王聶穎,張 輝,隋 陽(yáng),2,陳約余,胡 南,戴仲然,丁德馨*

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介導(dǎo)植酸鹽水解促進(jìn)U(VI)-PO43-生物礦化

王聶穎1,張 輝1,隋 陽(yáng)1,2,陳約余1,胡 南1,戴仲然1,丁德馨1*

(1.南華大學(xué),鈾礦冶生物技術(shù)國(guó)防重點(diǎn)學(xué)科實(shí)驗(yàn)室, 極貧鈾資源綠色開發(fā)技術(shù)湖南省重點(diǎn)實(shí)驗(yàn)室,湖南 衡陽(yáng) 421001；2.南華大學(xué)核科學(xué)技術(shù)學(xué)院,湖南 衡陽(yáng) 421001)

從廣東某鈾尾礦庫(kù)水下沉積物中分離篩選出了一株能水解植酸鹽的真菌M5-1,對(duì)其菌落形態(tài)、ITS序列、最適生長(zhǎng)pH值、對(duì)鈾的耐受性及其水解植酸鹽的效果進(jìn)行了分析,隨后對(duì)M5-1生物礦化鈾過(guò)程中pH值、正磷酸鹽濃度、鈾濃度、鈾去除率的變化進(jìn)行了監(jiān)測(cè),對(duì)礦化產(chǎn)物的主要元素和礦物組成進(jìn)行了分析.證實(shí)了真菌M5-1為(MH978623),其最適生長(zhǎng)pH值范圍為6~7,對(duì)鈾(~0.84mmol/L)具有較強(qiáng)的耐受性;介導(dǎo)植酸鹽水解促進(jìn)U(VI)-PO43-礦化62d后,鈾的去除率達(dá)95.2%;介導(dǎo)U(VI)-PO43-礦化過(guò)程中可能形成了難溶的氫鈾云母和變鈉鈾云母礦物.結(jié)果表明,能有效水解植酸鹽釋放可溶性正磷酸鹽,從而促進(jìn)U(VI)-PO43-礦化.研究結(jié)果為采用介導(dǎo)植酸鹽水解原位修復(fù)鈾污染地表水提供了試驗(yàn)依據(jù).

鈾污染地表水；U(VI)；；生物礦化；植酸鹽水解

在核燃料循環(huán)過(guò)程中,都會(huì)有少量的鈾釋放到環(huán)境中,使部分水體受到一定程度的污染[1-2].進(jìn)入水體的鈾可通過(guò)食物鏈富集到人體內(nèi),以重金屬毒性和內(nèi)照射2種方式誘發(fā)慢性放射性疾病,對(duì)人體的健康造成威脅[3-4].因此,鈾污染水體的修復(fù)方法已成為環(huán)境領(lǐng)域研究的重點(diǎn).

原位生物修復(fù)技術(shù)與傳統(tǒng)修復(fù)方法相比,具有設(shè)計(jì)簡(jiǎn)單、可行性高、對(duì)環(huán)境干擾小和經(jīng)濟(jì)性好等優(yōu)點(diǎn),是最具發(fā)展前景的鈾污染水體修復(fù)技術(shù)之一,受到了廣泛關(guān)注[5-6].近年來(lái),國(guó)內(nèi)外研究人員在鈾的原位生物還原方面開展了大量研究.但是,研究發(fā)現(xiàn)生物還原過(guò)程形成的U(IV)礦物容易被O2、Fe3+、硝酸鹽等氧化劑氧化,使鈾再次釋放遷移[7-9].而且,一些鈾污染地區(qū)具有pH值低、硝酸鹽濃度高、處于氧化狀態(tài)等特點(diǎn),從而限制了生物還原技術(shù)的應(yīng)用[10].

