Analysis of the Constituents of Scent Secretions from Dolycoris baccarum Using GC-MS
ZHANG Yan',GUO RuiSheng',HOU JianGan2 ('DepartmentofBiological Science and Technology,Jinzhong University,Jinzhong,Shanxi3619,China;2nstitute of Applied Biology, Shanxi University, Taiyuan, Shanxi O30oo6, China)
Abstract: 【Objective】 This study aimed to analyze the constituents of scent secretions from Dolycoris baccarum and the differences between the male and female adults to provide references for development of specific control measures against the insect.【Method】 The adults of D baccarum were collcted from Jinzhong University and Shanxi Uinverisity Town area, and their scent secretions were sampled and analyzed using a solid-phase microextraction (SPME) device coupled with gas chromatography-mass spectrometry (GC-MS). 【Result】 A total of six categories of compounds, namely aldehydes,ketones, alkanes, alcohols, ester and cyclics,including 38 distinct compounds were detected in the scent secretions from both male and female adults of the insect. The components found in the females only included 14 compounds of 5 categories: hexanal belonging to aldehydes; 1,5,5-trimethyl-bicyclo[2.2.1heptan-2-ol belonging to alcohols; L-fenchone belonging to ketones; 4-methyl-decane, dodecane, 3,7-dimethyl-decane, pentadecane, 2,6-dimethyl-undecane, 4- methyl-dodecane, 2,4,6-trimethyl-decane, 4,6-dimethyl-dodecane and 2,7,10-trimethyl-dodecane belonging to alkanes; and toluene and 1-methylene- 1H. -indene belonging to cyclics. Seven compounds of five categories were obtained only in the males: fenchone belonging to ketones; 2-methyl-decane, 4,8-dimethyl-undecane, and 2,6,1l-trimethyl-dodecane belonging to alkanes; (1S,3R,5S,6R)-(-)-5- caranol belonging to alcohols; 2-ethylhexyl ester formic acid belonging to esters; and naphthalene belonging to cyclics. Seventeen compounds of 5 categories were found common in both of the male and females: heptanal and (E) -(3,3-dimethyl-cyclohexylidene)-acetaldehyde belonging to aldehydes; L-(-)-camphor belonging to ketones; n-decane, undecan and tetradecane belonging to alkanes; ethanol, 2-ethyl-l-hexanol and endo-borneol belonging to alcohols; phenol, ethylbenzene, m-xylene, benzaldehyde,3-ethylene toluene, acetylbenzene, 1,3-dichlorbenzene and 2,4-ditert-butyl phenol belonging to cyclics.【Conclusion】The 38 compounds identified enriched the information of scent secretions from D . baccarum, among which, the 14 compounds of 5 categories obtained only in females can lay the foundation for further exploitation of the sexual atractants of the insect by using pheromones; and the 7 compounds of 5 categories obtained only in males provide a reference for further specialized biological control of the bug.
