As水質(zhì)毒性發(fā)光細(xì)菌及微生物燃料電池在線監(jiān)測(cè)技術(shù)的比選研究

易 雯, 嚴(yán)惠華, 林秀榕, 張 琳, 韓士松, 王仕琦

廣東省環(huán)境監(jiān)測(cè)中心，廣東 廣州 510308

As水質(zhì)毒性發(fā)光細(xì)菌及微生物燃料電池在線監(jiān)測(cè)技術(shù)的比選研究

易 雯, 嚴(yán)惠華, 林秀榕, 張 琳, 韓士松, 王仕琦

廣東省環(huán)境監(jiān)測(cè)中心，廣東 廣州 510308

針對(duì)實(shí)際應(yīng)用需要，對(duì)比研究了發(fā)光細(xì)菌和微生物燃料電池兩種不同毒性預(yù)警技術(shù)對(duì)典型類金屬砷離子的響應(yīng)差異，并考察了豐水期、枯水期河水水質(zhì)背景干擾對(duì)響應(yīng)信號(hào)的影響。結(jié)果表明，發(fā)光細(xì)菌法對(duì)砷離子響應(yīng)較為靈敏，微生物燃料電池法對(duì)砷離子的響應(yīng)非常靈敏。發(fā)光細(xì)菌法受水體水質(zhì)背景干擾大，且不同生產(chǎn)廠家的儀器差異顯著，推測(cè)可能是由于菌株來源不同、儀器響應(yīng)靈敏度差異所造成。微生物燃料電池法對(duì)背景干擾的抗性較強(qiáng)，穩(wěn)定性較高，河水加標(biāo)與純水加標(biāo)樣本的劑量-效應(yīng)曲線差別不大。基于上述研究結(jié)果，分析了兩種技術(shù)監(jiān)測(cè)類金屬砷的優(yōu)缺點(diǎn)并提出實(shí)際在線運(yùn)行的相關(guān)建議。

砷；發(fā)光細(xì)菌；微生物燃料電池；生物監(jiān)測(cè)

Abstract:Research of the two biomonitoring technologies were conducted for the needs of their practical applications. The comparison between their response differences to arsenic ions as well as the effects of matrix interferences of the river water in the wet and dry seasons on the responses were presented. The study found that the luminescent bacteria toxicity test is sensitive to the arsenic ions; however, still being inferior to the microbial fuel cells toxicity methods. The toxicity test based on the luminescent bacteria is easy to be interfered by water background. The online instruments from the different manufacturers showed different response signals to the toxicant at the same concentration. We speculated that the difference in the sources of bacterial strain and the sensitivities of instruments led to the different response signals. In practical, the microbial fuel cells toxicity method was found with advantages such as good anti-interference ability and high stability. Based on the abovementioned results, the advantages and disadvantages of the two technologies have been concluded and some recommendations were given for their practical on-line operation.

Keywords:arsenic;luminescent bacteria;microbial fuel cell;bio-monitoring

發(fā)光細(xì)菌法是利用一類能夠發(fā)射可見熒光，且發(fā)光量的強(qiáng)弱程度與其活體數(shù)量和代謝活性有關(guān)的細(xì)菌建立的一種生物毒性在線監(jiān)測(cè)方法。當(dāng)水體受到污染時(shí)，會(huì)導(dǎo)致發(fā)光細(xì)菌代謝活性受到抑制甚至死亡，發(fā)光強(qiáng)度減弱。污染越嚴(yán)重，發(fā)光細(xì)菌代謝活性抑制越強(qiáng)、死亡數(shù)量越多，發(fā)光強(qiáng)度也越弱，從而可以判斷水質(zhì)的綜合毒性。目前，國(guó)內(nèi)常用的3種發(fā)光細(xì)菌為明亮發(fā)光桿菌(PhotobacteriumphosphoreumT3spp)[10]、費(fèi)氏弧菌(Vibriofischeri)[11]、青?；【?Vibrioqinghaiensissp. nov , strain Q67)[12-14]。鑒于發(fā)光細(xì)菌法是一種快速、靈敏、經(jīng)濟(jì)的監(jiān)測(cè)方法，現(xiàn)已廣泛用于環(huán)境污染物綜合毒性監(jiān)測(cè)中[15-20]。

微生物燃料電池是以微生物為陽(yáng)極催化劑，將化學(xué)能轉(zhuǎn)化成電能的裝置[21-24]。其原理是微生物直接將水中有機(jī)物分解產(chǎn)生電子，代謝過程中的電子轉(zhuǎn)化成電流。當(dāng)毒性物流入時(shí)，微生物活性降低，產(chǎn)生電流減少，利用電流強(qiáng)度與污染物濃度呈線性關(guān)系而建立生物毒性在線監(jiān)測(cè)方法。該方法響應(yīng)速度較快、重復(fù)性較好，國(guó)內(nèi)外已有很多研究者用其開發(fā)水質(zhì)毒性綜合監(jiān)測(cè)的傳感器[25-30]。

