Primula polyantha
Creators: name: Tatsuzawa, Fumi
name: Mizuno, Takayuki
name: Kikuchi, Ryo
name: Kato, Kazuhisa
name: Ota, Toru
name: Murai, Yoshinori
name: Yangzom, Rinchen
name: Iwashina, Tsukasa
Publisher: name: Zenodo
Year: 2021
Subjects: Biodiversity
Taxonomy
Plantae
Tracheophyta
Magnoliopsida
Ericales
Primulaceae
Primula
Primula polyantha

description: 3.1. Flower colors and copigmentation in P. × polyantha and P. primulina
The petals from 19 P × polyantha cultivars were classified into six groups (G1–G6) according to their colors and spectral data (Table 1). The pH values of the petal homogenates were within the ranges of 5.7–6.0 for the nine violet-blue cultivars (G1) and 5.0–5.4 for other cultivars (G2–G6) (Table 1), indicating that the pH of the petals was an important factor for the expression of the violet-blue color. Regarding the distribution of major anthocyanins among the six groups, G1 and G2 were very similar, especially the presence of 6. However, they were different compared to the other groups (Table 2). On the other hand, the distribution of the flavonols was the same among six groups except for a few minor compounds (Table 2), indicating that flavonol dose not affect the flower color.
In G1, nine cultivars were violet-blue according to the RHS Color Chart, and their hue values (b*/a*) ranged from 2.40 to 1.64. We found that their major anthocyanin 6 (39.4%–64.6%) is hirsutidin 3- O - galactoside-5- O -glucoside (Table 2, Fig. 3). It has been reported that the violet-blue color of the flowers of old P. × polyantha cultivars is expressed through the intermolecular copigmentation between hirsutidin 3,5-di- O -glucoside (2) and quercetin 3- O -glucosyl-(1 → 2)-glucosyl-(1 → 6)-glucoside (A) (Saito et al., 1990; Harborne, 1968). However, in our nine modern cultivars with violet-blue flowers, hirsutidin 3- O -galactoside-5- O -glucoside (6) was the major anthocyanin, while 2 was a minor component. Therefore, we conducted a flower color reproduction experiment using 6 and A, which were major components of the modern violet-blue flower cultivars. When 6 alone was dissolved in MacIlvaine (phosphate-citrate buffer) solution at pH 5.8, the shape of the absorption spectrum of the fresh petals could not be reproduced. On the other hand, when 6 and A were mixed in the pH 5.8 buffer solution, the spectral shape was reproduced (Fig. 6). This verified that the shape of the absorption spectrum of the modern violet-blue cultivars was due to intermolecular copigmentation between 6 and A, similar to the cases of the old cultivars.
Only one cultivar belonged to G2 and was purple according to the RHS Color Chart, and its hue value (b*/a*) was 0.53 (Table 1). Although the major anthocyanin and flavonol patterns of G2 were the same as those of G1 (Table 2), the pH of its petal homogenates was equivalent to those of the red-purple and red cultivars (G3 and G4) but lower than that of the violet-blue cultivars (G1) (Table 1). This indicates that the flower color expression of the purple cultivar was influenced by the pH of its petal (Table 1).
Three cultivars belonged to G3 and matched red-purple of the RHS Color Chart, with hue values (b*/a*) ranging from 0.09 to 0.24 (Table 1). Their major anthocyanins, 2 (15.2%–20.9%), 3 (33.0%– 34.6%), and 7 (24.0%–35.7%), were identified as cyanidin 3- O -glucoside, peonidin 3,5-di- O -glucoside, and peonidin 3- O -glucoside, respectively (Table 2, Fig. 4).
Two cultivars belonged to G4 and were red according to the RHS Color Chart, and their hue values (b*/a*) were 0.42 and 1.00, respectively (Table 1). Their major anthocyanins, 2 (14.8% and 15.9%), 5 (21.7% and 8.5%), and 7 (58.8% and 70.7%), were identified to be cyanidin 3- O -glucoside, peonidin 3- O -gentiobioside, and peonidin 3- O - glucoside, respectively (Table 2, Fig. 4). It has been reported that major anthocyanins of two cultivars of G4 produced a dusky color effect (Saito et al., 1998) due to a higher concentration of anthocyanidin 3- O -glycosides (95.3% and 85.1% in the respective cultivars), which was higher than that in the three G3 cultivars (43.2%, 48.2%, and 57.3%, respectively) (Table 2).
Three cultivars belonged to G5 and were yellow-orange and yellow according to the RHS Color Chart, and their hue values (b*/a*) ranged from 4.17 to 22.45. Anthocyanins were not found in these cultivars (Tables 1 and 2). The yellow-flowered cultivars tended to be more reddish due to high amounts of carotenoids (Table 5). G6 was formed by only one cultivar and was white according to the RHS Color Chart. Anthocyanins were not detected from G6 cultivar (Tables 1, 2 and 5).
The coloration of the yellow flowers of P. × polyantha has been reported to be due to carotenoids (Yamamizo et al., 2011), such a Dendranthema × grandiflorum and Rosa cultivars, which are the most common flowers (Park et al., 2015; Wan et al., 2019). In this study, carotenoids were detected in fresh red (G4) and yellow (G5) flowers (Table 1). Additionally, in the acetone:MeOH (1:1) extract of the dried petals, a small amount of carotenoid was detected in the red-purple (G3) and white (G6) petals, but their effect on the flower color was considered as minimal (Table 5).
We also isolated two anthocyanins (1 and 6) and two flavonols (B and D) from the flower of P. primulina and identified by MS and coHPLC. Since 1 is a major component of the blue flower of P. × polyantha and was used in the flower color reconstruction, it could be presumed that a similar mechanism is involved in that blue flower.
In summary, by studying cultivars, the violet-blue coloration of the flowers of P. × polyantha was elucidated. Seventy series with ‘Halloween Purple’ was due to the intermolecular copigmentation between 6 and A and a higher pH value than that of other cultivars (Fig. 7). Additionally, the lower anthocyanin concentration in the ‘Seventy Mid Blue’ resulted in having a lower b*/a* value than that of the ‘Seventy Blue’ (Table 1), which was also considered to cause the bluing effects of flower coloration (Tatsuzawa et al., 2012b). Moreover, it was found that the changes of flowers from red to red-purple and yellow depended on a decrease of the anthocyanin concentration (causing the bluing effect), an increase of anthocyanidin 3- O -glycoside (causing a dusky effect), and the presence of a trace amount of carotenoids (causing reddish coloration) (Fig. 7).
These results give us a clearer understanding of the molecular basis behind the coloration expressed by the flowers of different P. × polyantha cultivars. Flavonoids are known to have health-benefiting effects, therefore, our identification of these novel anthocyanins and flavonols would add to the existing database of plant flavonoids, which are important resources applied in the pharmaceutical and cosmetic industries. Our findings would also be useful for plant breeders to create new cultivars with more complex colorations for the ornamental plant market.
description: Published as part of Tatsuzawa, Fumi, Mizuno, Takayuki, Kikuchi, Ryo, Kato, Kazuhisa, Ota, Toru, Murai, Yoshinori, Yangzom, Rinchen & Iwashina, Tsukasa, 2021, Flavonoids in the flowers of Primula × polyantha Mill. and Primula primulina (Spreng.) H. Hara (Primulaceae), pp. 1-13 in Phytochemistry (112827) (112827) 189 on pages 5-8, DOI: 10.1016/j.phytochem.2021.112827, http://zenodo.org/record/8259359