微藻及其分泌物對(duì)溶解性有機(jī)磷礦化的促進(jìn)效應(yīng)

張曉燕,羅專溪,王振紅*

微藻及其分泌物對(duì)溶解性有機(jī)磷礦化的促進(jìn)效應(yīng)

張曉燕1,羅專溪2,王振紅1*

(1.閩南師范大學(xué)化學(xué)化工與環(huán)境學(xué)院,福建省現(xiàn)代分離分析科學(xué)與技術(shù)重點(diǎn)實(shí)驗(yàn)室,污染監(jiān)測(cè)與控制福建省高校重點(diǎn)實(shí)驗(yàn)室,福建 漳州 363000；2.華僑大學(xué)化工學(xué)院,福建 廈門 361021)

為較好認(rèn)識(shí)水生態(tài)環(huán)境中溶解性有機(jī)磷(DOP)的礦化過程及其影響因素,本實(shí)驗(yàn)選取典型的DOP類型(腺苷-5'-三磷酸二鈉鹽(ATP)、β-甘油磷酸鈉(βP)和D-葡萄糖-6-磷酸二鈉鹽(GP))、溫度(4,15,25℃)、銅綠微囊藻及其分泌物占比(& EPS)(0%、50%、90%)、砷酸鹽(As(V))濃度(0,10,100μg/L)4種環(huán)境因素,通過L9(34)正交試驗(yàn)探究影響DOP礦化過程的主要影響因素、水平及礦化過程中的環(huán)境變化.結(jié)果表明:& EPS作為主要影響因素可顯著促進(jìn)DOP的初期(前2d)礦化,之后被溫度取代,溫度升高有利于DOP的礦化.典型DOP礦化過程中藻細(xì)胞增殖對(duì)最佳溫度下(25℃)DOP的礦化起促進(jìn)作用,并使得βP與ATP環(huán)境下的pH值和氧化還原電位(ORP)產(chǎn)生較大波動(dòng).含As(V)環(huán)境中DOP礦化伴隨著As(V)形態(tài)的轉(zhuǎn)化,GP和ATP環(huán)境中均有明顯的三價(jià)砷(As(III))產(chǎn)生,使得砷的生態(tài)風(fēng)險(xiǎn)增強(qiáng),而溫度與& EPS占比增高可促進(jìn)βP環(huán)境中As(V)的甲基化.單獨(dú)DOP礦化可導(dǎo)致環(huán)境中以類蛋白質(zhì)組分為主的溶解性有機(jī)質(zhì)(DOM)含量顯著升高,而存在& EPS時(shí)則以可溶性微生物產(chǎn)物類DOM為主.研究結(jié)果對(duì)認(rèn)識(shí)水體中不同DOP的礦化過程及其可能導(dǎo)致的水華爆發(fā)和水環(huán)境變化所帶來的生態(tài)風(fēng)險(xiǎn)的科學(xué)管控具有重要意義.

正交試驗(yàn)；有機(jī)磷礦化；溫度；銅綠微囊藻；砷形態(tài)

近年來隨著對(duì)水體富營(yíng)養(yǎng)化的管控,外源性磷輸入受限,內(nèi)源性有機(jī)磷尤其是溶解性有機(jī)磷(DOP)在水體總磷中占比日漸突出(約占總磷的25%～ 50%)[1],使得作為水體潛在無機(jī)磷源之一的DOP礦化受到普遍關(guān)注.水體中DOP以植酸鹽、磷酸酯、核酸、核苷酸、磷脂、肌醇和有機(jī)磷農(nóng)藥等形態(tài)存在[2-3],其中以不穩(wěn)定的磷酸單酯類居多[4].DOP礦化伴隨的能量轉(zhuǎn)移和形態(tài)轉(zhuǎn)化,不僅能提供有效的磷源,同時(shí)其釋出的溶解性有機(jī)物亦可成為微生物的可利用性碳源[5],并由此可能引起藻華爆發(fā).

