Polyphenol metabolism in differently colored cultivars of red currant (Ribes rubrum L.) through fruit ripening.

MAIN CONCLUSION: Rare red currants colors caused by low anthocyanin content in the pink and a lack of anthocyanins in the white cultivar correlated with low ANS gene expression, enzyme activity, and increased sugar/acid ratios. Changes in the contents of polyphenols, sugars, and organic acids in berries of the three differently colored Ribes rubrum L. cultivars ('Jonkheer van Tets', 'Pink Champagne', and 'Zitavia') were determined by LC-MS and HPLC at 4 sampling times during the last month of fruit ripening. The activities of the main flavonoid enzymes, chalcone synthase/chalcone isomerase (CHS/CHI), flavanone 3-hydroxylase (FHT), and dihydroflavonol 4-reductase (DFR), and the expression of anthocyanidin synthase (ANS) were additionally measured. Despite many attempts, activities of flavonol synthase and glycosyltransferase did not show reliable results, the reason of which they could not be demonstrated in this study. The pink fruited cultivar 'Pink Champagne' showed generally lower enzyme activity than the red cultivar 'Jonkheer van Tets'. The white cultivar 'Zitavia' showed very low CHS/CHI activity and ANS expression and no FHT and DFR activities were detected. The DFR of R. rubrum L. clearly preferred dihydromyricetin as substrate although no 3',4',5'-hydroxylated anthocyanins were present. The anthocyanin content of the red cultivar slightly increased during the last three weeks of ripening and reached a maximum of 890 mg kg-1 FW. Contrary to this, the pink cultivar showed low accumulation of anthocyanins; however, in the last three weeks of ripening, their content increased from 14 to 105 mg kg-1 FW. Simultaneously, the content of polyphenols slightly decreased in all 3 cultivars, while the sugar/acid ratio increased. The white cultivar had no anthocyanins, but the sugar/acid ratios were the highest. In the white and pink cultivars, reduction/lack of anthocyanins was mainly compensated by increased relative concentrations of hydroxycinnamic acids and flavonols.

Fruit Quality Characteristics and Biochemical Composition of Fully Ripe Blackberries Harvested at Different Times.

We investigated how the quality of blackberry fruit changes during the ripening period. Since it is difficult to predict picking dates for blackberries, we were interested in how the quality of fully ripe fruit changed depending on the sampling date (from 28 July to 1 September). Blackberries (at full ripeness) were sampled at six weekly intervals and the contents of sugars, vitamin C, organic acids and phenolic components were analysed by high performance liquid chromatography combined with mass spectrometry. The colour parameters, total soluble solids and weight of the fruits were also measured. The results showed that the fruits at the last sampling had a significantly lower fruit weight and higher soluble solids. 'Cacanska Bestrna' had the highest fruit weight and vitamin C content (11.43 mg/100 g). The main sugars in blackberries were fructose (13.8-33.4 g/kg FW) and glucose (13.0-33.2 g/kg FW). 'Loch Ness' and 'Navaho' had a sweeter taste since they had the highest ratio of sugars and acids (S/A approx. 5.8) and 'Smoothstem' and 'Thornfree' had the sourest taste, with a ratio of S/A 2.5. Blackberries harvested at the first two samplings had lower anthocyanin contents than fruits from later sampling times. There were no significant differences in the content of flavonols, ellagitannins, flavanols or hydroxycinnamic acids during the ripening period. The content of vitamin C in the fruits did not change among samplings, but the fruits had a higher content of organic acids at the first two or three samplings. The results may be useful for both the processing industry and growers to set quality standards for each variety and to determine the optimal harvest time.

Are Processed Bilberry Products a Good Source of Phenolics?

Bilberries are processed into different food products and beverages. Therefore, we studied how various bilberry items (puree, infusion, liqueur, and fresh and pasteurized juices) differ in secondary metabolites as compared to the fresh fruit. For this purpose, we identified phenolics with a liquid chromatography-mass spectrophotometry (LC-MS) system and compared phenolic profiles and phenolic contents in various bilberry products prepared from the same raw material. Our results show that processing significantly affected the biochemical pattern and levels of phenolics. Total analyzed phenolics decreased with processing, mostly due to anthocyanins, which had the highest share in phenolic profile of all bilberry-based products. The highest differences among products in flavanol (63.7-fold), followed by flavonol contents (26.3-fold). Fresh fruits had the highest content of total anthocyanins (5,190 mg/100 g dry weight [DW]), followed by puree (2,719 mg/100 g DW), infusion (2,469 mg/100 g DW), and liqueur (1,830 mg/100 g DW), whereas fresh and pasteurized juices had the lowest content of total anthocyanins (921 and 1,099 mg/100 g DW). PRACTICAL APPLICATION: Although we confirmed the common belief among consumers that the consumption of fresh berries is more beneficial than consuming processed products, this study showed that various bilberry-derived products can also be valuable source of phenolic compounds all year round.

