謝 姣，鄧麗莉，明 建，姚世響，曾凱芳※

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導(dǎo)致柑橘果實(shí)油胞病的橘油揮發(fā)組分分析

謝 姣1,2，鄧麗莉1,3，明 建1,3，姚世響1,3，曾凱芳1,3※

（1. 西南大學(xué)食品科學(xué)學(xué)院，重慶 400715；2. 貴州醫(yī)科大學(xué)公共衛(wèi)生學(xué)院，貴州 550025； 3. 西南大學(xué)食品貯藏與物流研究中心，重慶 400715）

油胞病是一種主要的柑橘生理性病害，其典型特征是病斑區(qū)油胞凸起、油胞周?chē)M織壞死并形成綠色、黃色甚至褐色病斑，這種癥狀不僅引起果皮外觀品質(zhì)下降，還嚴(yán)重影響了柑橘果實(shí)的商業(yè)價(jià)值。目前研究認(rèn)為油胞病發(fā)病與橘油泄露有關(guān)。因此，該研究分別采用市售橘油、提取橘油、橘油非揮發(fā)性成分及26種揮發(fā)性成分單品分別處理錦橙、臍橙和椪柑果實(shí)，結(jié)果發(fā)現(xiàn)市售橘油和提取橘油處理油胞病發(fā)病率均為100.00%；橘油非揮發(fā)性成分及單品-蒎烯、石竹烯、朱欒倍半萜、-金合歡烯和-金合歡烯處理其發(fā)病率均為0；而其余21種揮發(fā)性成分單品處理均出現(xiàn)油胞病癥狀，且發(fā)病率≥73.33%。研究橘油成分對(duì)柑橘果實(shí)油胞病的影響將為進(jìn)一步探索油胞病的防治措施提供理論基礎(chǔ)。

果實(shí)；有機(jī)揮發(fā)性成分；病害；錦橙；臍橙；椪柑；油胞??；橘油

0 引 言

柑橘因其外觀、風(fēng)味以及營(yíng)養(yǎng)價(jià)值而深受消費(fèi)者的青睞，是世界上最受歡迎的亞熱帶水果之一[1-3]。然而由于柑橘果實(shí)油胞病產(chǎn)生的果皮外觀缺陷不僅大大降低柑橘果實(shí)外觀品質(zhì)，還對(duì)采后貯藏造成影響，往往引起嚴(yán)重的經(jīng)濟(jì)損失[4-6]。柑橘油胞病，又稱油斑病，是一種常見(jiàn)的柑橘果皮生理性病害，常見(jiàn)于果實(shí)成熟期以及采后貯藏期[7]。目前大多數(shù)柑橘品種均發(fā)現(xiàn)有油胞病的產(chǎn)生，且不同品種柑橘對(duì)油胞病的敏感性不同[8]。油胞病典型發(fā)病癥狀為發(fā)病區(qū)整體果皮下陷、油胞周?chē)M織細(xì)胞壞死以及發(fā)病區(qū)油胞凸起甚至出現(xiàn)油胞破裂現(xiàn)象，而發(fā)病區(qū)色澤呈現(xiàn)綠色、黃色甚至褐色[9-11]。

據(jù)報(bào)道，柑橘果實(shí)在發(fā)育、成熟以及貯藏期產(chǎn)生油胞病的主要原因?yàn)橄x(chóng)害、機(jī)械損傷以及氣候變化[6]。其中，對(duì)模擬機(jī)械損傷而引起油胞病的研究表明其引起的油胞病產(chǎn)生的原因可能是機(jī)械損傷引起位于果皮黃皮層中的油胞破裂而釋放出橘油所致[12]。另外，劉麗丹等[13-15]的研究證明了使用市售橘油能誘導(dǎo)柑橘果皮油胞病的產(chǎn)生。此外，利用-檸檬烯處理同樣能誘導(dǎo)油胞病的產(chǎn)生[16]。以上研究結(jié)果表明油胞病的產(chǎn)生與橘油存在一定的關(guān)聯(lián)性[17]。