水體中DOP的礦化受溫度、pH值、細(xì)菌、溶解氧、磷酸酶等多種因素影響[6-7].研究表明DOP的礦化率隨溫度升高而增加[8-9],且在缺氧環(huán)境下被顯著促進(jìn)[10],偏酸或偏堿環(huán)境均有利于其礦化[11].浮游藻類因可分泌磷酸酶將DOP礦化為自身生長(zhǎng)代謝所需的營(yíng)養(yǎng)物質(zhì)而在DOP的礦化中扮演重要角色[12-15].銅綠微囊藻()因在適宜溫度下能快速增殖而成為富營(yíng)養(yǎng)化水體中常見的藍(lán)藻水華優(yōu)勢(shì)藻種之一[16],其增殖的同時(shí)可向環(huán)境中釋出包括多糖、蛋白質(zhì)、脂質(zhì)、腐殖質(zhì)及少量的堿性磷酸酶(AP)等多種胞外分泌物(EPS)[17],其中AP能水解DOP中的磷酸單酯[18],并由此增強(qiáng)微藻對(duì)低磷環(huán)境的適應(yīng)性[13].

砷(As)作為地殼中的微量過渡金屬元素在水體中普遍存在,砷酸鹽(As(V))因與磷酸鹽(PO43-)具有相似的化學(xué)結(jié)構(gòu),被認(rèn)為可通過PO43-轉(zhuǎn)運(yùn)途徑在藻體內(nèi)累積與轉(zhuǎn)化[19].研究認(rèn)為EPS中的AP能誘導(dǎo)藻體對(duì)As(V)的轉(zhuǎn)化并減少As(V)的累積[20],這使得As(V)對(duì)DOP的礦化因銅綠微囊藻及其分泌物(EPS)的制約而相互產(chǎn)生影響成為可能.同時(shí)不同類型DOP的礦化過程在一定程度上也存在差異[21].基于此本研究選取DOP類型、溫度、銅綠微囊藻及其分泌物(EPS)、砷酸鹽(As(V))濃度作為影響DOP礦化的主要影響因素,采用正交試驗(yàn)設(shè)計(jì)方法探討影響DOP礦化的主要因素及相關(guān)環(huán)境的變化,以更好地理解水環(huán)境中DOP的礦化過程及該過程可能產(chǎn)生的水生態(tài)環(huán)境風(fēng)險(xiǎn).

1 材料與方法

1.1 正交試驗(yàn)設(shè)計(jì)

選取有機(jī)多聚磷—腺苷-5'-三磷酸二鈉鹽(ATP)和小分子單酯磷—β-甘油磷酸鈉(βP)與D-葡萄糖-6-磷酸二鈉鹽(GP)為典型DOP類型,As(V)濃度參考《生活飲用水衛(wèi)生標(biāo)準(zhǔn)(GB/5749-2006)》和《地表水環(huán)境質(zhì)量標(biāo)準(zhǔn)(GB/3838-2002)》中As的規(guī)定限值并考慮無As污染作為對(duì)照設(shè)定[22-23],溫度則結(jié)合DOP的保存條件與藻的實(shí)際生長(zhǎng)環(huán)境確定,& EPS依據(jù)實(shí)際水體藻細(xì)胞狀況并以無& EPS作為對(duì)照設(shè)定,具體各因素及水平如表1所示,采用L9(34)正交試驗(yàn)設(shè)計(jì)(表2)探討影響DOP礦化的主要因素及相應(yīng)環(huán)境條件.

表1 正交試驗(yàn)的環(huán)境因素與水平

表2 L9(34)正交試驗(yàn)設(shè)計(jì)

1.2 M. a & EPS的來源與處理

銅綠微囊藻()FACHB-905,購(gòu)自中國(guó)科學(xué)院水生生物研究所,取1.5L正常培養(yǎng)的生長(zhǎng)狀態(tài)良好的藻液以4000r/min離心15min,棄掉上清液,加入同體積滅菌去離子水洗滌2次以除去磷酸鹽.將其轉(zhuǎn)移至滅菌的無磷BG11培養(yǎng)液中,置于培養(yǎng)箱中(25℃,光照強(qiáng)度3000lx,光暗比16h:8h)進(jìn)行3d的磷饑餓培養(yǎng),使藻細(xì)胞處于磷饑餓狀態(tài)[24].取磷饑餓培養(yǎng)后的藻細(xì)胞以4000r/min離心15min,棄去離心管底部的藻體,上清液為EPS,上清液中藻細(xì)胞密度經(jīng)流式細(xì)胞儀(美國(guó)BD C6Plus)測(cè)定為1.2×106cells/mL.