本研究從實(shí)際應(yīng)用需要出發(fā)，比較研究基于發(fā)光細(xì)菌和微生物燃料電池兩種不同水質(zhì)生物毒性在線監(jiān)測(cè)技術(shù)對(duì)典型類金屬砷的響應(yīng)差異，探討水體背景干擾對(duì)響應(yīng)信號(hào)的影響，為水質(zhì)生物毒性在線監(jiān)測(cè)技術(shù)的規(guī)范化運(yùn)行提供基礎(chǔ)數(shù)據(jù)，服務(wù)于水質(zhì)安全預(yù)警技術(shù)體系和監(jiān)督檢測(cè)體系的完善和業(yè)務(wù)化運(yùn)行。

1 實(shí)驗(yàn)部分

1.1儀器與試劑

兩種國(guó)產(chǎn)發(fā)光細(xì)菌水質(zhì)毒性在線監(jiān)測(cè)儀和進(jìn)口微生物燃料電池水質(zhì)毒性在線監(jiān)測(cè)儀，儀器型號(hào)及使用情況見表1。

表1 儀器型號(hào)及使用情況

試劑和材料：含1%硝酸的砷標(biāo)準(zhǔn)品；容量瓶；移液管；洗耳球；酸缸；儀器維護(hù)耗材等。

1.2實(shí)驗(yàn)方法

取濃度為100 mg/L的As標(biāo)準(zhǔn)溶液10.00 mL于100 mL容量瓶，用純水稀釋定容濃度為10 mg/L的As標(biāo)準(zhǔn)中間使用液；吸取10 mg/L的As標(biāo)準(zhǔn)中間使用液5.00、10.00、30.00 mL各3組，吸取濃度為100 mg/L的As標(biāo)準(zhǔn)溶液5.00 mL 3組，分別用純水、豐水期河水、枯水期河水稀釋定容至1 000 mL，配制成As3+標(biāo)準(zhǔn)溶液濃度分別為0.05、0.1、0.3、0.5 mg/L的3種不同背景的測(cè)試樣品(其中，豐水期河水背景As3+濃度為0.3、0.5 mg/L的測(cè)試樣品按同樣方法分別換成0.15、0.2 mg/L)。

趙某散步時(shí)和劉某家的狗迎面相遇，劉某的狗突然躥至趙某身后咬了一口，事后，趙某經(jīng)就醫(yī)治療共花費(fèi)數(shù)千元。經(jīng)過民警調(diào)解，雙方未達(dá)成一致意見，故趙某將劉某訴至法院，要求其賠償醫(yī)療費(fèi)等各項(xiàng)費(fèi)用共計(jì)3000余元。

考察兩種水質(zhì)生物毒性在線監(jiān)測(cè)技術(shù)(3臺(tái)儀器)對(duì)砷離子的信號(hào)響應(yīng)，每種水樣測(cè)試3次，以抑制率平均值和極差表征實(shí)驗(yàn)結(jié)果。

2 結(jié)果與討論

2.1純水背景下兩種在線監(jiān)測(cè)技術(shù)對(duì)類金屬砷的毒性響應(yīng)對(duì)比

兩種水質(zhì)毒性在線監(jiān)測(cè)技術(shù)對(duì)砷離子毒性效應(yīng)的比較如圖1所示。以費(fèi)氏弧菌為菌株的兩種生物毒性在線監(jiān)測(cè)儀器數(shù)據(jù)取平均值，與微生物燃料電池法水質(zhì)毒性在線監(jiān)測(cè)儀器數(shù)據(jù)進(jìn)行對(duì)比。

圖1 兩種水質(zhì)毒性在線監(jiān)測(cè)技術(shù)對(duì)不同濃度砷的毒性效應(yīng)(均值-極差)Fig.1 The toxicity effects of two different toxicity on-line monitoring technologies of water quality on different concentrations of arsenic (average-range)

從圖1可以看出，在實(shí)驗(yàn)室純水配水狀態(tài)下，兩種技術(shù)對(duì)砷的響應(yīng)都比較靈敏。高濃度條件下(濃度大于等于0.5 mg/L)，微生物燃料電池法的響應(yīng)靈敏度與發(fā)光細(xì)菌法相當(dāng)，在低濃度情況下明顯優(yōu)于發(fā)光細(xì)菌法(濃度小于等于0.1 mg/L)?？傮w上，微生物燃料電池法較發(fā)光細(xì)菌法更為靈敏。