于是,研究人員又開始研究鈾的原位生物礦化,并發(fā)現(xiàn)與生物還原相比,它的適用范圍更廣、產(chǎn)物更穩(wěn)定,被認(rèn)為是一種最具發(fā)展?jié)摿Φ拟櫸廴舅w修復(fù)方法[11].U(VI)的生物礦化通常是指U(VI)被微生物代謝過(guò)程中產(chǎn)生的PO43-等配位體礦化形成難溶的沉淀[11].近年來(lái),研究人員發(fā)現(xiàn),很多細(xì)菌和真菌在鈾的脅迫下能產(chǎn)生磷酸酶,促進(jìn)有機(jī)磷的水解以及釋放正磷酸鹽[12].Liang等[13]研究了黑曲霉和爪哇擬青霉以G2P為唯一磷源時(shí)固定溶液中鈾的行為,發(fā)現(xiàn)在菌絲表面沉淀了大量的U-P礦物.植酸鹽是一種經(jīng)濟(jì)高效的天然有機(jī)磷,可以被磷酸酶催化水解為低級(jí)肌醇衍生物(IP,=1~5)和正磷酸鹽[14].以植酸鹽為有機(jī)磷源,利用微生物介導(dǎo)植酸鹽水解促進(jìn)U-PO43-礦化,是一種實(shí)現(xiàn)鈾污染水體修復(fù)的可行方法.

本研究采用平板分離法從廣東韶關(guān)某鈾尾礦庫(kù)水下沉積物中分離篩選了能高效水解植酸鹽的微生物,探索了該微生物以植酸鹽為唯一磷源時(shí)對(duì)溶液中鈾的礦化作用,監(jiān)測(cè)了礦化過(guò)程中pH值、正磷酸鹽和鈾濃度的變化,并利用SEM-EDS和XRD分析了礦化產(chǎn)物的主要元素和礦物組成,以期為采用微生物介導(dǎo)植酸鹽水解原位修復(fù)鈾污染地表水提供試驗(yàn)依據(jù).

1 材料與方法

1.1 材料

1.1.1 樣品 試驗(yàn)所用的沉積物樣品,于2017年12月25日采自廣東某鈾尾礦庫(kù)2m深蓄水池底部的沉積物. 沉積物取樣后立即裝入無(wú)菌密封取樣袋中,覆蓋冰袋保存.運(yùn)回實(shí)驗(yàn)室后放入-20℃冰箱保存.

1.1.2 試驗(yàn)用培養(yǎng)基 平板篩選培養(yǎng)基[15]:植酸鈣0.3%, MgSO4·7H2O 0.05%, MnSO4·4H2O 0.05%, FeSO4·7H2O 0.003%, KCl 0.05%, NH4NO30.05%,葡萄糖 1.5%,瓊脂粉1.8%.

改良馬丁液體培養(yǎng)基[16]:蛋白胨0.5%,酵母浸膏0.2%,葡萄糖2%, KH2PO40.1%, MgSO4·7H2O 0.05%.

1.2 方法

1.2.1 菌種分離與篩選 取沉積物10g,放入盛有90mL無(wú)菌生理鹽水和數(shù)粒玻璃珠的無(wú)菌錐形瓶中,放入搖床震蕩30min,取出后分別制成10-3,10-4, 10-5,10-6,10-7,10-8,10-9稀釋度的懸液.將不同稀釋度的懸液分別涂布于pH值為4,5,6的平板篩選培養(yǎng)基上(此處將pH值設(shè)為4,5,6是由于鈾尾礦庫(kù)水中的pH值約為5),每個(gè)實(shí)驗(yàn)設(shè)3個(gè)平行樣.將涂布好的平板放入恒溫培養(yǎng)箱中30℃培養(yǎng),每隔12h觀察是否產(chǎn)生水解圈.

挑取水解圈與菌落直徑之比較大的菌落多次接種,平板劃線分離,獲得一系列單菌落,再挑取單菌落重復(fù)以上操作,直至獲得純的培養(yǎng)菌株,同時(shí)將純化后的目標(biāo)菌株用甘油冷凍(-80℃)保存.

1.2.2 菌株M5-1菌落特征及其分子生物學(xué)鑒定 M5-1菌落特征:將1.2.1節(jié)分離到的一株可以高效水解植酸鹽的真菌編號(hào)為M5-1.用無(wú)菌接種環(huán)將M5-1接種于改良馬丁固體培養(yǎng)基上,30℃恒溫培養(yǎng)5d,觀察并描述菌落形狀、質(zhì)地和顏色等形態(tài)特征.