1.3 試驗(yàn)的準(zhǔn)備與操作

考慮無機(jī)磷及總磷的實(shí)際測(cè)定及藻細(xì)胞生長(zhǎng)的需要,設(shè)定設(shè)置GP、ATP與βP的濃度為1.0mg/L (以P計(jì)),使用Na3AsO4.12H2O配置As(V)標(biāo)準(zhǔn)儲(chǔ)備液.根據(jù)表2中的實(shí)驗(yàn)條件,分別配置100mL不同DOP溶液,各試驗(yàn)均設(shè)置3個(gè)平行.所有玻璃儀器須在10% HNO3溶液中浸泡24h后經(jīng)滅菌處理,以上實(shí)驗(yàn)過程均在無菌環(huán)境下完成.

1.4 最佳實(shí)驗(yàn)條件下DOP向無機(jī)磷的礦化過程

將GP、ATP和βP溶液分別與& EPS和滅菌超純水按1:9的比例進(jìn)行混合(總體積為100mL,& EPS占比為90%,P濃度為1.0mg/L),置于25℃培養(yǎng)箱中,連續(xù)觀測(cè)6d,于每天取樣10mL,測(cè)定其溶解性無機(jī)磷、pH值和氧化還原電位(ORP)的變化.

1.5 測(cè)定方法

采用流式細(xì)胞儀測(cè)定藻細(xì)胞密度,同時(shí)用雷磁pH計(jì)和ORP測(cè)量?jī)x(上?？祪x)分別測(cè)定溶液的pH值和ORP.將上清液經(jīng)0.22μm纖維素針式過濾器過濾,參照先前文獻(xiàn)中的方法采用高效液相色譜-等離子體質(zhì)譜聯(lián)用儀(HPLC-ICP-MS,美國(guó) Agilent 1200)測(cè)定砷形態(tài),流動(dòng)相為10mmol/L硝酸銨與10mmol/L磷酸氫二胺,并用優(yōu)級(jí)純硝酸調(diào)節(jié)pH至6.2,流速為1mL/min[25-26].采用鉬酸銨分光光度法測(cè)定溶解性無機(jī)磷和總磷的濃度.

DOP礦化率的計(jì)算公式為:

式中:為DOP礦化率,%;表示總磷濃度,mg/L;X為時(shí)刻的溶解性無機(jī)磷濃度,mg/L.

1.6 三維熒光光譜測(cè)定

將第0d和第6d含藻試驗(yàn)組溶液進(jìn)行離心分離,取上清液并經(jīng)0.45μm纖維素針式過濾器過濾后,采用熒光分光光度計(jì)(瓦里安Cary Eclipse)測(cè)定的三維熒光光譜,同時(shí)測(cè)定超純水.掃描條件:激發(fā)波長(zhǎng)(Ex)為200～450nm,掃描間距為10nm;發(fā)射波長(zhǎng)(Em)為250～550nm,掃描間距為2nm;狹縫寬度為20nm,掃描速度為1200nm/min.使用MATLAB軟件中的DOMFluor工具箱處理三維熒光數(shù)據(jù)并進(jìn)行平行因子(PARAFAC)分析,三維熒光光譜測(cè)定結(jié)果扣除超純水?dāng)?shù)據(jù)以去除拉曼散射的影響,結(jié)果表明3個(gè)組分模型能很好地通過殘差分析、拆半檢驗(yàn)、隨機(jī)初始化分析和 Tucker’s congruence coefficient(TCC)檢驗(yàn),說明結(jié)果具有可靠性[27-28].

1.7 數(shù)據(jù)分析及處理

采用 GraphPad Prism 8、Origin2019、MATLAB繪圖,正交實(shí)驗(yàn)結(jié)果用Minitab18進(jìn)行單因素方差分析(ANOVA),并用以下公式計(jì)算每個(gè)因素的貢獻(xiàn)百分比(PC)[29]:

式中:SST為總平方和;SSF為階乘平方和;Er為誤差的方差;DOF為自由度.

