陳婷婷,唐崇儉,鄭 平(浙江大學(xué)環(huán)境工程系,浙江 杭州 310029)

制藥廢水厭氧氨氧化脫氮性能與毒性機(jī)理的研究

陳婷婷,唐崇儉,鄭 平*(浙江大學(xué)環(huán)境工程系,浙江 杭州 310029)

采用上流式厭氧氨氧化污泥床反應(yīng)器考察了制藥廢水的生物脫氮性能,并采用發(fā)光細(xì)菌急性毒性試驗(yàn)研究了制藥廢水、厭氧氨氧化處理進(jìn)出水的生物毒性,以及制藥廢水對(duì)厭氧氨氧化污泥的蓄積毒性.結(jié)果表明,當(dāng)制藥廢水稀釋30倍以上時(shí),毒性物質(zhì)濃度低于毒性抑制濃度閾值,厭氧氨氧化反應(yīng)器運(yùn)行性能良好,平均氨氮和亞硝氮去除率分別達(dá)87.8%和95.6%,平均總氮容積負(fù)荷可達(dá)10.38 kg/(m3·d);但當(dāng)進(jìn)水稀釋小于20倍時(shí),毒性物質(zhì)濃度高于毒性抑制濃度閾值,反應(yīng)器運(yùn)行性能惡化,平均氨氮和亞硝氮去除率降至24.6%和26.0%,直到完全消失.制藥廢水、厭氧氨氧化反應(yīng)器進(jìn)出水均具有較強(qiáng)的生物毒性,在相對(duì)發(fā)光度為50%時(shí),所對(duì)應(yīng)的制藥廢水、反應(yīng)器進(jìn)水、出水的稀釋倍數(shù)分別為70.5,5.19,7.77倍.經(jīng)厭氧氨氧化處理后,出水毒性增強(qiáng),說明制藥廢水毒性物質(zhì)可在厭氧氨氧化污泥中蓄積,具有蓄積毒性.

厭氧氨氧化；制藥廢水；毒性機(jī)理

Abstract：In order to develop effective process to remove nitrogen from pharmaceutical wastewater, the performance of anaerobic ammonia oxidation (anammox) for nitrogen removal from pharmaceutical wastewater was investigated using upflow anaerobic sludge bed (UASB) reactor. The acute toxicity of pharmaceutical wastewater, the influent and effluent of anammox UASB reactor as well as the cumulative toxicity of pharmaceutical wastewater to anammox sludge were also studied. The reactor operation results indicated that when the pharmaceutical wastewater was diluted more than 30 times, the toxicant concentration below the threshold and the operation performance of anammox reactor was very good, the average removal rate of ammonia and nitrite reached 87.8% and 95.6% respectively, the average volumetric total nitrogen loading rate was up to 10.38 kg/(m3·d). However, when the pharmaceutical wastewater was diluted less than 20 times, the toxicant concentration higher than the threshold and the operation performance of anammox reactor deteriorated, the average removal rate of ammonia and nitrite reduced to 24.6% and 26.0%, and then completely disappeared. The acute toxicity by photobacteria tests showed that the pharmaceutical wastewater, the influent and effluent of anammox process all had intense biotoxicity. When the relative luminosity was 50%, the dilution factors of the three samples were 70.5, 5.19, 7.77 respectively. After anammox treatment, the toxicity of effluent enhanced. The cumulative toxicity by photobacteria tests suggested that the toxicants in pharmaceutical wastewater could cumulate in anammox sludge, leading to higher toxicity in the test Anammox sludge than that in the seed sludge.

Key words：anammox process；pharmaceutical wastewater；toxic mechanism

厭氧氨氧化是以氨為電子供體,亞硝酸鹽為電子受體的生物反應(yīng),最早發(fā)現(xiàn)于流化床反應(yīng)器中[1].厭氧氨氧化工藝具有總氮容積負(fù)荷高以及經(jīng)濟(jì)性好的優(yōu)點(diǎn)[2],目前相關(guān)研究主要集中在反應(yīng)器啟動(dòng)[3-4],工藝性能[5-6],富集培養(yǎng)[7-8],分子生物學(xué)[9]等方面,有關(guān)厭氧氨氧化工藝處理實(shí)際廢水的研究,迄今少見報(bào)道.