Keywords:Dolycoris baccarum; scent secretions; GC-MS
0 引言
【研究意義】昆蟲信息素具有生物活性高、使用簡便、專一性強(qiáng)的優(yōu)點,已經(jīng)廣泛用于害蟲監(jiān)測和防控中(DOMINGUEZetal.,2019;崔艮中等,2023;XUetal.,2023)。從信息素角度對蝽類昆蟲尋找有效的生物防治方法越來越受到人們的重視。對斑須蝽[Dolycoris baccarum(Linnaeus)]雌雄蟲臭腺分泌物進(jìn)行比較分析有很大的生物學(xué)意義。對其后胸臭腺分泌物進(jìn)行研究,不僅可以加深對斑須蝽信息素的了解,而且可以為從信息素角度對斑須蝽進(jìn)行有效的生物防治提供基礎(chǔ)信息?!厩叭搜芯窟M(jìn)展】斑須蝽隸屬半翅目異翅亞目蝽科斑須蝽屬,是一種植食性害蟲。若蟲和成蟲均以刺吸式口器吸食寄主植物的汁液,特別喜歡嫩葉、嫩芽、嫩莖,有群聚性、弱趨光性和假死性,常危害棉、玉米、高梁、粟、大豆、煙、麻、稻、麥、蘋果、梨等經(jīng)濟(jì)作物和水果。斑須蝽成蟲在落葉、雜草和墻縫中越冬,早春僅在晴朗無風(fēng)的午間活動,其余時間潛藏在植株下部(董慈祥等,2003)。4月下旬至5月上旬成蟲開始在附近蔬菜地為害,以在藜科蔬菜上為害最嚴(yán)重;5月中旬至6月上中旬遷徙到玉米和果樹上為害,給農(nóng)業(yè)生產(chǎn)帶來嚴(yán)重經(jīng)濟(jì)損失(孫宏偉等,2016)。斑須蝽成蟲體長 ,體灰黃色或赤褐色,頭部前端至小盾片有黃色細(xì)線,前胸背板有黃色窄邊,腹部每節(jié)背面中央和兩側(cè)都有黑色斑(劉峰,2008)。成蟲產(chǎn)卵期較長,當(dāng)環(huán)境溫度在 20℃左右,相對濕度在
時,產(chǎn)卵達(dá)到高峰。蝽類昆蟲具有臭腺,當(dāng)其受驚時會從中噴射出大量具有強(qiáng)烈刺激性氣味的化學(xué)物質(zhì),這些化學(xué)物質(zhì)可以發(fā)揮防御天敵、性信息素以及告警信息素的功能(ISHIWATARI,1974;LOCKWOODandSTO-RY,1985;ALDRICH,1988;MORAESetal.,2008)。目前,對半翅目蝽科昆蟲雌雄個體臭腺分泌物化學(xué)物質(zhì)進(jìn)行組分分析的研究較多。王玉潔等(2009)對荔枝蝽[Tessaratoma papillosa(Drury)]雌雄蟲臭腺分泌物的乙醚提取物分析發(fā)現(xiàn)共有化合物
-辛烯醛對成蟲有顯著的驅(qū)避作用。張濤(2011)發(fā)現(xiàn)丁酸己酯同時存在于綠盲蝽雌雄蟲中,但在雄蟲中的含量遠(yuǎn)高于雌蟲,雄蟲在交配過程中通過釋放丁酸己酯可以抑制其他雄蟲的靠近,ZHANG和ALDRICH(2003)還發(fā)現(xiàn)這種現(xiàn)象存在于松植盲蝽(Phytocorisdifficilis)中。羅靜(2017)首次從分子生物學(xué)角度對半翅目中黑盲蝽信息素生物合成相關(guān)基因進(jìn)行探索,鑒定出818條可能參與信息素生物合成的候選基因。馬春燕等(2013)通過對斑須蝽雌雄蟲不同個體間酯酶同工酶的酶譜分析發(fā)現(xiàn)雌蟲比雄蟲在陽極端多1條帶,性別差異明顯,且大于個體差異。張嫣等(2024)對斯氏珀蝽雌雄成蟲后胸臭腺分泌物中的差異組分進(jìn)行了比較分析?!颈狙芯壳腥朦c】目前,對斑須蝽的防治仍以化學(xué)防治為主,由此帶來的耐受藥劑種類不斷增多,抗藥性現(xiàn)象也日漸突出(張麗陽和劉承蘭,2016)。