2 結(jié)果與討論

2.1 影響DOP礦化的主要因素

表3 不同時(shí)間下各因素對(duì)礦化率的影響主效應(yīng)

由正交試驗(yàn)得出6d內(nèi)各因素對(duì)DOP礦化率影響的主效應(yīng)結(jié)果如表3所示,可以看出不同時(shí)間下影響因素的排秩具有差異性,& EPS占比為DOP礦化第1d時(shí)的主要影響因素,之后則被溫度取代,溫度升高可促進(jìn)DOP的礦化,表現(xiàn)為25℃> 15℃>4℃.& EPS的存在有利于DOP礦化,而As(V)則對(duì)DOP礦化具有抑制效應(yīng).前3d內(nèi)ATP較GP和βP表現(xiàn)出更佳的礦化效果,之后則以GP的礦化效果更好.綜合得出25℃時(shí)& EPS占比為90%的無As(V)環(huán)境更有利于DOP的礦化.從不同DOP礦化率的均值主效應(yīng)圖(圖1)可以看出,影響DOP礦化的因素主次排秩為:T>& EPS>As (V)>DOP類型,溫度升高可促進(jìn)DOP的礦化,且25℃較15℃時(shí)DOP礦化率顯著增高.總體來看除溫度外其余3種因素對(duì)DOP礦化的影響均相對(duì)較小,且表現(xiàn)為隨& EPS占比增高礦化率增加的趨勢(shì),表現(xiàn)為微量藻的存在有利于DOP的礦化,這可能受藻細(xì)胞能分泌AP進(jìn)而對(duì)DOP礦化起促進(jìn)作用[30]. As(V)對(duì)DOP礦化的抑制效應(yīng)與其濃度呈正相關(guān),ATP表現(xiàn)出較GP和βP更易被礦化的趨勢(shì). DOP最佳礦化條件為:25℃時(shí)& EPS占比為90%不含As(V)的ATP環(huán)境,其礦化率于第5d達(dá)到100%.

圖1 不同DOP的礦化率的均值主效應(yīng)

由單因素方差分析得出不同時(shí)間下各因素對(duì)DOP礦化率的影響在不同水平間均具有顯著性差異(< 0.05),其貢獻(xiàn)占比如圖2所示,可以看出& EPS在轉(zhuǎn)化前2d作為主要影響因素影響DOP的礦化,第1d其占4個(gè)因素總貢獻(xiàn)的51.5%,是溫度貢獻(xiàn)的3倍,第2d則與溫度影響接近,占總貢獻(xiàn)的36.0%,之后則以溫度影響占絕對(duì)優(yōu)勢(shì),占總貢獻(xiàn)的90.5%以上,且隨時(shí)間趨于穩(wěn)定.& EPS在礦化第1d表現(xiàn)出隨其占比增高DOP礦化率顯著增加的趨勢(shì),其對(duì)DOP礦化率的促進(jìn)是其他因子的2倍,第2d對(duì)礦化率的促進(jìn)作用與溫度基本一致,且表現(xiàn)為其占比為50%和90%時(shí)無顯著差異,均較占比為0%時(shí)的顯著增高.

圖2 不同時(shí)間下各影響因素對(duì)DOP礦化率的貢獻(xiàn)占比

圖3 不同溫度下不同類型DOP礦化率隨時(shí)間的變化特征

3種類型DOP在不同溫度下礦化率隨時(shí)間變化如圖3所示, 25℃時(shí)除GP環(huán)境中的T5試驗(yàn)組前2d礦化率相對(duì)較低,且低于T4(15℃)和T6(4℃)試驗(yàn)組,其余兩種DOP環(huán)境下均顯著高于15和4℃試驗(yàn)組,且DOP礦化率隨時(shí)間呈顯著增高趨勢(shì),該溫度下第5d時(shí)T5(GP)試驗(yàn)組礦化率達(dá)到88.1%之后趨于平穩(wěn),T3(βP)和T7(ATP)試驗(yàn)組則分別于第6d時(shí)礦化率達(dá)到100%.15℃時(shí)T9(ATP)試驗(yàn)組幾乎沒發(fā)生礦化,說明該溫度下As(V)的存在不會(huì)影響ATP的礦化,而T2(βP)與T4(GP)試驗(yàn)組均分別高于4℃時(shí)的T1和T6試驗(yàn)組,第6d時(shí)礦化率分別達(dá)到14.4%和28.4%說明即使EPS占比增加但該溫度下DOP的礦化率并不高,這可能與該溫度下藻的生長(zhǎng)相對(duì)緩慢有關(guān)[31].4℃時(shí),T1試驗(yàn)組的βP幾乎沒有礦化,T6(GP)和T8(ATP)試驗(yàn)組因EPS的存在其礦化率在第1d時(shí)略有升高,之后則保持平穩(wěn)態(tài)勢(shì),GP和ATP的平均礦化率分別為(6.0%±1.7%)和(10.5%±1.8%),這可能是由于初始時(shí)EPS中攜帶的AP對(duì)DOP的礦化作用所致,之后則由于低溫環(huán)境藻的生長(zhǎng)與生物化學(xué)反應(yīng)停止,使其礦化率保持穩(wěn)定.