M5-1分子生物學(xué)鑒定:采用真菌核糖體內(nèi)轉(zhuǎn)錄間隔區(qū)通用引物ITS1和ITS4進(jìn)行PCR擴(kuò)增.PCR采用50μL反應(yīng)體系: PCRMix 25μL,ITS1(10μmol/L) 2.0μL,ITS4(10μmol/L)2.0μL,DNA template 1.0μL,加無(wú)菌ddH2O補(bǔ)足.PCR擴(kuò)增條件: 98℃ 2min;98℃ 10s,54℃ 10s,72℃ 10s,35個(gè)循環(huán);72℃ 5min.由上海美吉生物科技有限公司提供PCR擴(kuò)增產(chǎn)物測(cè)序.所得ITS序列數(shù)據(jù)在NCBI在線數(shù)據(jù)庫(kù)進(jìn)行Blast序列相似性比對(duì),使用MEGA4.1生成系統(tǒng)發(fā)育樹.將測(cè)序數(shù)據(jù)提交至GenBank,申請(qǐng)序列號(hào).

1.2.3 M5-1最適生長(zhǎng)pH值和對(duì)鈾的耐受性 250mL錐形瓶中加入100mL改良馬丁液體培養(yǎng)基,將培養(yǎng)基的初始pH值分別調(diào)至3,4,5,6,7,8,9,高壓蒸汽滅菌之后加入0.1mL OD600值為1.0的孢子懸液,每組實(shí)驗(yàn)設(shè)3個(gè)平行樣.放入恒溫?fù)u床中,180r/min、30℃培養(yǎng)7d后用濾紙過(guò)濾收集菌絲體,60℃烘干至恒重,稱量菌絲體干質(zhì)量.

250mL錐形瓶中加入100mL改良馬丁液體培養(yǎng)基,向錐形瓶中加入一定體積4.20mmol/L的鈾母液,分別配制成鈾濃度為0.084,0.21,0.42,0.63, 0.84mmol/L的液體培養(yǎng)基,以不添加鈾母液的處理為對(duì)照組,將液體培養(yǎng)基的pH值調(diào)為5,過(guò)濾滅菌.向每個(gè)錐形瓶中加入0.1mL OD600值為1.0的孢子懸液,每組實(shí)驗(yàn)設(shè)3個(gè)平行樣.放入恒溫?fù)u床中, 180r/min、30℃培養(yǎng)7d后用濾紙過(guò)濾收集菌絲體, 60℃烘干至恒重,稱量菌絲體干質(zhì)量.

1.2.4 M5-1水解植酸鹽效果 向250mL錐形瓶中加入100mL不添加KH2PO4的改良馬丁液體培養(yǎng)基,8.4mmol/L植酸鈉,調(diào)節(jié)初始pH值為5(此處將pH值設(shè)為5是因?yàn)殁櫸驳V庫(kù)水中的pH值約為5,且M5-1分離自pH值為5的固體培養(yǎng)基上),過(guò)濾滅菌后加入0.1mL OD600值為1.0的孢子懸液,以不接種M5-1的處理為對(duì)照組,每組實(shí)驗(yàn)設(shè)3個(gè)平行樣.放入恒溫?fù)u床中,180r/min、30℃培養(yǎng),每隔一段時(shí)間取上清液測(cè)pH值和正磷酸鹽濃度.

1.2.5 鈾和植酸鹽的非生物沉淀 為了確保生物礦化實(shí)驗(yàn)中植酸鹽不與鈾以非生物方式形成沉淀,有必要研究鈾和植酸鹽之間的化學(xué)相互作用.250mL錐形瓶中加入100mL 168μmol/L的鈾標(biāo)準(zhǔn)溶液,按 [Na12-IP6]:[U]的物質(zhì)的量比為1:4,1:2, 1:1,5:1, 10:1,15:1,25:1,50:1添加植酸鈉,調(diào)節(jié)溶液pH值為5,每組實(shí)驗(yàn)設(shè)3個(gè)平行樣.室溫下平衡7d后取上清液測(cè)鈾濃度.

