A metabolomics approach to elucidate apple fruit responses to static and dynamic controlled atmosphere storage
代謝組學方法闡明蘋果果實對靜態和動態受控氣氛存儲的反應
Stefano Brizzolaraa,*, Claudio Santuccib, Leonardo Tenoric, Maarten Hertogd,
Bart Nicolaid,e, Stefan Stürzf, Angelo Zanellaf, Pietro Tonuttia
a Istituto di Scienze della Vita, Scuola Superiore Sant’Anna, Pisa, Italy
b CERM, University of Firenze, Firenze, Italy
c Fondazione FiorGen ONLUS, Firenze, Italy
d Division of Mechatronics, Biostatistics and Sensors (MeBioS), Department of Biosystems (BIOSYST), KU Leuven, Leuven, Belgium
e Flanders Centre of Postharvest Technology (VCBT), Leuven, Belgium
f Laimburg Research Centre for Agriculture and Forestry, Ora, Bolzano, Italy
ABSTRACT
The response of apple fruit to storage conditions based on low oxygen protocols depends on their genetic background. In order to elucidate common and divergent processes characterizing the metabolic changes under hypoxia, fruit of two apple (Malus domestica) varieties (‘Granny Smith’, GS, and ‘Red Delicious’, RD) were stored under two different low oxygen protocols (Ultra Low Oxygen, ULO, at 0.9 kPa oxygen, and Dynamic Controlled Atmosphere based on chlorophyll fluorescence, DCA-CF, between 0.2 and 0.55 kPa oxygen) for up to 200 and 214 days of storage for GS and RD samples, respectively. Through an integrated metabolomics approach (1H NMR, GC–MS, HS-SPME-GC–MS analyses) a total of 130 metabolites (volatiles and non-volatiles) were identified. Most of them (117) were common to both c*rs; 95 were significantly different between both c*rs when comparing the whole set of data (ULO + DCA-CF), whereas 13 volatile compounds, identified via HS-SPME-GC–MS, were specific for either GS or RD. M*riate analyses (PCA and PLS) of the whole dataset allowed to clearly discriminate between GS and RD samples. When storage condition was used as a categorical response variable, a lower percentage explained variance was obtained as this effect was overshadowed by the large effect of storage time. After 4 months of storage, RD underwent more pronounced metabolic compositional changes of the cortex, possibly associated with the evolution of ripening. Based on the accumulation pattern of pyruvate- derived metabolites (ethanol, acetaldehyde, lactate, alanine) it can be hypothesized that there are two main metabolic reconfiguration strategies in GS and RD to regenerate NAD+ and cope with energy crisis under hypoxia. GS showed more pronounced responses through changes in the nitrogen metabolism and limited induction of the ethanol fermentation while the latter was highly induced in RD under both ULO and DCA-CF. Marked differences were detected between the VOC profiles of the two c*rs regardless storage conditions. Ethyl esters and 2-methylbutyl derivatives appeared finely modulated by the oxygen level in GS and RD apples, respectively.
蘋果果實對低氧貯藏條件的反應取決于其遺傳背景。為了闡明在缺氧條件下代謝變化的共同和不同過程，兩個蘋果品種(Granny Smith，GS和Red Delicious，RD)的果實在兩種不同的低氧條件下貯藏(超低氧，ULO，0.9Kpa氧氣，以葉綠素熒光為基礎的動態控制大氣，DCA-CF，0.2-0.55kpa氧氣)，分別對GS和RD樣品進行長達200天和214天的儲存。通過綜合代謝組學方法(1hnmr、GC-MS、HS-SPME-GC-MS分析)，共鑒定出130種代謝物(揮發物和非揮發物)。其中大多數(117個)是兩個品種共有的；在比較整個數據集(ULO+DCA-CF)時，兩個品種之間的95個顯著不同，而通過HS-SPME-GC-MS鑒定的13個揮發性化合物，對GS或RD都有特異性。整個數據集的多變量分析(PCA和PLS)可以清楚區分GS和RD樣本。當存儲條件被用作分類響應變量時，由于存儲時間的巨大影響掩蓋了這一影響，因此得到的解釋方差百分比較低。貯藏4個月后，RD的皮層代謝成分發生了更明顯的變化，可能與成熟的進化有關。根據丙酮酸代謝產物(乙醇、乙醛、乳酸、*)的積累模式，可以推測GS和RD有兩種主要的代謝重構策略來再生NAD+，應對缺氧下的能量危機。GS通過氮代謝的變化和乙醇發酵的有限誘導而表現出更明顯的響應，后者在ULO和DCA-CF兩種條件下均在RD中被高度誘導，兩個品種的VOC譜在不同貯藏條件下存在顯著差異。在GS和RD蘋果中，乙酯和2-甲基丁基衍生物分別受到氧含量的精細調節。
Keywords:Hypoxic metabolism，Ultra low oxygen (ULO)，Dynamic controlled atmosphere (DCA)，Malus domestica，Postharvest，Metabolic profiling
關鍵詞：低氧代謝、超低氧(ULO)、動態控制氣氛(DCA)、蘋果、采后、代謝譜
