.01, respectively, and ns indicating no significant difference.

**Table 4.** The 3-year (2010–2012) isoflavone content range and mean for different maturity group elite breeding lines created at the agricultural institute Osijek in comparison to standard cultivars.

from the cross between high-isoflavone and low-isoflavone cultivars of which 6, 7, 10 and 11 QTLs were associated with daidzein, glycitein, genistein and TI, respectively. Akond et al. [193] identified three QTLs on three different linkage groups, one controlling daidzein content and two controlling glycitein content. Furthermore, QTL epistatic interactions are also thought to contribute to isoflavone variation [205]. Although many QTL have been discovered for isoflavone content, these are mainly minor effect often influenced by the environment, so discovering sufficient loci associated with isoflavone content stable in different environments that are going to significantly aid in selection is yet to be accomplished [219].
