Effect of different terroir conditions on endogenous hormones and berry quality of Chardonnay at the East Helan Mountains

【Objective】This study aimed to investigate the changes of endogenous hormone content and fruit quality of Chardonnay grape cultivated in different terroir conditions, and to clarify the relationship between the endogenous hormones and fruit quality in order to provide a theoretical basis for Chardonnay flavor formation in each sub-production region of the Eastern foothills of the Helan Mountain in Ningxia.【Methods】We used Chardonnay (Vitis vinifera L.) in three sub-production areas of ZhenBeiBu (Gravelly soil, ZBB, Yuanshi vineyard), YuQuanYing (Aeolian soils, YQY, Chateau yuquan) and GeZiShan (Sierozem, GZS, Xige estate) as experimental materials. The berries were sampled at five periods, E-L 33 (Berries still hard and green), E-L 35 (Berries begin to colour and enlarge), E-L 37 (Ber-ries not quite ripe) and E-L 38 (Berries harvest- ripe), and were frozen with liquid nitrogen. The berry skin, pulp and seed were ground to powder in liquid nitrogen and 10 mL of 80% methanol was added to 1 g of the powder. The solution was extracted overnight in the dark at 4 ℃ in a shaker. The mixture was then centrifuged at 12 000 × g and 4 ℃ for 5 min and the supernatant was collected. The extraction process was repeated three times and the collected supernatant was pooled and the methanol was evaporated off by nitrogen blowing apparatus. The remaining material was extracted by adding 1.5 times of petroleum ether, and the ether phase was discarded. The same volume of methyl for mate was added for extraction and the ester phase was collected. The ester phase obtained was blown dry by nitrogen blowing apparatus, dissolved with chromatographic methanol, passed through a 0.22 μm filter membrane and stored at - 20 ℃ . The endogenous hormone content of abscisic acid (ABA), indole acetic acid (IAA), gibberellin A3 (GA3), isopentene adenine (2- IP), and trans zeatin riboside (tZR) in fruit skin, flesh and seed during fruit development were determined by UPLC-MS (Ultra Performance Liquid Chromatography Tandem Mass Spectrometry). The contents of glucose, fructose, malic acid and tartaric acid in the fruits were determined by HPLC. The contents of individual sugar, organic acid were determined by HPLC according to a previously described method with minor modifications. The juice was contrifuged at 12 000 ×g for 10 min, The supernatant was extracted and filtered through a 0.22 µm filter and used for subsequent analysis. The separating column were Hypersil GOLDTM Amino (250 mm×4.6 mm, 5 µm) and C18 (250 mm×4.6 mm, 5 µm) column. The acetonitrile-water mixture (75∶25) and methanolKH2PO4 mixture (3∶97) were the mobile phase at flow rate of 0.5 mL·min-1 and 0.8 mL·min-1 . The column temperature of 25 ℃ and injection volume of 10 μL were adopted. The analysis was carried out with three technical replicates and the corresponding sugar and acid content were calculated using a standard curve. The 100- grain weight, titratable acid and total soluble solids content of grape berries were determined. Quantitative real-time PCR (qRT-PCR) was used to detect the expression levels genes related to abscisic acid metabolism, total RNA was used as the template for reverse transcription according to the Prime ScriptTM RT Reagent Kit with gDNA Eraser Reverse Transcription Kit instruction. The VvActin was selected as the internal reference, and the primers of the VvNCED2, VvNCED4, VvAAO3 and VvHYD2 genes were designed with Primer 5.【Results】The fruit quality indexes of the same variety cultivated in the three areas were significantly different. The glucose content of Chardonnay fruit at harvest was 84 mg·g-1 in YQY, 76 mg·g-1 in GZS, and 75 mg·g-1 in ZBB, and the fructose content was 105 mg·g-1 in YQY and 84 mg·g-1 in ZBB. The malic acid continued to decline in Chardonnay fruit development, with 9.6 mg · g-1 in the fruit of ZBB at maturity, 39% higher than that of GZS. The content of the tartaric acid did not change significantly at the later stage of fruit development, with the highest content in YQY and the lowest in GZS at maturity. During the growth of Chardonnay fruit, the ABA content in the fruit tissues of the three areas was highest during the E-L 35 period and gradually decreased as the fruit matured, and the ABA content in the fruit of the GZS was higher than that of the other areas; the GA3 content in the peel had a peak during the E-L 36 period, the GA3 content in the flesh showed an “M”change trend. The IAA content in the peel and flesh showed a decreasing trend, the IAA content in the peel in ZBB appellation was the highest and it was lower in the seeds after E-L 35, with insignificant differences among areas; the 2-IP content in the peel gradually increased in GZS and YQY and decreased at harvest; the tZR content in the flesh of Chardonnay showed an increasing trend, with the lowest tZR content in YQY. The endogenous hormone content of Chardonnay fruits in the three sub-appellations differed from each other, and the differences were significant at the E-L 35. The VvNCED expression during E-L 35 was significantly higher in the fruit of GZS than those of the other two production areas, and the VvHYD2 expression in the fruit of ZBB was significantly higher than those of GZS andYQY.【Conclusion】The sugar content and the solidity-acid ratio of Chardonnay grown in YQY aeolian soils were higher than that in other regions, indicating that Chardonnay was better adapted to YQY’s terroir. The ZBB gravel soil has the highest nitrogen content and rich organic matter, and the fruit of Chardonnay grown there had higher IAA and tZR content, lower ABA content than the fruits in other producing areas, and had lower sugar content, higher acidity content and lower ripeness than the fruits in the other two areas.