1.2.6 M5-1介導(dǎo)植酸鹽水解促進(jìn)U-PO43-礦化實(shí)驗(yàn) 實(shí)驗(yàn)分為A、B 2組,每組設(shè)3個(gè)平行樣.250mL錐形瓶中加入100mL改良馬丁液體培養(yǎng)基,實(shí)驗(yàn)條件如表1所示.A、B 2組放入恒溫?fù)u床中,180r/min、30℃培養(yǎng).每隔一段時(shí)間取上清液測(cè)pH值、鈾濃度和可溶性正磷酸鹽濃度.最后的沉淀用超純水洗滌8次后用于SEM-EDS和XRD分析.

表1 實(shí)驗(yàn)分組和條件

1.2.7 檢測(cè)方法 溶液pH值采用pH計(jì)測(cè)量(HACH,HQ430D,USA);鈾濃度采用5-Br-PADAP分光光度法測(cè)量(T6,新世紀(jì),北京)[17];可溶性正磷酸鹽濃度采用改進(jìn)的鉬酸銨分光光度法測(cè)量(T6,新世紀(jì),北京)[18];沉淀的元素組成用掃描電鏡能譜儀分析(FEI,Quanta 250,USA);沉淀的礦物類型用X射線全自動(dòng)衍射儀分析(Bruer Axs,D8Advance,GER).

1.2.8 數(shù)據(jù)處理與分析 用Microsoft Excel 2016軟件整理試驗(yàn)數(shù)據(jù),計(jì)算平均值及標(biāo)準(zhǔn)差;用Origin 2016繪圖;用Jade 6.0匹配XRD圖譜;用SPSS 20.0進(jìn)行方差分析,通過(guò)Tukey多重比較法檢驗(yàn)處理間的差異顯著性(<0.05),用不同小寫字母表示差異顯著性.

2 結(jié)果與討論

2.1 菌株M5-1菌落形態(tài)

(a)在平板篩選培養(yǎng)基上形成的水解圈;(b)正面的形態(tài);(c)反面的形態(tài)

2.2 M5-1鑒定結(jié)果及系統(tǒng)進(jìn)化分析

采用平板篩選培養(yǎng)基分離、純化植酸鹽水解圈與菌落直徑之比較大的菌株,得到M5-1(圖1a). M5-1在改良馬丁固體培養(yǎng)基上培養(yǎng),第2d長(zhǎng)出比較明顯的菌落,質(zhì)地絲絨狀,初生時(shí)為白色,菌落背面為淡黃色,呈放射狀.第3d少量菌絲頂端開始長(zhǎng)出分生孢子,孢子最開始為黃褐色,隨著培養(yǎng)時(shí)間增加,孢子顏色逐漸加深.第5d菌落直徑達(dá)5~6cm,整個(gè)菌落呈現(xiàn)黑褐色,菌落背面呈黃褐色.初期菌落形態(tài)正面和反面如圖1b、1c所示.

將M5-1的ITS序列在NCBI在線數(shù)據(jù)庫(kù)進(jìn)行Blast序列相似性比對(duì),選擇相關(guān)性較高的序列進(jìn)行分析,建立了如圖2所示的系統(tǒng)進(jìn)化樹.菌株M5-1與(MH185806.1)關(guān)系最緊密,位于同一分支中,相似度為99%.結(jié)合菌株M5-1的菌落特征,參考中國(guó)真菌志,可將菌株M5-1歸為[19].其分類學(xué)地位是子囊菌門,散囊菌綱,散囊菌目,曲霉科,曲霉屬,是一種好氧或兼性厭氧真菌,具有溶磷、釋鉀和固氮的功能[20]. GenBank序列號(hào)為MH978623.

圖2 基于ITS序列的菌株M5-1系統(tǒng)發(fā)育樹

2.3 Aspergillus tubingensis最適生長(zhǎng)pH值和對(duì)鈾的耐受性

環(huán)境pH值對(duì)微生物的增殖和活性會(huì)造成一定的影響.培養(yǎng)基初始pH值對(duì)菌絲干重的影響如圖3a所示,當(dāng)pH值為3~9時(shí),均有不同程度的生長(zhǎng).當(dāng)初始pH值為6~7時(shí),其菌絲干重顯著高于初始pH值為3,4,5,8,9的實(shí)驗(yàn)組(<0.05),說(shuō)明的最適生長(zhǎng)pH范圍為6~7.