制藥廢水COD、鹽度、氨氮含量均很高,成分復(fù)雜并含毒性物質(zhì),屬于難處理工業(yè)廢水[10],如何處理該類廢水是當(dāng)今環(huán)境保護(hù)的一個(gè)難題.眾多學(xué)者已研究證實(shí),制藥廢水對(duì)生物具有急性毒性作用[11-13],但對(duì)蓄積毒性的研究報(bào)道較少[14-15].Carucci等[16]研究表明,制藥廢水中的某些組分,如林肯霉素,會(huì)對(duì)傳統(tǒng)脫氮技術(shù)產(chǎn)生抑制作用.為此,本研究采用實(shí)驗(yàn)室裝置試驗(yàn)了厭氧氨氧化工藝處理制藥廢水的性能,并采用發(fā)光細(xì)菌急性毒性試驗(yàn)研究了制藥廢水的急性毒性和蓄積毒性.

1 材料與方法

1.1材料

試驗(yàn)用水取自浙江省某制藥公司,其水質(zhì)指標(biāo)為:pH 9.30~9.72,NH4+-N濃度為5000~ 5500mg/L,COD為2000~4000mg/L,NO2--N 0.82~ 1.03mg/L, NO3--N 8.50~10.06mg/L,廢水COD/ NH4+-N為0.4~0.8.

毒性試驗(yàn)菌種為明亮發(fā)光桿菌(Photobacterium phosphoreum)T3變種,購(gòu)自中國(guó)科學(xué)院南京土壤研究所.培養(yǎng)液及培養(yǎng)基配方參照文獻(xiàn)[17].菌劑復(fù)蘇方法如下:將裝有凍干粉的安培瓶從4℃冰箱取出,打開瓶頸,用1mL注射器注入2%NaCl,置于漩渦振蕩器上充分混勻,2min后菌劑即復(fù)蘇發(fā)光,作為試驗(yàn)菌劑備用.

毒性試驗(yàn)污泥為用于反應(yīng)器接種的厭氧氨氧化污泥及處理制藥廢水后的厭氧氨氧化污泥.

1.2儀器

DXY-2型生物毒性(污染)測(cè)試儀(中國(guó)科學(xué)院南京土壤所);YXQ-SG41-280型手提式壓力蒸汽滅菌鍋(上海華線醫(yī)用核子儀器有限公司); FE20型pH計(jì)(梅特勒-托利多儀器(上海)有限公司);85-2型磁力攪拌器(杭州儀表電機(jī)廠);微量注射器(10μL)(上海光正醫(yī)療儀器有限公司); Beckman高速冷凍離心機(jī).

1.3試驗(yàn)裝置與運(yùn)行

上流式厭氧污泥床(UASB)反應(yīng)器如圖1所示.由有機(jī)玻璃制成,內(nèi)徑50mm,反應(yīng)區(qū)有效容積為1.1L,外裹黑布,以防光照的負(fù)面影響.廢水通過蠕動(dòng)泵連續(xù)從底部泵入反應(yīng)器,氣體從反應(yīng)器頂部氣室引出,凈化水從反應(yīng)器上部溢流堰排放.進(jìn)水采用氬氣除氧30min,以確保缺氧狀態(tài),操作溫度控制在(35±1)℃.在進(jìn)行本試驗(yàn)前,該厭氧氨氧化反應(yīng)器已采用模擬含氨廢水穩(wěn)定運(yùn)行數(shù)月,總氮容積去除速率為25.04kg N/(m3·d)[18],反應(yīng)器中平均污泥濃度(以VSS計(jì))為21.1g/L.

圖1 厭氧氨氧化UASB反應(yīng)器示意Fig.1 Schematic drawing of the anammox UASB reactor

考慮到制藥廢水的生物毒性和廢水中的有機(jī)物濃度,采用逐漸縮小稀釋倍數(shù)的方法考察厭氧氨氧化工藝的脫氮性能.在整個(gè)試驗(yàn)過程中,向進(jìn)水中添加亞硝酸鈉,控制進(jìn)水亞硝酸鹽濃度為240~300mg/L,以避免亞硝酸鹽自身對(duì)厭氧氨氧化的抑制作用[18].根據(jù)厭氧氨氧化工藝的化學(xué)計(jì)量關(guān)系,在稀釋倍數(shù)較大時(shí)(制藥廢水中氨氮濃度不足),向廢水中添加硫酸銨,以保證厭氧氨氧化所需的NH4+-N濃度;另外,按文獻(xiàn)[18]的配方,在進(jìn)水中添加營(yíng)養(yǎng)元素和微量元素.每個(gè)稀釋倍數(shù)下穩(wěn)定運(yùn)行1周,待反應(yīng)器性能穩(wěn)定后再進(jìn)一步縮小稀釋倍數(shù).