2.2河水背景下兩種在線監(jiān)測(cè)技術(shù)對(duì)類金屬砷的毒性響應(yīng)對(duì)比

分別以廣東省韶關(guān)樂昌三溪水站枯水期、豐水期河水為背景基質(zhì)進(jìn)行兩種技術(shù)的對(duì)比，河水水質(zhì)基本理化參數(shù)如表2所示。可以看出，由于該水站上游存在大量有色金屬礦，水體本底金屬離子含量較高，其中砷、銻、鎘3種金屬在豐水期接近或超過《地表水環(huán)境質(zhì)量標(biāo)準(zhǔn)》(GB 3838—2002) III類水質(zhì)標(biāo)準(zhǔn)限值。

比較兩種在線監(jiān)測(cè)技術(shù)對(duì)不同濃度砷離子河水加標(biāo)樣本的信號(hào)與純水加標(biāo)樣本的信號(hào)，表3、圖2為抑制率響應(yīng)對(duì)比的情況。

表2 枯水期和豐水期河水水質(zhì)參數(shù)

注：“*”表示《地表水環(huán)境質(zhì)量標(biāo)準(zhǔn)》(GB 3838—2002)中基本項(xiàng)目III類水質(zhì)標(biāo)準(zhǔn)限值或特定項(xiàng)目標(biāo)準(zhǔn)限值; “—”表示未測(cè)試或無相關(guān)要求;“ND”表示未檢出。

表3 不同水質(zhì)毒性在線監(jiān)測(cè)技術(shù)在河水背景和純水背景下對(duì)不同濃度砷的毒性響應(yīng)

注：“—”表示未測(cè)試。

如圖2(a)所示，H公司生產(chǎn)的H-2010型發(fā)光細(xì)菌法水質(zhì)毒性在線監(jiān)測(cè)儀在河水背景下與純水背景相比，抑制信號(hào)呈增強(qiáng)趨勢(shì)，增強(qiáng)百分比為58.7%～175.3%。圖2(b)中，Z公司生產(chǎn)的Z-8000型發(fā)光細(xì)菌法水質(zhì)毒性在線監(jiān)測(cè)儀在河水背景下則呈現(xiàn)出相反的抑制信號(hào)減弱趨勢(shì)，與純水背景相比，抑制信號(hào)減弱29.7%～152.0%。此外，從圖2(a)和圖2(b)中可以看出，兩種發(fā)光菌法水質(zhì)毒性在線監(jiān)測(cè)儀在純水背景下砷的水質(zhì)毒性劑量-效應(yīng)關(guān)系有較好的一致性，這是第2.1小節(jié)中以費(fèi)氏弧菌為菌株的生物毒性在線監(jiān)測(cè)儀器數(shù)據(jù)取平均值的基礎(chǔ)，也是本小節(jié)中兩種發(fā)光細(xì)菌法水質(zhì)毒性在線監(jiān)測(cè)儀在河水背景與純水背景下對(duì)砷水質(zhì)毒性響應(yīng)比較的基礎(chǔ)，從而進(jìn)一步說明以費(fèi)氏弧菌為菌株的發(fā)光細(xì)菌生物毒性在線監(jiān)測(cè)儀數(shù)據(jù)在豐水期和枯水期兩種河水背景下的響應(yīng)信號(hào)與純水背景樣本相比均有較大差異，不同廠家儀器靈敏度差異顯著。

圖2 不同水質(zhì)毒性在線監(jiān)測(cè)技術(shù)在河水背景和純水背景下對(duì)不同濃度砷的毒性響應(yīng)Fig.2 The toxicity effects of different toxicity on-line monitoring technologies of water quality on different concentrations of arsenic in the background of river and pure water

圖2(c)中，國(guó)外某公司生產(chǎn)的S-2000型微生物燃料電池法水質(zhì)毒性在線監(jiān)測(cè)儀則表現(xiàn)出較好的穩(wěn)定性，河水背景與純水背景樣本的抑制響應(yīng)曲線差別不大，除在豐水期河水背景下，砷離子濃度為0.05 mg/L時(shí)與純水背景相比數(shù)據(jù)波動(dòng)較大外，枯水期數(shù)據(jù)和砷離子濃度為0.1 mg/L，豐水期數(shù)據(jù)與純水背景相比波動(dòng)僅在0～22.2%范圍變化。