對斑須蝽成蟲臭腺分泌物進(jìn)行測定分析以用于該蟲的監(jiān)測和防治,具有重要的前景和意義,但目前對斑須蝽成蟲臭腺分泌物的組分還不清楚?!緮M解決的關(guān)鍵問題】本研究擬采用固相微萃取裝置偶聯(lián)氣相色譜一質(zhì)譜聯(lián)用儀進(jìn)行氣味測定的方法(ZHANGandPAWL-ISZYN,1993;王大偉等,2018;張嫣等,2020),分析探討斑須蝽雌雄成蟲臭腺分泌物的化學(xué)組分情況及相關(guān)組分在不同性別間的差異,以期為開展該蟲的專一性綠色防治提供參考。
1.1 供試?yán)ハx
本試驗所用斑須蝽野生成蟲捕捉于晉中學(xué)院校園及山西大學(xué)城區(qū)域,對捕捉到的成蟲活體立即進(jìn)行后胸臭腺分泌物收集,隨后在實驗室完成組分測定分析。
1.2 試驗儀器與設(shè)備
試驗所用儀器與設(shè)備及其制造商見表1。
1.3斑須蝽臭腺分泌物收集與分析
1.3.1 臭腺分泌物的收集
將采集到的活力強(qiáng)的供試?yán)ハx活體標(biāo)本,迅速放入制作好的氣體收集瓶中,做震動、搖晃處理。保證蟲體處于生命體征活躍的狀態(tài),使其較充分地釋放臭腺分泌物。采集分泌物后,將蟲體取出,保存在 75% 的乙醇溶液中,并與分泌物氣體樣品一對一做好標(biāo)記。在光學(xué)顯微鏡下對昆蟲標(biāo)本進(jìn)行分類鑒定,將所采集的分泌物樣品保存在 冰箱中,直至分析進(jìn)樣(張嫣等,2020)。
1.3.2 萃取針活化
將SPME探針插入GC進(jìn)樣口250℃活化 5min (首次使用要在GC進(jìn)樣口250℃作老化處理60min)。本研究采用(ZZ-SPME-06)探針,其萃取材質(zhì)為活性炭/聚二甲基硅氧烷/聚二乙烯基苯(ACAR/PDMS/DVB)復(fù)合涂層,為雙極性萃取層,適合較寬范圍的揮發(fā)性有機(jī)物,有利于對未知組分的分析(KRAJICEKetal.,2016)。
1.3.3臭腺分泌物富集
設(shè)置空白對照試驗,設(shè)置固相微萃取裝置溫度為40℃,待溫度穩(wěn)定后將活化后的固相微萃取針管逐個插入樣品瓶中操作探針萃取頭伸出,保證樣品瓶密封,萃取 到達(dá)平衡相(張嫣等,2020)。
1.3.4 氣體測定分析
氣相色譜(GC)條件:色譜柱為TG-5MS,石英毛細(xì)管柱規(guī)格為 。柱箱溫度在
下運行
,然后以
升高至250℃,并在該溫度下保持
。載氣為氣,柱流量為
。進(jìn)樣口采用分流方式,分流比為 1:10 。進(jìn)樣器和傳輸線溫度均保持在 250℃(張嫣等,2020)。
質(zhì)譜(MS)條件:離子源電子能量為 的標(biāo)準(zhǔn)EI源,傳輸線溫度250℃,質(zhì)譜數(shù)據(jù)采集模式為全掃描,掃描離子范圍為
amu(張嫣等,2020)。
2 結(jié)果與分析
2.1斑須蝽雌雄蟲后胸臭腺分泌物的色譜分析結(jié)果
對斑須蝽雌雄蟲后胸臭腺分泌物色譜分析,結(jié)果如圖1、圖2所示。結(jié)果發(fā)現(xiàn),同一出峰時間,斑須蝽雌蟲和雄蟲后胸臭腺分泌物色譜圖的出峰形狀是相似的,只是峰高有差異。這說明雌雄蟲臭腺分泌物的主要組分相似,但是含量有很大的差異。
2.2斑須蝽雌雄蟲后胸臭腺分泌物的初步定性結(jié)果分析
將實驗測定的雌雄蟲臭腺分泌物結(jié)果,去除背景信號,與標(biāo)準(zhǔn)譜庫(NIST2018數(shù)據(jù)庫)質(zhì)譜圖進(jìn)行比對,篩選相似性指數(shù) (即匹配度gt;80.0% )且相對含量 gt;0.01% 的化合物。對關(guān)鍵性的化合物需參照文獻(xiàn)記錄進(jìn)行二次比對,進(jìn)而完成測定結(jié)果中化合物種類的初步定性。由此,分析得到斑須蝽兩性成蟲分泌物化學(xué)組分,如表2所示。