1.4測(cè)定方法

1.4.1水樣的發(fā)光細(xì)菌急性毒性 以采集的制藥廢水及厭氧氨氧化反應(yīng)器運(yùn)行至第43d的進(jìn)出水進(jìn)行試驗(yàn),控制溫度為20~25℃,原水pH值調(diào)節(jié)至7.3,反應(yīng)器進(jìn)出水pH值不加調(diào)節(jié).將待測(cè)水樣用3% NaCl稀釋配制成5個(gè)濃度梯度,每個(gè)濃度設(shè)3組平行,以2mL 3%NaCl作為空白對(duì)照.取10μL試驗(yàn)菌劑于各玻璃管中,振蕩混勻,15min后用生物毒性測(cè)試儀測(cè)定發(fā)光強(qiáng)度.通過空白對(duì)照發(fā)光度(相對(duì)發(fā)光單位RLU表示)的平均值RLU0和各濃度3組平行樣發(fā)光度的平均值RLU,計(jì)算發(fā)光細(xì)菌的相對(duì)發(fā)光度X(%),即:

用直線內(nèi)插法求出X為50%時(shí)所對(duì)應(yīng)的溶液稀釋倍數(shù).

1.4.2發(fā)光細(xì)菌蓄積毒性 分別稱取接種污泥及處理制藥廢水后的厭氧氨氧化污泥2g,通過勻漿、過濾(孔徑1~3μm)、10000r/min離心之后,取上清液用3% NaCl定容至50mL.用3% NaCl稀釋配制成5個(gè)濃度梯度,每個(gè)濃度設(shè)3組平行,以2mL 3% NaCl做為空白對(duì)照.測(cè)定及計(jì)算方法同急性毒性試驗(yàn),由于提取液有色,測(cè)量時(shí)進(jìn)行色度校正[19].

1.4.3其他指標(biāo) 氨氮采用水楊酸-次氯酸鹽光度法測(cè)定;亞硝氮采用N-(1-萘基)-乙二胺光度法測(cè)定;硝氮采用紫外分光光度法;VSS采用重量法測(cè)定[20];pH值采用FE20型pH計(jì)測(cè)定;COD采用COD儀測(cè)定.

1.5數(shù)據(jù)處理及分析方法

利用Originlab 7.5進(jìn)行數(shù)據(jù)統(tǒng)計(jì)分析.采用重鉻酸鉀法測(cè)定COD時(shí),NO2--N可被重鉻酸鉀氧化而使測(cè)定值比實(shí)際值偏高,因此廢水中實(shí)際COD應(yīng)扣除由NO2--N所致的COD.在廢水COD計(jì)算中,采用下式扣除亞硝氮的影響[21-22]:

2 結(jié)果與討論

2.1反應(yīng)器運(yùn)行性能

維持反應(yīng)器進(jìn)水NH4+-N、NO2--N濃度為180~300mg/L,控制HRT為0.96h,將稀釋倍數(shù)由60倍逐漸降低為20倍.在整個(gè)運(yùn)行過程中,反應(yīng)器的進(jìn)水pH值控制為6.8~7.3;出水pH值穩(wěn)定在8.0~8.36.由圖2可知,當(dāng)進(jìn)水稀釋倍數(shù)為30倍以上時(shí),反應(yīng)器平均出水NH4+-N、NO2--N和NO3--N濃度分別為27.4,11.2,25.4mg/L,平均NH4+-N和NO2--N去除率分別達(dá)87.8%和95.6%,總氮容積去除速率可達(dá)10.38kg N/(m3·d);但當(dāng)進(jìn)水稀釋倍數(shù)降低至20倍以下時(shí),反應(yīng)器性能降低,出水NH4+-N、NO2--N分別達(dá)192.5, 114.2mg/L,平均NH4+-N和NO2--N去除率降低至24.6%和26.0%.運(yùn)行到27d,出水NH4+-N濃度高于進(jìn)水濃度,反應(yīng)器的厭氧氨氧化功能消失.鑒于此,第28,29d,連續(xù)用清水沖洗反應(yīng)器,待出水NH4+-N和NO2--N濃度降低至5mg/L以下后,將HRT延長(zhǎng)到16h,繼續(xù)以稀釋倍數(shù)為20倍的進(jìn)水運(yùn)行.反應(yīng)器的運(yùn)行性能不佳,至40d,反應(yīng)器的出水NH4+-N和NO2--N濃度再次高于進(jìn)水濃度.