目前關(guān)于柑橘果實(shí)油胞病主要致力于果實(shí)油胞病發(fā)生后果皮組織的變化機(jī)理研究[12-14,16]以及果皮色素變化的研究[4,18]，而具體哪些橘油成分能引起柑橘果實(shí)油胞病的發(fā)生的研究鮮有報(bào)道。柑橘精油主要由85%~99%的揮發(fā)性成分和1%~15%的非揮發(fā)性成分組成[19-21]。其中柑橘精油中揮發(fā)性成分主要是單萜（包括-側(cè)柏烯、-蒎烯、檜烯、-月桂烯以及-檸檬烯）、倍半萜烯（包括順式--羅勒烯、甘香烯、-石竹烯和-金合歡烯）、醛類(lèi)（包括辛醛、香茅醛、橙花醛和香葉醛）、醇類(lèi)（包括松油醇、芳樟醇、香茅醇和香葉醇）、酯類(lèi)（包括乙酸香茅酯、乙酸橙花酯和乙酸香葉酯）、氧化物類(lèi)（包括石竹烯氧化物、順式-檸檬烯氧化物和反式-檸檬烯氧化物）以及酮類(lèi)（包括諾卡酮、長(zhǎng)葉薄荷酮和香芹酮）等氧化衍生物的混合物[22-26]，而精油中非揮發(fā)性成分主要由碳?xì)浠衔?、固醇?lèi)、脂肪酸、類(lèi)胡蘿卜素、香豆素、補(bǔ)骨脂素和黃酮類(lèi)等組成[20-21]。

因此，本研究以錦橙、臍橙和椪柑為研究對(duì)象，分別利用市售橘油和提取橘油（分別從錦橙、臍橙和椪柑果皮中提取）處理以上3個(gè)品種柑橘果實(shí)來(lái)確定油胞病的產(chǎn)生與橘油的關(guān)聯(lián)性。另外，采用橘油非揮發(fā)性成分分別處理3個(gè)品種柑橘果實(shí)，從而明確橘油中對(duì)柑橘油胞病起誘導(dǎo)作用的成分。此外，前期試驗(yàn)利用固相微萃取結(jié)合GC-MS分析市售橘油和提取橘油發(fā)現(xiàn)，2類(lèi)橘油中揮發(fā)性成分同樣是由萜烯烴類(lèi)（包括單萜和倍半萜烯類(lèi)）、醛類(lèi)、醇類(lèi)、酯類(lèi)、氧化物類(lèi)以及酮類(lèi)組成，因此，Xie等[5]通過(guò)研究錦橙、臍橙和椪柑健康果皮和油胞病果皮中的揮發(fā)性成分，確定了3個(gè)品種柑橘油胞病果皮中發(fā)生主要變化的是26種揮發(fā)性成分即10種萜烯烴類(lèi)化合物即-蒎烯、-石竹烯、朱欒倍半萜、-金合歡烯、-金合歡烯、-檸檬烯、月桂烯、檜烯、-萜品油烯和-萜品油烯，5種萜醇類(lèi)化合物芳樟醇、-松油醇、-松油醇、香葉醇和香茅醇，6種醛類(lèi)化合物壬醛、癸醛、十一醛、十二醛、香茅醛和香葉醛，2種氧化物類(lèi)化合物順-檸檬烯氧化物和反-檸檬烯氧化物，2種酯類(lèi)乙酸橙花酯和乙酸香葉酯以及酮類(lèi)化合物香芹酮。因而，本研究最后利用這26種揮發(fā)性成分分別處理3個(gè)品種柑橘果實(shí)，從而明確橘油中具體揮發(fā)性成分對(duì)3個(gè)品種柑橘果實(shí)油胞病的影響，這為進(jìn)一步研究油胞病的發(fā)病機(jī)制及控制措施提供理論指導(dǎo)。

1 材料與方法

1.1 材料與試劑

試驗(yàn)柑橘品種為花期220 d左右的錦橙（Osbeck cv.#447#），臍橙（Osbeck cv. Fengji）和椪柑（Blanco cv. Ponkan），采自重慶市北碚區(qū)歇馬鎮(zhèn)柑橘果園，挑選無(wú)物理?yè)p傷、無(wú)病蟲(chóng)害、大小均勻一致的健康果實(shí)以及油胞病果實(shí)，室溫下洗凈，晾干，備用。