不同小寫字母表示不同處理對(duì)菌絲干重影響差異顯著(<0.05)

鈾既是重金屬元素又是放射性元素,具有毒性,對(duì)微生物的生長(zhǎng)具有一定的影響,因此研究對(duì)鈾的耐受性十分重要.如圖3b所示,鈾濃度為0~0.84mmol/L時(shí),均能生長(zhǎng).鈾濃度為0.21mmol/L時(shí),的菌絲干重略高于對(duì)照組,其菌絲干重相比對(duì)照組只提高了5.41%;而其它鈾濃度實(shí)驗(yàn)組的菌絲干重相比對(duì)照組雖有所降低,但均無(wú)顯著差異.這些結(jié)果表明,對(duì)0~0.84mmol/L的鈾具有耐受性.

2.4 Aspergillus tubingensis水解植酸鹽效果

由圖4a可知,對(duì)照組(未接種)溶液pH值基本保持不變,維持在5左右.然而實(shí)驗(yàn)組(接種了),在0~2d, pH值從5迅速降至3.33,第15d,pH值降至最低值2.73.造成培養(yǎng)液pH值降低的原因可能是: 1)在生長(zhǎng)過(guò)程中分泌了有機(jī)酸[21-22]; 2)經(jīng)呼吸作用產(chǎn)生了CO2, CO2溶于水形成碳酸[23-24],導(dǎo)致pH值降低.15~62d,pH值逐漸上升,最終升至5.79,pH值上升主要與植酸鈉水解釋放正磷酸鹽有關(guān).

水解植酸鈉過(guò)程中正磷酸鹽濃度的變化如圖4b所示.第10d開始,實(shí)驗(yàn)組(接種了)開始產(chǎn)生正磷酸鹽,且正磷酸鹽濃度隨著時(shí)間的增加持續(xù)上升,這表明能有效水解植酸鹽釋放正磷酸鹽.

2.5 非生物沉淀

鈾濃度恒定為168 μmol/L時(shí),不同[Na12-IP6]:[U]物質(zhì)的量比下,溶液中水溶態(tài)鈾濃度及其所占比例如圖5所示.[Na12-IP6]:[U]￡5:1時(shí),溶液中水溶態(tài)鈾所占比例隨著[Na12-IP6]:[U]物質(zhì)的量比的增加而減少.然而[Na12-IP6]:[U]35:1時(shí),溶液中水溶態(tài)鈾所占比例隨著[Na12-IP6]:[U]物質(zhì)的量比的增加而增加.在1:2￡[Na12-IP6]:[U]￡15:1時(shí),溶液中的鈾主要以U-IP6形式沉淀.然而,當(dāng)[Na12-IP6]:[U]325:1時(shí),溶液中水溶態(tài)鈾所占比例增加,80%以上的鈾存在于溶液中.該現(xiàn)象與John等[25]的研究結(jié)果吻合.鑒于[Na12-IP6]: [U]325:1時(shí),80%以上的鈾存在于溶液中,本研究采用[Na12-IP6]:[U]=50:1進(jìn)行介導(dǎo)植酸鹽水解促進(jìn)U-PO43-礦化的實(shí) 驗(yàn).

圖5 溶液平衡7d后水溶態(tài)鈾的濃度及其所占比例

由圖4a可知,在水解植酸鹽的過(guò)程中,溶液pH值變化較大.為確定不同pH值下植酸鈉溶液中水溶態(tài)鈾的濃度及其所占比例,本文研究了在初始鈾濃度為168μmol/L,[Na12-IP6]: [U]= 50:1,溶液pH值分別為2,3,4,5,6,7的條件下,溶液平衡7d后,其中水溶態(tài)鈾的濃度及其所占的比例.試驗(yàn)結(jié)果如圖6所示.當(dāng)pH值分別為2,3,4,5,6,7時(shí),溶液中水溶態(tài)鈾的濃度和其所占比例相差甚微,說(shuō)明當(dāng)[Na12-IP6]:[U]=50:1時(shí),pH值對(duì)溶液中水溶態(tài)鈾的濃度及其所占比例影響很小.