Do optimally ripe blackberries contain the highest levels of metabolites?

Five blackberry cultivars were selected for the study ('Chester Thornless', 'Cacanska Bestrna', 'Loch Ness', 'Smoothstem' and 'Thornfree') and harvested at three different maturity stages (under-, optimal- and over-ripe). Optimally ripe and over-ripe blackberries contained significantly higher levels of total sugars compared to under-ripe fruit. 'Loch Ness' cultivar was characterized by 2.2-2.6-fold higher levels of total sugars than other cultivars and consequently, the highest sugar/acids ratio. 'Chester Thornless' stands out as the cultivar with the highest level of vitamin C in under-ripe (125.87mgkg(-1)) and optimally mature fruit (127.66mgkg(-1)). Maturity stage significantly affected the accumulation of phenolic compounds. The content of total anthocyanins increased for 43% at optimal maturity stage and cinnamic acid derivatives for 57% compared to under-ripe fruit. Over-ripe blackberries were distinguished by the highest content of total phenolics (1251-2115mg GAE kg(-1) FW) and greatest FRAP values (25.9-43.2mM TE kg(-1) FW).

A wild 'albino' bilberry (Vaccinium myrtillus L.) from Slovenia shows three bottlenecks in the anthocyanin pathway and significant differences in the expression of several regulatory genes compared to the common blue berry type.

Relative expressions of structural genes and a number of transcription factors of the anthocyanin pathway relevant in Vaccinium species, and related key enzyme activities were compared with the composition and content of metabolites in skins of ripe fruits of wild albino and blue bilberry (Vaccinium myrtillus) found in Slovenia. Compared to the common blue type, the albino variant had a 151-fold lower total anthocyanin and a 7-fold lower total phenolic content in their berry skin, which correlated with lower gene expression of flavonoid 3-O-glycosyltransferase (FGT; 33-fold), flavanone 3-hydroxylase (FHT; 18-fold), anthocyanidin synthase (ANS; 11-fold), chalcone synthase (CHS, 7.6-fold) and MYBPA1 transcription factor (22-fold). The expression of chalcone isomerase (CHI), dihydroflavonol 4-reductase (DFR), leucoanthocyanidin reductase (LAR), anthocyanidin reductase (ANR) and MYBC2 transcription factor was reduced only by a factor of 1.5-2 in the albino berry skins, while MYBR3 and flavonoid 3',5'-hydroxylase (F3'5'H) were increased to a similar extent. Expression of the SQUAMOSA class transcription factor TDR4, in contrast, was independent of the color type and does therefore not seem to be correlated with anthocyanin formation in this variant. At the level of enzymes, significantly lower FHT and DFR activities, but not of phenylalanine ammonia-lyase (PAL) and CHS/CHI, were observed in the fruit skins of albino bilberries. A strong increase in relative hydroxycinnamic acid derivative concentrations indicates the presence of an additional bottleneck in the general phenylpropanoid pathway at a so far unknown step between PAL and CHS.

White versus blue: Does the wild 'albino' bilberry (Vaccinium myrtillus L.) differ in fruit quality compared to the blue one?

Wild albino and blue bilberry fruit were analyzed to compare different fruit characteristics linked to the composition of primary and secondary metabolites. Compounds were identified using HPLC-MS and standard quality parameters were determined. Albino berries were significantly smaller, accumulated less water and were characterized by 23% higher dry weight. Soluble solids content and pH value were higher in albino bilberry and their surface was lighter and characterized by a yellow hue. Both bilberry forms accumulated identical individual sugars and organic acids; however, the albino form had 33% higher content of total sugars and 9% higher content of total organic acids compared to the blue type. Fifteen anthocyanins were identified in both forms, but in albino bilberries, individual anthocyanins were only detected in traces. Blue bilberry contained 1.6-fold higher levels of flavanols, 2.1-fold higher levels of flavonols, 2.5-fold higher levels of hydroxycinnamic acid derivatives and consequently, 4.6-fold higher total phenolic content.