柑橘精油（純度為100%，由擠壓法獲得），英國(guó)植優(yōu)雅香精油公司；無(wú)水硫酸鈉，分析純，成都市科龍化工試劑廠。標(biāo)品：-檸檬烯、-萜品油烯、-萜品油烯、-石竹烯、壬醛、癸醛、香葉醛、十二醛、十一醛、芳樟醇、-松油醇、-松油醇、香茅醇、香葉醇、橙花醇乙酸酯、乙酸香葉酯和香芹酮購(gòu)自中國(guó)上海TCI公司；-蒎烯、香檜烯、-月桂烯、-金合歡烯、-金合歡烯、瓦倫西亞橘烯、香茅醛購(gòu)自美國(guó)sigma公司；順-檸檬烯氧化物，反-檸檬烯氧化物購(gòu)自日本W(wǎng)AKO公司。以上標(biāo)品除月桂烯（溶于甲基叔丁基醚中保持穩(wěn)定）和乙酸香葉酯外其他標(biāo)品純度均大于90%。

1.2 方 法

1.2.1 冷壓法柑橘果皮橘油收集

參照Sawamura等[27-28]的方法并加以改進(jìn)。分別將3個(gè)品種柑橘切成8等份，去除果肉部分，仔細(xì)去除果皮中白皮層，保留黃皮層。使用裝有飽和氯化鈉溶液的離心管收集利用槍頭壓出的油滴5 mL，整個(gè)收集工作在冰上操作完成；收集液于4 ℃、4 000×離心15 min后，再用無(wú)水硫酸鈉于5 ℃條件下干燥以上離心后的上層油狀液24 h，過(guò)濾，所得3個(gè)品種柑橘橘油于-20 ℃保存，待用。

1.2.2 市售橘油處理

參照Knight等[16,29-30]方法并加以改進(jìn)。處理組采用直接涂抹方法，將15L純度為100%的橘油直接涂抹至果實(shí)赤道周?chē)s3 cm2面積，晾干；對(duì)照果實(shí)未用橘油處理；將晾干的處理組和對(duì)照組果實(shí)用厚度為0.015 mm的聚乙烯袋單果包裝，黑暗處理，并于25 ℃、相對(duì)濕度為85%條件下貯藏。樣品每組10個(gè)果實(shí)，每組設(shè)3個(gè)平行，試驗(yàn)重復(fù)2次。

1.2.3 提取橘油處理

將從1.2.1中分別制取的錦橙、臍橙和椪柑果皮中的精油分別涂抹至相應(yīng)品種柑橘果皮上，其后參照1.2.2處理方式處理。樣品每組10個(gè)果實(shí)，每組設(shè)3個(gè)平行，試驗(yàn)重復(fù)2次。

1.2.4 橘油非揮發(fā)性性成分處理

將從1.2.1制取的3個(gè)品種柑橘橘油5 mL置于10 mL離心管中，于室溫條件下敞口放置（黑暗處理）以使橘油中揮發(fā)性成分揮發(fā)完全，周期約20 d；分別用雙重蒸餾水和乙醇溶液溶解揮發(fā)后的橘油干物質(zhì)（體積溶解比例為1∶1），過(guò)濾，參照1.2.2處理方法處理對(duì)應(yīng)的柑橘品種，對(duì)照組只使用相應(yīng)的溶劑（雙重蒸餾水和乙醇溶液）進(jìn)行處理。樣品每組10個(gè)果實(shí)，每組設(shè)3個(gè)平行，試驗(yàn)重復(fù)2次。

1.2.5 橘油揮發(fā)性成分處理

利用1.1所列橘油揮發(fā)性成分的標(biāo)品處理柑橘果實(shí)，處理方法參照1.2.2。樣品每組10個(gè)果實(shí)，每組設(shè)3個(gè)平行，試驗(yàn)重復(fù)2次。

1.2.6 發(fā)病率測(cè)定

以上柑橘果實(shí)處理后72 h進(jìn)行觀察，柑橘果實(shí)上出現(xiàn)油胞病癥狀認(rèn)定為發(fā)病，其油胞病發(fā)病率參參照Garcia-Martin等[31]的方法，其計(jì)算公式如式（1）所示。