圖6 不同pH值下溶液平衡7d后水溶態(tài)鈾的濃度及其所占比例

2.6 Aspergillus tubingensis介導(dǎo)植酸鹽水解促進(jìn)U-PO43-礦化過(guò)程中溶液pH值、正磷酸鹽濃度、鈾濃度和鈾去除率的變化

介導(dǎo)植酸鹽水解促進(jìn)U-PO43-礦化過(guò)程中溶液pH值隨時(shí)間的變化如圖7a所示.由圖可知:0~5d,A組pH值幾乎沒有變化,B組pH值從5迅速降至2.84.A組溶液pH值沒有變化的原因可能是在生長(zhǎng)初期對(duì)含鈾的培養(yǎng)基有一段調(diào)整期,生長(zhǎng)比較緩慢,釋放的有機(jī)酸較少[26].B組溶液中pH值降低的原因是,植酸鈉的存在,縮短了對(duì)含鈾培養(yǎng)基的調(diào)整期,并且促進(jìn)了的生長(zhǎng),它在生長(zhǎng)過(guò)程中分泌了有機(jī)酸,還有它的呼吸作用產(chǎn)生的CO2溶于水形成了碳酸[21-24]. 5~62d, A組pH值最終降至2.38,B組pH值持續(xù)上升至5.58.A組pH值快速降低的原因也是,經(jīng)過(guò)了調(diào)整期后生長(zhǎng)較快,分泌了大量的有機(jī)酸以及呼吸作用產(chǎn)生了較多的CO2[21-24].B組pH值上升可能與植酸鈉水解釋放正磷酸鹽有關(guān).

介導(dǎo)植酸鹽水解促進(jìn)U-PO43-礦化過(guò)程中正磷酸鹽濃度隨時(shí)間的變化如圖7b所示.第5d開始,B組開始產(chǎn)生正磷酸鹽,且隨著時(shí)間的增加正磷酸鹽濃度逐漸上升,最終達(dá)到19.80mmol/L.與水解植酸鹽實(shí)驗(yàn)中正磷酸鹽的變化情況相比,可發(fā)現(xiàn)鈾的存在刺激了正磷酸鹽的產(chǎn)生.

A:只添加鈾;B:添加鈾和植酸鈉

介導(dǎo)植酸鹽水解促進(jìn)U-PO43-礦化過(guò)程中鈾濃度和鈾的去除率隨時(shí)間的變化如圖7c所示.0~20d,A組和B組鈾濃度都出現(xiàn)波動(dòng).菌絲對(duì)鈾的吸附與溶液pH值兩者的耦合作用可能是導(dǎo)致A組鈾濃度出現(xiàn)波動(dòng)的主要原因;而B組鈾濃度出現(xiàn)波動(dòng)主要與溶液pH值、非生物沉淀和U-PO43-礦化三者的耦合作用有關(guān)[13,27-28].20~62d,A組鈾濃度上升至初始值,B組鈾濃度降至最低值,鈾的去除率最終達(dá)95.2%.A組鈾濃度回升至初始值的原因可能是溶液的pH值較低,吸附在表面的鈾被解吸[29].B組鈾濃度下降可能有如下兩方面的原因:1)溶液中的鈾可能被固定在了的菌絲上;2)溶液中釋放的正磷酸鹽與鈾結(jié)合可能形成了難溶于水的U-PO43-沉淀[13].

2.7 掃描電鏡/能譜分析

從圖8a可以看出,沉淀中不存在晶體,且圖8b中沒有出現(xiàn)U的信號(hào)峰,說(shuō)明A組實(shí)驗(yàn)沒有生成含U的礦物.然而從圖8c可以看出,沉淀中存在多面體晶體,且多面體晶體被包裹在菌絲體中間,表明鈾的礦化產(chǎn)物可在菌絲體表面形成[30].從圖8d可以看出,沉淀主要含U、P、Na、O等元素,且能譜中出現(xiàn)了較明顯的U、P信號(hào)峰,說(shuō)明介導(dǎo)U-PO43-礦化過(guò)程中可能形成了難溶的U-P礦物[31].