從表2可以看出,僅在雌蟲臭腺分泌物中測得的化合物有5類14種,包括醛類的己醛,酮類的小茴香酮,烷類的4-甲基癸烷、十二烷、3,7-二甲基癸烷、十五烷、2,6-二甲基十一烷、4-甲基十二烷、2,4,6-三甲基癸烷、4,6-二甲基十二烷、2,7,10-三甲基十二烷,醇類的異酮,以及環(huán)類的甲苯、1-亞甲基-1H-節(jié)。僅在雄蟲臭腺分泌物中測得的化合物有5類7種,包括酮類的酮,烷類的2-甲基癸烷、4,8-二甲基十一烷、2,6,11-三甲基十二烷,醇類的 (1R,2S,4R,6S)-4,7,7. -三甲基-雙環(huán)[4.1.0]庚烷,酯類的2-乙基甲酸己酯,以及環(huán)類的萘。雌雄蟲臭腺分泌物中共同含有的化合物有5類17種,包括醛類的庚醛、 (E) -(3,3-二甲基環(huán)己亞基)-乙醛,酮類的左旋樟腦,烷類的癸烷、十一烷、十四烷,醇類的乙醇、2-乙基己醇、冰片,以及環(huán)類的苯酚、乙苯、間二甲苯、苯甲醛、3-乙基甲苯、苯乙酮、間二氯苯、2,4-二叔丁基苯酚。
3 討論
昆蟲信息素是物種進(jìn)行信息交流的化學(xué)語言,可以實現(xiàn)種內(nèi)或種間個體信息傳遞的調(diào)控。性信息素是由昆蟲某一個體分泌,被同種異性個體的感受器接受,并引起異性個體產(chǎn)生一定生殖行為反應(yīng)的微量化合物(吳帆等,2021,2023)。利用昆蟲性信息素監(jiān)測害蟲,具有高靈敏度、高選擇性、環(huán)境友好等優(yōu)點。利用性信息素誘殺求偶交配期的雄蟲,可以使雌蟲無法高效繁殖后代,從而起到減藥控害的效果(王亞紅等,2018;鄭麗霞等,2018)。雄蟲對雌蟲性信息素的接收是實現(xiàn)昆蟲種內(nèi)性信息通訊和完成性行為的重要環(huán)節(jié)(趙新成等,2003)。本研究獲得的僅在斑須蝽雌蟲臭腺分泌物中的化合物組分有5類14種,若從中篩選出對雄蟲具有高效引誘活性的化合物組分,可有效用于該蝽性引誘劑的研發(fā)。
有報道指出,有少數(shù)昆蟲的性信息素由雄蟲分泌引誘雌蟲(BILLETERandWOLFNER,2018;VALTEROVAetal.,2019),因此本研究獲得的僅在雄蟲臭腺分泌物中的5類7種化合物組分,有望為開發(fā)該蟲的性信息素研究奠定基礎(chǔ)。隨著昆蟲交配通訊機(jī)理研究的逐步深入,人們發(fā)現(xiàn)雄性昆蟲的性信息在種間生殖隔離中發(fā)揮重要作用,尤其對于某些具有相同雌性信息素的物種來說更是必不可少(李紅軍等,2008)。因此,通過對這些化合物開展進(jìn)一步的室內(nèi)生物學(xué)活性測定,可以為實現(xiàn)該蝽的專一性綠色防治提供參考(REYN-OLDSetal.,2016;RIZVIetal.,2021;雷子寅等,2023)。
4結(jié)論
本研究分析獲得的斑須蝽雌雄兩性成蟲共有的化合物有5類17種,它們可能與斑須蝽雌雄成蟲進(jìn)行相互識別有關(guān),是實現(xiàn)種群繁衍進(jìn)行信息交流的重要成分,可以發(fā)揮聚集信息素或報警信息素的功能。下一步可在實驗室開展行為反應(yīng)測定實驗或在斑須蝽為害嚴(yán)重的田間開展誘集試驗,為開發(fā)和掌握聚集信息素在種群監(jiān)測和大量誘殺方面的應(yīng)用奠定基礎(chǔ)。
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