1.3 數(shù)據(jù)分析

采用IBM SPSS Statistics 22.0軟件對(duì)實(shí)驗(yàn)數(shù)據(jù)進(jìn)行分析，用-test方法進(jìn)行差異顯著性分析；采用GraphPad Prism 7以及Photoshop 6.0進(jìn)行圖片的繪制。

2 結(jié)果與分析

2.1 市售橘油和提取橘油處理對(duì)柑橘果實(shí)油胞病發(fā)病率及發(fā)病癥狀的影響

如表1所示，市售橘油處理后的錦橙、臍橙和椪柑分別于25 ℃、相對(duì)濕度85%條件下貯藏72 h后，3個(gè)品種的柑橘果實(shí)油胞病發(fā)病率均為100.00%，且相對(duì)對(duì)照組在相同的貯藏條件下的油胞病發(fā)病率具有顯著性差異（<0.05）。另外，利用1.2.1冷壓法制取的錦橙、臍橙和椪柑果皮橘油即提取橘油分別處理對(duì)應(yīng)柑橘果實(shí)并貯藏72 h后，發(fā)現(xiàn)3個(gè)品種柑橘果實(shí)油胞病發(fā)病率均為100.00%，且相對(duì)對(duì)照組具有顯著性差異（<0.05）。

如圖1所示，市售橘油分別處理錦橙、臍橙和椪柑果實(shí)后，3個(gè)品種柑橘果實(shí)產(chǎn)生的油胞病具有相似的發(fā)病癥狀，即發(fā)病區(qū)整體塌陷，油胞凸起，油胞周?chē)M織下陷，發(fā)病區(qū)顏色呈現(xiàn)褐色。另外，觀察到3個(gè)品種柑橘油胞病發(fā)病面積與處理面積即3 cm2基本相同。以上結(jié)果揭示了市售橘油處理其果皮出現(xiàn)了油胞病的典型癥狀，結(jié)合市售橘油處理3個(gè)品種柑橘果實(shí)統(tǒng)計(jì)的油胞病發(fā)病率均為100.00%，說(shuō)明市售橘油處理能誘導(dǎo)3個(gè)品種柑橘油胞病的產(chǎn)生。

表1 市售橘油和提取橘油處理對(duì)柑橘果實(shí)油胞病發(fā)病率的影響（平均值±標(biāo)準(zhǔn)差）

注：數(shù)字的統(tǒng)計(jì)學(xué)差異（<0.05）用不同的字母表示，下同。

Note: Statistics differences (<0.05) of number are indicated by different letters, the same as below.

圖1 市售橘油處理對(duì)柑橘果實(shí)油胞病發(fā)病癥狀的影響

如圖2所示，分別利用1.2.1中制取的3個(gè)品種柑橘橘油處理對(duì)應(yīng)柑橘品種并貯藏72 h, 結(jié)果發(fā)現(xiàn)其誘導(dǎo)3個(gè)品種柑橘果皮油胞病的發(fā)病癥狀與市售橘油處理引起的油胞病有相似的癥狀即發(fā)病區(qū)整體塌陷，油胞凸起，油胞周?chē)M織下陷，發(fā)病區(qū)顏色為淺褐色。另外，提取橘油處理3個(gè)品種柑橘同樣觀察到其發(fā)病面積與處理面積相同。以上結(jié)果揭示了提取橘油處理其果皮同樣出現(xiàn)了油胞病的典型癥狀，結(jié)合提取橘油處理其果實(shí)油胞病發(fā)病率，說(shuō)明提取橘油能誘導(dǎo)3個(gè)品種柑橘油胞病的產(chǎn)生。據(jù)報(bào)道，油胞病的形成是位于柑橘果皮有色層中的油胞所含橘油泄露從而造成果皮的光毒性損傷而引起的果皮生理性失調(diào)[32-33]。這種果皮損傷主要表現(xiàn)為發(fā)病區(qū)整體組織下陷，油胞相對(duì)油胞間組織凸起，油胞周?chē)M織出現(xiàn)壞死，形成綠色、黃色甚至褐色病斑?；陂儆偷男孤兑鹩桶〉漠a(chǎn)生這一結(jié)論[32-33]以及劉麗丹等利用市售橘油誘導(dǎo)臍橙油胞病的產(chǎn)生[13-15]，說(shuō)明橘油本身與油胞病可能存在一定的關(guān)聯(lián)性[17]。目前，對(duì)于橘油誘導(dǎo)油胞病主要以臍橙為研究對(duì)象并以合成橘油研究其誘導(dǎo)作用，但并不明確市售橘油對(duì)其他柑橘品種以及柑橘所制取的橘油（提取橘油）對(duì)相應(yīng)柑橘品種果實(shí)油胞病的誘導(dǎo)作用。而結(jié)合以上市售橘油及提取橘油誘導(dǎo)3個(gè)品種柑橘果實(shí)出現(xiàn)油胞病的癥狀符合以上油胞病典型癥狀，且其發(fā)病率均為100.00%，這可進(jìn)一步說(shuō)明橘油與油胞病的形成存在一定的關(guān)聯(lián)性。