SEM圖比例尺:10μm;A:只添加鈾;B:添加鈾和植酸鈉

2.8 X射線衍射分析

介導(dǎo)植酸鹽水解促進(jìn)U- PO43-礦化過(guò)程中A、B 2組實(shí)驗(yàn)第62d沉淀的X射線衍射圖譜如圖9所示.A組的XRD圖顯示出較大的突起,說(shuō)明該組實(shí)驗(yàn)形成的沉淀是非晶形結(jié)構(gòu)[35].通過(guò)與XRD的PDF標(biāo)準(zhǔn)圖譜進(jìn)行比對(duì)發(fā)現(xiàn),B組形成的沉淀中可能含有氫鈾云母(H2(UO2)2(PO4)2·8H2O)和變鈉鈾云母(Na(UO2)(PO4)·3H2O)2種礦物.一些研究者也發(fā)現(xiàn)在微生物的作用下鈾可與PO43-結(jié)合形成氫鈾云母和變鈉鈾云母礦物[1,30-31,34].

圖9 Aspergillus tubingensis介導(dǎo)植酸鹽水解促進(jìn)U-PO43-礦化過(guò)程中A、B 2組第62d沉淀的XRD圖譜

A:只添加鈾;B:添加鈾和植酸鈉

3 結(jié)論

3.1 經(jīng)形態(tài)觀察和分子生物學(xué)鑒定,菌株M5-1為( MH978623).

3.2 菌株M5-1在pH值3~9范圍內(nèi)可以生長(zhǎng),其最適生長(zhǎng)pH值范圍為6~7.M5-1對(duì)鈾具有較強(qiáng)的耐受性,可耐受的鈾濃度高達(dá)0.84mmol/L.

3.3可以使植酸鹽水解釋放可溶性正磷酸鹽,從而將鈾原位固定,當(dāng)初始鈾濃度為168mmol/L時(shí),溶液中鈾的去除率達(dá)95.2%.

3.4 SEM-EDS和XRD分析表明,介導(dǎo)植酸鹽水解促進(jìn)U-PO43-礦化過(guò)程中可能形成了難溶的氫鈾云母和變鈉鈾云母礦物.

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Biomineralization of U(VI)-PO43-promoted bymediated phytate hydrolysis.

WANG Nie-ying1, ZHANG Hui1, SUI Yang1,2, CHEN Yue-yu1, HU Nan1, DAI Zhong-ran1, DING De-xin1*

(1.Key Discipline Laboratory for National Defense for Biotechnology in Uranium Mining and Hydrometallurgy, Hunan Province Key Laboratory of Green Development Technology for Extremely Low Grade Uranium Resources, University of South China, Hengyang 421001, China；2.School of Nuclear and Technology, University of South China, Hengyang 421001, China)., 2019,39(5)：2161~2169

The fungus M5-1capable of hydrolyzing phytate was isolated from the sediments of a uranium tailings repository in Guangdong Province. Its colony morphology, ITS sequences, suitable growth pH value, tolerance against uranium and effect on hydrolysis of phytate were systematically studied. The variations of pH value, orthophosphate concentration, uranium concentration and removal efficiency of uranium were monitored. The main elements and mineral components of the biomineralization products were also analyzed. It was found that the strain M5-1was(MH978623) with optimal growth at pH 6~7, andwith high tolerance against uranium (~0.84mmol/L). After biomineralization of U(VI)-PO43-promoted bymediated phytate hydrolysis for62days, the removal efficiency of uranium reached 95.2%. SEM-EDS and XRD analyses indicated that the insoluble chernikovite and metanatroautunite were formed during the biomineralization of U(VI)-PO43-The results showed thatcould effectively hydrolyze phytate to release soluble orthophosphate, which promoted the mineralization of U(VI)-PO43-. These results provided an experimental basis for the in-situ bioremediation of uranium contaminated surface water by biomineralization of U(VI)-PO43-promoted bymediated phytate hydrolysis.

uranium contaminated surface water；U(VI)；；biomineralization；phytate hydrolysis

X591

A

1000-6923(2019)05-2161-09

王聶穎(1993-),女,湖南常德人,南華大學(xué)碩士研究生,主要從事鈾污染水體生物修復(fù)技術(shù)研究.

2018-10-24

國(guó)家自然科學(xué)基金資助項(xiàng)目(U1401231,11775106)

*責(zé)任作者, 教授, dingdxzzz@163.com