2.2 最佳條件下DOP的礦化率

根據(jù)正交試驗(yàn)確定的DOP礦化的最佳條件:25℃時(shí)& EPS占比為90%的不含As(V)環(huán)境,得到的該環(huán)境下ATP、GP和βP的實(shí)際礦化率隨時(shí)間變化如圖4所示.可以看出ATP、GP和βP 3種DOP的礦化率均隨時(shí)間呈增高趨勢(shì),在第6d時(shí)礦化率分別達(dá)到86.9%、87.3%和90.0%,較該環(huán)境下的預(yù)測(cè)值100%均相對(duì)偏低,其中ATP礦化率在第4d達(dá)到83.7%,分別高出βP和GP的29.8%和35.4%.與前期研究相比較得出& EPS對(duì)βP和GP在前4d的礦化率與單獨(dú)水環(huán)境的空白組相比無顯著差異,從第5d起則顯著高于空白組,而ATP則表現(xiàn)為& EPS的添加與空白組隨時(shí)間均無顯著差異[32].較單獨(dú)EPS環(huán)境相比較& EPS的添加均顯著促進(jìn)了DOP的礦化,與前期研究得到的EPS在一定程度會(huì)抑制DOP的礦化結(jié)果不同[32],說明藻細(xì)胞的存在對(duì)DOP的礦化具有驅(qū)動(dòng)作用.

圖4 最佳條件下不同類型DOP礦化率隨時(shí)間的變化特征

2.3 DOP礦化過程中主要的環(huán)境變化

2.3.1 含& EPS試驗(yàn)組中的藻細(xì)胞密度變化 含& EPS試驗(yàn)組中微藻細(xì)胞密度在礦化前后變化如圖5所示,& EPS占比為50%(T2、T6和T7試驗(yàn)組)和90%(T3、T4和T8試驗(yàn)組)對(duì)應(yīng)的藻細(xì)胞密度分別為6.00×105cells/mL和1.08×106cells/ mL.可以看出第6d時(shí)T3和T7試驗(yàn)組的藻細(xì)胞密度較初始相比均相對(duì)增高,顯著大于其余試驗(yàn)組(<0.05),分別達(dá)到5.29×106cells/mL和1.03× 106cells/mL,是第0d的4.90倍和1.72倍,而其他試驗(yàn)組則顯著降低.由先前研究可知銅綠微囊藻能以βP、ATP和GP分別作為單一磷源進(jìn)行增殖,且對(duì)藻的生長(zhǎng)無顯著差異[19],因此適宜微藻增殖的溫度下(25℃),T3較T7試驗(yàn)組的& EPS占比高,使得T3試驗(yàn)組藻細(xì)胞顯著高于T7試驗(yàn)組,該溫度下微藻細(xì)胞增殖驅(qū)動(dòng)了DOP的高礦化率,這與Bai等研究得出銅綠微囊藻可促進(jìn)磷酸單酯(Mono-P)的轉(zhuǎn)化相一致[6,8].15℃(T2和T4試驗(yàn)組)和4℃(T4和T8試驗(yàn)組)環(huán)境下藻細(xì)胞增殖受到抑制,不同試驗(yàn)組間無顯著差異(>0.05),這與Yang等研究發(fā)現(xiàn)低溫抑制微囊藻的生長(zhǎng)一致[33].