圖2 提取橘油處理對(duì)柑橘果實(shí)油胞病發(fā)病癥狀的影響

2.2 橘油非揮發(fā)性物質(zhì)處理對(duì)柑橘果實(shí)油胞病發(fā)病率及發(fā)病癥狀的影響

據(jù)報(bào)道，冷壓法收集的橘油由非揮發(fā)性成分和揮發(fā)性成分組成，而橘油中這2類(lèi)成分對(duì)柑橘果實(shí)油胞病的誘導(dǎo)作用未有報(bào)道。此外，橘油非揮發(fā)性成分有脂溶性和水溶性2類(lèi)化合物。因此，本試驗(yàn)分別利用蒸餾水和乙醇溶解非揮發(fā)性物質(zhì)處理3個(gè)品種柑橘來(lái)確非揮發(fā)性成分對(duì)油胞病的誘導(dǎo)作用。從圖3可知，3個(gè)品種柑橘果皮橘油中的非揮發(fā)性物質(zhì)分別使用蒸餾水和乙醇溶解，利用其溶解物分別處理3個(gè)品種柑橘果實(shí)，結(jié)果發(fā)現(xiàn)2種溶解物處理未出現(xiàn)油胞病典型癥狀，且其發(fā)病率均為0并相對(duì)對(duì)照組無(wú)顯著性差異（>0.05）。以上結(jié)果闡明了非揮發(fā)性成分的溶解物（脂溶性和水溶性溶解物）對(duì)3個(gè)品種的柑橘果實(shí)油胞病均不具誘導(dǎo)作用，這進(jìn)一步說(shuō)明橘油中對(duì)3個(gè)品種柑橘具有誘導(dǎo)作用的是橘油中的揮發(fā)性成分而不是其非揮發(fā)性成分。

2.3 橘油揮發(fā)性成分處理對(duì)柑橘果實(shí)油胞病發(fā)病率及發(fā)病癥狀的影響

由于橘油非揮發(fā)性成分對(duì)柑橘?zèng)]有誘導(dǎo)作用，因此可以推測(cè)橘油中的揮發(fā)性成分是誘導(dǎo)柑橘油胞病發(fā)生的主要原因。此外，根據(jù)柑橘果皮釋放具有光毒性的橘油是導(dǎo)致油胞病的主要原因[28-29]，這一理論進(jìn)一步說(shuō)明柑橘果皮油胞病發(fā)生后其發(fā)病區(qū)揮發(fā)性成分變化顯著的物質(zhì)可能是誘導(dǎo)油胞病產(chǎn)生的原因。Xie等[5]研究油胞病和健康果皮精油中揮發(fā)性成分發(fā)現(xiàn)在油胞病果皮中的10種萜烯烴類(lèi)化合物，6種醛類(lèi)化合物，2種氧化物類(lèi)化合物，2種酯類(lèi)以及酮類(lèi)化合物含量相對(duì)健康果皮具有顯著性的差異。因此，本研究利用以上26種揮發(fā)性成分分別處理3個(gè)品種柑橘來(lái)確定具體成分對(duì)果實(shí)油胞病的誘導(dǎo)作用。從表2可知，橘油揮發(fā)性性成分中的10種萜烯烴類(lèi)化合物除-蒎烯、-石竹烯、朱欒倍半萜、-金合歡烯、-金合歡烯不能引起柑橘果實(shí)發(fā)生油胞病以外，其余5種萜烯烴類(lèi)化合物均能引起柑橘果實(shí)發(fā)生油胞病且相對(duì)對(duì)照組均有顯著性差異（<0.05），其中，-檸檬烯和月桂烯處理分別誘導(dǎo)臍橙果實(shí)的油胞病發(fā)病率為93.33%與73.33%，檜烯、-萜品油烯和-萜品油烯誘導(dǎo)3個(gè)品種柑橘果實(shí)的油胞病發(fā)病率均為100.00%。另外，橘油揮發(fā)性性成分中5種萜醇類(lèi)化合物即芳樟醇、-松油醇、-松油醇、香葉醇和香茅醇，6種醛類(lèi)化合物即壬醛、癸醛、十一醛、十二醛、香茅醛和香葉醛，2種氧化物類(lèi)化合物即順-檸檬烯氧化物和反-檸檬烯氧化物誘，2種酯類(lèi)化合物即乙酸香葉酯和乙酸橙花酯以及酮類(lèi)化合物香芹酮誘導(dǎo)3個(gè)品種柑橘果實(shí)的油胞病發(fā)病率均為100.00%且相對(duì)對(duì)照組均有顯著性差異（<0.05）。