當(dāng)進(jìn)水稀釋倍數(shù)為30倍以上時(shí),COD平均去除率為49.0%;當(dāng)進(jìn)水稀釋倍數(shù)降低至20倍以下時(shí),隨著運(yùn)行時(shí)間的延長(zhǎng),COD的去除也逐漸變差,至試驗(yàn)?zāi)┢?出水COD與進(jìn)水持平,有時(shí)甚至高于進(jìn)水(圖3).

由此可見,當(dāng)進(jìn)水稀釋倍數(shù)為30倍以上時(shí),毒性效應(yīng)不顯著,說明在稀釋倍數(shù)較大時(shí),廢水中毒性物質(zhì)濃度低于可產(chǎn)生毒性作用的下限值(即毒性抑制濃度閾值),此時(shí)反應(yīng)器具有較高的總氮容積負(fù)荷、NH4+-N、NO2--N去除率及一定的COD去除率;但當(dāng)進(jìn)水稀釋倍數(shù)降低至20倍以下時(shí),毒性效應(yīng)顯現(xiàn),說明隨著稀釋倍數(shù)的降低,毒性物質(zhì)濃度增加且高于毒性抑制濃度閾值,此時(shí)反應(yīng)器性能惡化,總氮容積負(fù)荷以及NH4+-N、NO2--N、COD去除率降低且急劇下降,直到完全消失.出現(xiàn)毒性效應(yīng)后,延長(zhǎng)HRT、降低負(fù)荷也無法改變反應(yīng)器性能惡化的趨勢(shì).值得關(guān)注的是,隨著運(yùn)行時(shí)間的增加,恢復(fù)運(yùn)行后厭氧氨氧化功能因毒性效應(yīng)消失的速度加快,表現(xiàn)出蓄積毒性.

2.2制藥廢水的急性毒性

由圖4可知,發(fā)光細(xì)菌的發(fā)光強(qiáng)度與廢水稀釋倍數(shù)有較好的線性關(guān)系,相關(guān)系數(shù)為0.9237~ 0.9963.依據(jù)回歸方程計(jì)算,在相對(duì)發(fā)光度為50%時(shí),對(duì)應(yīng)的制藥廢水、厭氧氨氧化反應(yīng)器進(jìn)出水的稀釋倍數(shù)分別為70.5,5.19,7.77倍,出水的生物毒性高于進(jìn)水,值得深入研究.

由此可見,制藥廢水、厭氧氨氧化反應(yīng)器進(jìn)出水均具有較強(qiáng)的生物毒性.若不調(diào)節(jié)pH值,直接進(jìn)行制藥廢水的生物毒性試驗(yàn),即使稀釋40~200倍,相對(duì)發(fā)光度均為0,顯現(xiàn)出很強(qiáng)的生物毒性,這可歸因于高pH值(9.72)和制藥廢水中的毒性物質(zhì)的聯(lián)合作用,在高pH值下,氨主要以游離態(tài)形式存在,游離氨對(duì)微生物具有毒性作用[3].

圖2 厭氧氨氧化工藝處理制藥廢水運(yùn)行結(jié)果Fig.2 The operation results of pharmaceutical wastewater treated by anammox process

2.3蓄積毒性

以接種污泥(未處理制藥廢水的污泥)作對(duì)照,考察了毒性物質(zhì)在厭氧氨氧化污泥中的蓄積毒性.由圖5可見,在相同的稀釋倍數(shù)下,試驗(yàn)組的相對(duì)發(fā)光度明顯低于對(duì)照組,具有蓄積毒性.這一結(jié)果與反應(yīng)器運(yùn)行結(jié)果相吻合.