圖5 DOP礦化過程中含M. a & EPS組的藻細(xì)胞密度

2.3.2 含As(V)試驗(yàn)組中的砷形態(tài)轉(zhuǎn)化 各DOP經(jīng)6d礦化后其含As(V)試驗(yàn)組中的砷形態(tài)變化如圖6所示,可以看出DOP礦化同時(shí)伴隨As(V)形態(tài)的轉(zhuǎn)化,T2(βP,15℃,& EPS占比50%)和T3(βP, 25℃,& EPS占比90%)試驗(yàn)組均有二甲基砷(DMA)檢出,且T3試驗(yàn)組發(fā)現(xiàn)有單甲基砷(MMA),說明& EPS促進(jìn)βP礦化的同時(shí)也促進(jìn)介質(zhì)中As(V)的甲基化.T5、T6、T8和T9試驗(yàn)組中均有亞砷酸鹽As(III)檢出,其中T5試驗(yàn)組(GP,25℃,& EPS占比0%)中As(III)占總砷(TAs)的83.9%,說明該環(huán)境有利于As(V)的還原,究其原因可能是GP礦化生成還原性有機(jī)物而導(dǎo)致[34].T6(GP,4℃,& EPS占比50%)較T5試驗(yàn)組As(III)占比減少,占TAs的11.5%,說明盡管低溫抑制了GP的礦化,但& EPS的存在一定程度上也能引起As(V)的還原,這與Wang等研究發(fā)現(xiàn)GP可促進(jìn)As(V)的還原相一致[19]. T8(ATP,4℃,& EPS占比90%)和T9(ATP,15℃,& EPS占比0%)試驗(yàn)組中As(III)占比接近,分別為12.4%、14.0%,說明盡管ATP礦化率低,但溫度增高引起的As(V)的還原比& EPS占比為90%所導(dǎo)致的As(V)的還原相對(duì)要高,反映出溫度增高更有利于該環(huán)境中As(V)的還原.T5和T9試驗(yàn)組中As(III)的檢出表明DOP環(huán)境下溫度升高會(huì)增加水中As的生態(tài)風(fēng)險(xiǎn).

圖6 DOP礦化過程中含As(V)環(huán)境中的砷形態(tài)變化

2.3.3 環(huán)境中的pH值變化 不同試驗(yàn)組DOP礦化過程中水體pH值的變化如圖7所示,可以看出不含& EPS試驗(yàn)組中的pH值均相對(duì)較低,T1(βP)和T5(GP)試驗(yàn)組中的pH值在(6.62±0.17)范圍內(nèi)波動(dòng),而T9(ATP)試驗(yàn)組的pH值隨時(shí)間變化呈下降趨勢(shì);其余各試驗(yàn)組在第1d時(shí)pH值均顯著降低,說明環(huán)境中的DOP對(duì)& EPS引起pH值的增高具有緩沖效應(yīng).& EPS占比為50%時(shí),除第0d外,T2(βP)和T6(GP)試驗(yàn)組pH值為(7.22±0.05),但T7(ATP)試驗(yàn)組中的pH值波動(dòng)較大,第3d其pH值升高至7.91..& EPS占比為90%時(shí),T3(βP)試驗(yàn)組pH值呈先上升后下降的趨勢(shì);T4(GP)和T8(ATP)試驗(yàn)組的pH值為(7.52±0.22).T3(βP)和T7(ATP)試驗(yàn)組較大的pH值波動(dòng)可能是由于在適宜微藻生長(zhǎng)的溫度下,較高的& EPS占比提升了環(huán)境的堿度[35],使DOP處于堿性礦化環(huán)境.

2.3.4 環(huán)境中的ORP變化 不同試驗(yàn)組DOP礦化過程中水體ORP隨時(shí)間變化如圖8所示,各DOP環(huán)境中不含& EPS的T1、T5和T9試驗(yàn)組與其余試驗(yàn)組相比ORP均相對(duì)偏高,T1(βP)和T5(GP)環(huán)境中第0d的ORP分別為207mV和211mV,從第1d顯著升高隨后變化不大;T9(ATP)試驗(yàn)組ORP前3d相對(duì)穩(wěn)定,為(299±8)mV,第4d顯著增高至428mV,之后趨于穩(wěn)定.& EPS占比為50%時(shí),T2、T6和T7試驗(yàn)組中ORP除第1d顯著增高外,其余時(shí)間相對(duì)穩(wěn)定,分別為(212±23)mV、(247±14)mV和(238±16)mV.當(dāng)& EPS占比增高為90%時(shí),T3試驗(yàn)組中ORP從第2d起呈緩慢降低至第6d時(shí)為181mV,T4試驗(yàn)組除第1d顯著增高外其余時(shí)間保持穩(wěn)定為(247±13)mV,T8試驗(yàn)組中ORP隨時(shí)間變化略有升高,至第4d時(shí)最高為317mV之后又趨于降低.T3試驗(yàn)組ORP有所減小,可能是由于在適宜微藻生長(zhǎng)的溫度下使得環(huán)境的還原性增強(qiáng),更有利于DOP礦化.