注：圖3a和3b中從左至右的圖層分布分別為錦橙、臍橙和椪柑果實(shí)，從上至下分別為對(duì)照組和處理組。

表2 橘油揮發(fā)性成分處理對(duì)柑橘果實(shí)油胞病發(fā)病率的影響（平均值±標(biāo)準(zhǔn)差）

從圖4a、4b和4e可知，5種萜烯烴類(lèi)化合物、5種萜醇類(lèi)化合物和香芹酮引起3個(gè)品種柑橘果實(shí)發(fā)生油胞病的癥狀大致相似，即部分油胞在發(fā)病區(qū)凸起，油胞周?chē)M織向下有輕微塌陷。另外，發(fā)病區(qū)域部分油胞出現(xiàn)塌陷現(xiàn)象，這種現(xiàn)象在萜烯烴類(lèi)化合物誘導(dǎo)錦橙和椪柑果皮發(fā)生的油胞病中較為明顯，而在萜醇類(lèi)和酮類(lèi)化合物誘導(dǎo)臍橙果皮發(fā)生的油胞病中較為明顯。此外，5種萜烯烴類(lèi)化合物誘導(dǎo)錦橙和椪柑發(fā)病區(qū)域顏色為淺褐色，而臍橙發(fā)病區(qū)域有紫褐色病斑產(chǎn)生；5種萜醇類(lèi)化合物誘導(dǎo)3個(gè)品種柑橘發(fā)病區(qū)域顏色均為深褐色；香芹酮誘導(dǎo)錦橙和椪柑發(fā)病區(qū)域顏色為深褐色，而臍橙發(fā)病區(qū)域顏色為淺褐色。從圖4c可知，6種醛類(lèi)化合物引起3個(gè)品種柑橘果實(shí)發(fā)生油胞病的癥狀大致相似，即發(fā)病區(qū)油胞均出現(xiàn)塌陷現(xiàn)象，而油胞周?chē)M織向下嚴(yán)重下陷，且3個(gè)品種柑橘發(fā)病區(qū)域顏色均為深褐色。從圖4d和4e可知，2種氧化物類(lèi)化合物和2種酯類(lèi)化合物引起3個(gè)品種柑橘果實(shí)發(fā)生油胞病的癥狀大致相似，即發(fā)病區(qū)油胞凸起，油胞周?chē)M織向下有輕微下陷。另外，與以上揮發(fā)性物質(zhì)不同的是2種氧化物引起的臍橙和椪柑油胞病發(fā)病區(qū)域顏色均為紅褐色，而錦橙發(fā)病區(qū)域顏色為淺黃色；2種酯類(lèi)化合物引起的錦橙和椪柑油胞病發(fā)病區(qū)域顏色為深褐色，而臍橙發(fā)病區(qū)域顏色為淺褐色。以上結(jié)果說(shuō)明萜烯烴類(lèi)化合物中-檸檬烯、月桂烯、檜烯、-萜品油烯和-萜品油烯等處理3個(gè)品種柑橘具有油胞病的典型癥狀且發(fā)病率顯著高于對(duì)照組，揭示這5種萜烯烴類(lèi)化合物能誘導(dǎo)果實(shí)油胞病的形成，這一結(jié)果同-檸檬烯能誘導(dǎo)油胞病的產(chǎn)生的結(jié)果一致[30]，而其他5種萜烯烴類(lèi)化合物處理因不具油胞病典型癥狀說(shuō)明其不能引起柑橘果實(shí)發(fā)生油胞病。5種萜醇類(lèi)、6種醛類(lèi)、2種氧化物類(lèi)、2種酯類(lèi)和1種酮類(lèi)化合物均能引起柑橘果實(shí)油胞病的發(fā)生。綜上，橘油揮發(fā)性成分引起的柑橘果實(shí)油胞病均具有各自典型特征，3個(gè)品種柑橘果實(shí)油胞病發(fā)病癥狀略有差異，這可能是品種的敏感性不同所致。