從污泥自身性狀觀察,對(duì)照組偏紅色,試驗(yàn)組偏棕黃色.厭氧氨氧化菌富含血紅素[23-24],使污泥呈紅色,而效能惡化時(shí),污泥顏色變?yōu)樽攸S色.從 COD去除的變化趨勢(shì)(圖3)可知,隨著運(yùn)行時(shí)間的延長(zhǎng)及稀釋倍數(shù)的降低,COD去除率呈下降趨勢(shì),至試驗(yàn)?zāi)┢?出水COD高于進(jìn)水,這可能是由于廢水毒性導(dǎo)致菌體水解釋放出體內(nèi)物質(zhì),這也印證了制藥廢水的蓄積毒性.張蕾等[25]研究證實(shí),厭氧氨氧化菌富集培養(yǎng)物性狀黏稠,含有較多的胞外聚合物(EPS).EPS表面帶有許多負(fù)電荷基團(tuán)如羧基、磷?；土蛩岣?具有較強(qiáng)的吸附能力[26].而且,制藥廢水中成分復(fù)雜,其中包括一些不易生物降解的物質(zhì),而在進(jìn)行厭氧氨氧化處理時(shí),DO、pH值等因素發(fā)生變化,可能造成它們沉淀、富集在菌體上,各組分之間也可能相互發(fā)生化學(xué)反應(yīng)或經(jīng)生物轉(zhuǎn)化后生成毒性更強(qiáng)的物質(zhì),使之產(chǎn)生蓄積毒性效應(yīng)[27].

圖3 反應(yīng)器進(jìn)出水CODFig.3 COD concentration in the influent and effluent of the anammox reactor

圖4 廢水對(duì)發(fā)光細(xì)菌的急性毒性Fig.4 The acute toxicity of wastewater to photobacteria

制藥廢水中毒性物質(zhì)在厭氧氨氧化污泥中蓄積所產(chǎn)生的蓄積毒性,可能與菌體水解釋放體內(nèi)蓄積的毒性物質(zhì),EPS對(duì)制藥廢水中毒性物質(zhì)的吸附,制藥廢水中毒性物質(zhì)在菌體表面沉淀、富集,以及各組分之間復(fù)雜的生物化學(xué)反應(yīng)有關(guān).目前,對(duì)制藥廢水中各組分之間的生物化學(xué)反應(yīng)知之甚少,有待于進(jìn)一步研究.

圖5 厭氧氨氧化污泥的蓄積毒性Fig.5 The cumulative toxicity of anammox sludge

3 結(jié)論

3.1超過毒性抑制濃度閾值后,制藥廢水對(duì)厭氧氨氧化反應(yīng)器運(yùn)行性能具有顯著的毒性作用,隨著運(yùn)行時(shí)間的延長(zhǎng),這種毒性作用有加重的趨勢(shì).要使厭氧氨氧化反應(yīng)器正常工作,需將進(jìn)水中的毒性物質(zhì)濃度控制在閾值以下,對(duì)于供試廢水,宜將廢水稀釋30倍以上.

3.2發(fā)光細(xì)菌毒性試驗(yàn)表明,制藥廢水、厭氧氨氧化反應(yīng)器進(jìn)出水均具有較強(qiáng)的生物毒性,在相對(duì)發(fā)光度為50%時(shí),三者的稀釋倍數(shù)分別為70.5,5.19,7.77倍.經(jīng)生物處理后,出水毒性增強(qiáng).3.3由于制藥廢水中毒性物質(zhì)在厭氧氨氧化污泥中蓄積所產(chǎn)生的蓄積毒性以及生物處理對(duì)制藥廢水的毒性增強(qiáng)效應(yīng),采用厭氧氨氧化工藝進(jìn)行制藥廢水脫氮處理時(shí),宜持慎重態(tài)度.

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Nitrogen removal performance of anammox process and toxic mechanism of pharmaceutical wastewater.

CHEN Ting-ting, TANG Chong-jian, ZHENG Ping*(Department of Environmental Engineering, Zhejiang University, Hangzhou 310029, China). China Environmental Science, 2010,30(4)：504~509

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A

1000-6923(2010)04-0504-06

陳婷婷(1986-),女,四川廣安人,浙江大學(xué)環(huán)境工程系碩士研究生,研究方向?yàn)榄h(huán)境微生物與廢物生物處理.發(fā)表論文2篇.

2009-08-17

國(guó)家自然科學(xué)基金資助項(xiàng)目(30770039);國(guó)家“863”項(xiàng)目(2006AA06Z332);浙江省基金項(xiàng)目(Y507227)

* 責(zé)任作者, 教授, pzheng@zju.edu.cn