圖7 不同類型DOP礦化過程中水環(huán)境pH值隨時(shí)間變化

由各試驗(yàn)組中得到pH值、ORP、DOP礦化率與各影響因子間的相關(guān)分析可知:ORP與pH值、& EPS占比和DOP礦化率均顯著負(fù)相關(guān),相關(guān)系數(shù)分別為-0.667、-0.453和-0.223(<0.05),與As(V)濃度呈顯著正相關(guān),相關(guān)系數(shù)為0.206(<0.01);說明水體pH值升高和& EPS占比增加與DOP礦化率增高可導(dǎo)致水體ORP降低,同時(shí)As(V)濃度增高會(huì)引起水體ORP上升.& EPS占比與pH值顯著正相關(guān),與ORP顯著負(fù)相關(guān),表明& EPS占比增加可引起水體pH值升高與ORP顯著降低;DOP礦化率與溫度和pH值均顯著正相關(guān),相關(guān)系數(shù)分別為0.670和0.307(<0.01),與ORP顯著負(fù)相關(guān),相關(guān)系數(shù)為-0.223(<0.01),說明溫度升高和水體pH值升高均會(huì)促進(jìn)DOP礦化并引起水體ORP降低.總體得出環(huán)境溫度升高與& EPS介導(dǎo)的水體pH值升高能促進(jìn)DOP礦化.

圖8 不同類型DOP礦化過程中水環(huán)境中的ORP隨時(shí)間變化

2.4 三維熒光光譜分析

各試驗(yàn)組溶解性有機(jī)質(zhì)(DOM)的三維熒光光譜結(jié)果通過PARAFAC模型共解析出3個(gè)主要化學(xué)組分,模型可靠性通過拆半檢驗(yàn)和殘差分析得到充分驗(yàn)證,各組分的三維熒光光譜及載荷如圖9所示,C1在x/m=280/334nm處有單一激發(fā)和發(fā)射峰,歸類為可溶性微生物產(chǎn)物組分,C2在x/m= 220/334nm處有單一激發(fā)和發(fā)射峰,歸類為類蛋白質(zhì)組分,C3在x/m=340/436nm處有激發(fā)和發(fā)射主峰,x/m= 260/436nm處有二級(jí)峰,歸類為類腐殖酸組分[17,36].

圖9 不同試驗(yàn)組DOM中3個(gè)組分熒光光譜及載荷

用平行因子分析得到不同試驗(yàn)組DOM的最大熒光強(qiáng)度值(Fmax)表示各樣品中不同組分的含量[37].不同試驗(yàn)組DOP礦化前后各組分的Fmax及其相對(duì)占比如圖10所示,由圖可知,DOP經(jīng)6d礦化后其Fmax值均相對(duì)增高,礦化后的含& EPS試驗(yàn)組中DOM主要為C1組分(可溶性微生物產(chǎn)物),不同試驗(yàn)組礦化前后其在3個(gè)組分中的占比分別為(64.2%±6.4%)和(71.9%±6.8%);而無& EPS試驗(yàn)組中DOM主要為C2組分(類蛋白質(zhì)),礦化前后其占比分別為(90.4%±1.9%)和(78.6%±3.8%),主要是由于選取的這3種DOP均與類蛋白質(zhì)的合成有關(guān)[38].T3與T7試驗(yàn)組礦化后C1組分的Fmax值顯著增高,這主要是由于該試驗(yàn)組適宜的環(huán)境溫度(25℃)促進(jìn)了& EPS中藻細(xì)胞的增殖,進(jìn)而向環(huán)境分泌較多的可溶性微生物產(chǎn)物.T2和T4試驗(yàn)組礦化后C1組分的Fmax值及其占比都有所增加,可能是15℃下藻細(xì)胞分泌了少量的可溶性微生物產(chǎn)物.T6和T8試驗(yàn)組礦化后3個(gè)組分的Fmax值均無明顯變化,可能是低溫(4℃)抑制了藻細(xì)胞的代謝以及DOP的礦化.在& EPS占比為0%時(shí),T1、T5和T9試驗(yàn)組中3個(gè)組分的Fmax值均相對(duì)增大,其中C1組分占比增加,而C2組分有所降低,C3組分占比無明顯變化,說明DOP礦化產(chǎn)生了一些可以發(fā)熒光的官能團(tuán),C2組分代表生物降解來源的類色氨酸物質(zhì),與微生物降解產(chǎn)生的芳香性蛋白類結(jié)構(gòu)有關(guān)[35],說明βP、ATP和GP 3種DOP礦化后使得其相應(yīng)的蛋白類組分出現(xiàn)分解.