注：圖4a、4b、4c、4d、4e和4f中從上至下圖層分布為上層為錦橙、中間層為臍橙以及下層為椪柑果實(shí)。

3 結(jié) 論

通過(guò)研究市售橘油、提取橘油以及橘油非揮發(fā)性成分對(duì)柑橘果實(shí)油胞病的誘導(dǎo)作用，明確了橘油中能誘導(dǎo)柑橘果實(shí)油胞病的成分為揮發(fā)性成分。因此，本研究利用油胞病果皮中發(fā)生主要變化的26種揮發(fā)性成分來(lái)確定橘油具體成分對(duì)柑橘果實(shí)的誘導(dǎo)作用。結(jié)果發(fā)現(xiàn)，這些成分中除-蒎烯、-石竹烯、朱欒倍半萜、-金合歡烯和-金合歡烯不能誘導(dǎo)柑橘果實(shí)油胞病，其余21種揮發(fā)性物質(zhì)均能誘導(dǎo)油胞病并呈現(xiàn)出油胞病的典型癥狀。以上研究為闡述柑橘果皮油胞病的發(fā)生機(jī)制提供了理論基礎(chǔ)，并為進(jìn)一步研究油胞病的防治措施提供理論指導(dǎo)。

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Analysis of volatile components of citrus oil causing oleocellosis in citrus fruits

Xie Jiao1,2, Deng Lili1,3, Ming Jian1,3, Yao Shixiang1,3, Zeng Kaifang1,3※

(1.400715,; 2.550025,; 3.400715,)