根據(jù)文獻(xiàn)報(bào)道，由于河水背景中的營(yíng)養(yǎng)物質(zhì)和鹽分等因素，發(fā)光細(xì)菌的代謝加速，發(fā)光強(qiáng)度與純水相比會(huì)增強(qiáng)[31-32]，從而導(dǎo)致抑制信號(hào)減弱；另外，河水背景中仍然存在一些微量重金屬，加入砷后有可能會(huì)產(chǎn)生聯(lián)合毒性效應(yīng)，從而抑制信號(hào)增強(qiáng)。推測(cè)圖2(a)中在河水背景下抑制信號(hào)增強(qiáng)是由于該公司發(fā)光細(xì)菌菌種對(duì)重金屬的毒性效應(yīng)響應(yīng)比較靈敏，河水中存在的其他重金屬的毒性效應(yīng)也表現(xiàn)出來，增強(qiáng)了抑制信號(hào)。圖2(b)中在河水背景下抑制信號(hào)減弱推測(cè)是由于該公司發(fā)光細(xì)菌菌種對(duì)重金屬的毒性效應(yīng)響應(yīng)靈敏度一般，發(fā)光細(xì)菌在河水中的發(fā)光強(qiáng)度增強(qiáng)占了主導(dǎo)作用。綜上所述，發(fā)光細(xì)菌法受水體背景干擾大，不同生產(chǎn)廠家儀器測(cè)試的毒性效應(yīng)差異顯著。而微生物燃料電池法抗背景干擾能力強(qiáng)，有較強(qiáng)的穩(wěn)定性。

2.3對(duì)兩種生物毒性在線監(jiān)測(cè)儀的運(yùn)行建議

發(fā)光細(xì)菌法水質(zhì)生物毒性在線監(jiān)測(cè)技術(shù)受實(shí)際水樣的背景信號(hào)影響大，建議實(shí)際運(yùn)行時(shí)結(jié)合靈敏度需要用河水做標(biāo)線，建立該背景基質(zhì)條件下的劑量-效應(yīng)曲線，必要時(shí)可依此扣除背景基質(zhì)影響修正報(bào)警閾值。發(fā)光細(xì)菌法的不同設(shè)備廠家在線監(jiān)測(cè)儀器的信號(hào)差異顯著，推測(cè)是由于菌株來源不同、菌液濃度差異和儀器響應(yīng)靈敏度不同所造成。因此，在后續(xù)規(guī)范化運(yùn)行中，建議規(guī)范菌株來源，建立相應(yīng)的儀器標(biāo)準(zhǔn)和方法標(biāo)準(zhǔn)，以使儀器之間數(shù)據(jù)具有可比性和參考價(jià)值。

微生物燃料電池法水質(zhì)生物毒性在線監(jiān)測(cè)技術(shù)的缺點(diǎn)是微生物中毒后修復(fù)時(shí)間較長(zhǎng)，建議在設(shè)備中專門設(shè)置微生物培養(yǎng)罐，解決微生物被馴化和中毒后修復(fù)時(shí)間長(zhǎng)的問題。

3 結(jié)論

發(fā)光細(xì)菌法水質(zhì)生物毒性在線監(jiān)測(cè)技術(shù)對(duì)類金屬砷較為靈敏，但發(fā)光細(xì)菌法受水體背景干擾大，且不同生產(chǎn)廠家儀器的毒性效應(yīng)差異顯著。

微生物燃料電池法水質(zhì)生物毒性在線監(jiān)測(cè)技術(shù)對(duì)類金屬砷的響應(yīng)靈敏度高，且響應(yīng)信號(hào)受實(shí)際樣品背景干擾小，穩(wěn)定性高，河水加標(biāo)與純水加標(biāo)樣本的毒性響應(yīng)曲線差別不大。

兩種方法都有其優(yōu)勢(shì)與不足，因技術(shù)方法不同或相同方法使用不同廠家的菌種和儀器，造成同一濃度有毒物質(zhì)產(chǎn)生的毒性效應(yīng)有所差異。實(shí)際應(yīng)用于河流水污染毒性測(cè)試時(shí)應(yīng)根據(jù)所需監(jiān)測(cè)的毒性物質(zhì)、時(shí)間、成本等因素綜合比選技術(shù)方法與儀器設(shè)備。

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ComparisonResearchonTwoOn-lineBio-monitoringTechnologiesBasedonLuminescentBacteriaandMicrobialFuelCellsfortheWaterQualityToxicityofAs

YI Wen, YAN Huihua, LIN Xiurong, ZHANG Lin, HAN Shisong, WANG Shiqi

Guangdong Environmental Monitoring Centre, Guangzhou 510308，China

X830.2

A

1002-6002(2017)03- 0133- 06

10.19316/j.issn.1002-6002.2017.03.20

2016-06-01;

2017-03-02

中央重金屬污染防治資金項(xiàng)目“北江流域飲用水源水質(zhì)安全監(jiān)控預(yù)警關(guān)鍵技術(shù)研究與系統(tǒng)平臺(tái)建設(shè)”

易 雯(1963-)，女，湖南懷化人，教授級(jí)高級(jí)工程師。