3 結(jié)論

3.1& EPS在前2d對(duì)DOP的礦化影響顯著,之后則被溫度取代.溫度升高和& EPS占比增高可促進(jìn)DOP的礦化;同時(shí)ATP表現(xiàn)出較GP和βP更易礦化.25℃時(shí)& EPS占比為90%不含As(V)的最佳礦化條件下,3種DOP第6d時(shí)的礦化率均達(dá)到86.0%以上.

3.2 DOP礦化過程中適宜的溫度能促進(jìn)藻細(xì)胞增殖,使得βP與ATP環(huán)境中pH值增高、ORP降低.含As(V)環(huán)境中DOP礦化可引起As(V)形態(tài)轉(zhuǎn)化;βP環(huán)境中& EPS占比和溫度增高所導(dǎo)致的DOP礦化能促進(jìn)As的甲基化,而GP和ATP的礦化則伴隨著還原性As(III)的產(chǎn)生,使得As的水生態(tài)風(fēng)險(xiǎn)增加.

3.3 DOP礦化會(huì)引起水中DOM含量顯著增高,僅DOP時(shí)水體DOM以類蛋白質(zhì)組分為主,而& EPS存在時(shí)則以可溶性微生物產(chǎn)物組分為主.

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Promoting effect of microalgaend its secretions on dissolved organophosphate mineralization.

ZHANG Xiao-yan1, LUO Zhuan-xi2, WANG Zhen-hong1*

(1.School of Chemistry, Chemical Engineering and Environment, Minnan Normal University, Fujian Provincial Key Laboratory of Modern Separation and Analysis Science and Technology, Fujian Provincial Key Laboratory of Pollution Monitoring and Control, Zhangzhou 363000, China；2.School of Chemical Engineering, Huaqiao University, Xiamen 361021, China)., 2023,43(1)：341～350

In order to better understand the mineralization process of dissolved organic phosphorus (DOP) and its related influencing factors in the aquatic ecological environment, the typical DOP types (adenosine-5'-triphosphate disodium salt (ATP), sodium β-glycerophosphoric and D-glucose-6-phosphate disodium salt (GP)), temperatures (4, 15 and 25℃), proportions ofand its secretions (& EPS) (0%, 50% and 90%), arsenate (As(V)) concentration (0, 10 and 100μg/L) were selected as the main four environmental factors(three levels for each). The L9(34) orthogonal test was then used to explore the main influencing factors and levels of DOP mineralization process and the main environmental changes during the mineralization process. Results showed that& EPS, as the main influencing factor, could significantly promote the mineralization of DOP at the initial stage (the first 2days), and then was replaced by temperature. Herein the increase of temperature was beneficial to the mineralization of DOP. During the typical DOP mineralization process, the proliferation of algal cells promoted the mineralization of DOP at the optimal temperature(25℃), and caused large fluctuations in pH and ORP in the environment of βP and ATP. The mineralization of DOP in the As(V)-containing environment was accompanied by the transformation of As(V) species. In particular, the production of arsenite (As(III)) was significant in both GP and ATP environments, which could enhance the ecological risk of arsenic. The increased proportion of& EPS promoted the methylation of As(V) in the βP environment. Additionally, the mineralization of individual DOP led to a significant increase in the content of soluble organic matter (DOM) in the environment, which mainly accounted to protein-like components. Meanwhile, the mineralization of DOP combined with& EPS also increased the DOM but with soluble microbial products.The obtain results are of great significance for the comprehensive understanding of the mineralization process of different DOPs in water bodies and the scientific management and control of the ecological risks caused by the outbreak of algal blooms and changes in the water environment.

orthogonal test；organophosphorus mineralization；temperature；；arsenic species

X172

A

1000-6923(2023)01-0341-10

張曉燕(1996-),女,閩南師范大學(xué)碩士研究生,研究方向?yàn)樗h(huán)境化學(xué).發(fā)表論文1篇.

2011-06-07

國(guó)家自然科學(xué)基金項(xiàng)目(42177384);福建省自然科學(xué)基金資助項(xiàng)目(2020J01804);福建省高校產(chǎn)學(xué)合作項(xiàng)目(2022N5002)

* 責(zé)任作者, 教授, zhhwang1979@163.com