Citrus, consumed widely throughout the world since its appearance, flavor and nutritional value, is one of the most popular subtropical fruits in the world. However, oleocellosis that often occurs at harvest time and during postharvest storage hasa negative effect on the external quality of citrus fruits due to the appearance defects formed on the surface of the pericarp, eventually causes extensive economic damage to citrus industry. Oleocellosis, a major physiology disorder in citrus, is characterized by the appearance of outstanding oil glands of blemish area, tissue necrosis occurred in the adjacent epidermis of oil glands, and formation of green, yellow even brown spots in the disorder area. It was reported thatthe occurrence of oleocellosis is highly correlated with the release of peel oil from the glands located in citrus pericarp; oleocellosis results from various injuries such as friction from hail and the mechanical damage, and the injuries will induce the breakdown of glands to form oleocellosis development. Those studies about oleocellosis development revealed that there was a certain correlation between oleocellosis and citrus oil during the oleocellosis occurrance. At present, little information was available on which citrus oil components could cause the occurrence of oleocellosis in citrus fruits. In generally, the citrus oil obtained by the cold compression method consists of non-volatile components (including two kinds of compounds: lipo-soluble and water-soluble) and volatile components. The induction of these two components in citrus oil on oleocellosis development of citrus fruit has not been reported. In terms of that, the fruits of Jincheng, Navel orange and Ponkan mandarin fruits were used as raw material, and the commercial citrus oil and extracted citrus oils from the three citrus varieties were used to treat the three types of citrus fruit to confirm whether the oleocellosis could be induced by the two types of oils, respectively. The results showed that the oleocellosis symptoms both occurred in the three citrus types after the treatment of commercial citrus oil and extracted citrus oils, respectively, which indicated that the citrus oil could induce the oleocellosis. Additionally, non-volatile components in citrus oil which were treated with the distilled water- and ethanol-soluble also were used to induce the three citrus types, respectively. The results showed that the treatment of dissolving substances (including liposoluble and water-soluble) of non-volatile compositions in citrus oil on citrus fruits did not induce the occurrence of oleocellosis. From those results above, it could be concluded that the volatile components in the citrus oil were the main reason for oleocellosis development. Therefore, the twenty-six kinds of volatile componends that that significantly changed in oleocellosis peels compared to the healthy ones were applied to induce the three citrus types of fruits to study the oleocellosis incidence and symptoms based on this conclusion, respectively. The results showed that twenty-one volatile components contained five terpene hydrocarbons (including d-limonene, β-myrcene, sabinese, γ-terpinolene and α-terpinolene), five terpene alcohols (including linalool, β-terpineol, α-terpineol, geraniol and citronellol), six aldehydes (including nonanal, decanal, undecanal, dodecanal, citronellal and geranial), two oxides (including (E)-limonene oxide and (Z)-limonene oxide), two esters (including geranyl acetate and neryl acetate) and one ketone (including (-)-carvone) compounds could induce oleocellosis development of Jincheng, Navel orange and Ponkan fruits, however, the volatile compositions of α-pinene, β-caryophyllene, valencene, β-farnesene and α-farnesene could not incuce the oleocellosis development of the three citrus types, respectively. In summary, the volatile components in citrus oil which could induce oleocellosis development of citrus fruit were identified by studying the induction of commercial citrus oil, extracted citrus oils and non-volatile compositions in citrus oil on oleocellosis development of citrus fruit. In addition, oleocellosis development in Jincheng, Navel orange and Ponkan induced by the twenty-one volatile components of citrus oil has their own characteristics, for example, the phenomena that parts of oil glands in disease areas collapsed were more obvious in Jincheng and Ponkan oleocellosis fruits induced by terpene hydrocarbons compared to Navel orange, and the color of disease areas showed pale brown in the two types of oleocellosis fruits, however, Navel orange oleocellosis fruits showed purple brown.. However, the symptoms of oleocellosis in three varieties of citrus fruit had slightly different, which might be due to different varieties of sensitivity in citrus fruit. Studying the effect of specific components in citrus oil on the oleocellosis of citrus peels will provide a theoretical basis for explaining the mechanism of oleocellosis in citrus peel and theoretical guidance for further exploration on the prevention and control measures.

fruit; volatile organic components; disease; Jincheng; Navel orange; Ponkan mandarin; oleocellosis; citrus oil

2018-05-30

2018-09-11

國(guó)家自然科學(xué)基金面上項(xiàng)目：采后柑橘果實(shí)磷脂酶D對(duì)橘油的響應(yīng)及油胞病的調(diào)控機(jī)制（31471631）

謝 姣，博士生，從事農(nóng)產(chǎn)品加工及貯藏工程方面的研究。Email：lxj4516@126.com

曾凱芳，博士，博士生導(dǎo)師，教授，主要從事果蔬貯藏與保鮮的教學(xué)與研究工作。Email：zengkaifang@hotmail.com

10.11975/j.issn.1002-6819.2019.01.038

S609+.3; S666.2

A

1002-6819(2019)-01-0310-07

謝 姣，鄧麗莉，明 建，姚世響，曾凱芳. 導(dǎo)致柑橘果實(shí)油胞病的橘油揮發(fā)組分分析[J]. 農(nóng)業(yè)工程學(xué)報(bào)，2019，35(1)：310－316. doi：10.11975/j.issn.1002-6819.2019.01.038 http://www.tcsae.org

Xie Jiao, Deng Lili, Ming Jian, Yao Shixiang, Zeng Kaifang. Analysis of volatile components of citrus oil causing oleocellosis in citrus fruits[J]. Transactions of the Chinese Society of Agricultural Engineering (Transactions of the CSAE), 2019, 35(1): 310－316. (in Chinese with English abstract) doi：10.11975/j.issn.1002-6819.2019.01.038 http://www.tcsae.org