CN105821082B - 假交替单胞菌在制备纳米材料中的应用 - Google Patents
假交替单胞菌在制备纳米材料中的应用 Download PDFInfo
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- CN105821082B CN105821082B CN201610232204.7A CN201610232204A CN105821082B CN 105821082 B CN105821082 B CN 105821082B CN 201610232204 A CN201610232204 A CN 201610232204A CN 105821082 B CN105821082 B CN 105821082B
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- 241000519590 Pseudoalteromonas Species 0.000 title claims abstract description 20
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- NRDMFTSWEBBZFK-UHFFFAOYSA-N [P].[Cd] Chemical compound [P].[Cd] NRDMFTSWEBBZFK-UHFFFAOYSA-N 0.000 claims abstract description 24
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims abstract description 18
- QSOCBFRYIZAAJU-UHFFFAOYSA-N [S].[P].[Cd] Chemical compound [S].[P].[Cd] QSOCBFRYIZAAJU-UHFFFAOYSA-N 0.000 claims abstract description 16
- 241000519582 Pseudoalteromonas sp. Species 0.000 claims abstract description 12
- 150000003839 salts Chemical class 0.000 claims description 21
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- 229910052793 cadmium Inorganic materials 0.000 abstract description 33
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- BDOSMKKIYDKNTQ-UHFFFAOYSA-N cadmium atom Chemical compound [Cd] BDOSMKKIYDKNTQ-UHFFFAOYSA-N 0.000 abstract description 30
- 229910052698 phosphorus Inorganic materials 0.000 abstract description 26
- OAICVXFJPJFONN-UHFFFAOYSA-N Phosphorus Chemical compound [P] OAICVXFJPJFONN-UHFFFAOYSA-N 0.000 abstract description 23
- 239000011574 phosphorus Substances 0.000 abstract description 23
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- NLXLAEXVIDQMFP-UHFFFAOYSA-N Ammonia chloride Chemical compound [NH4+].[Cl-] NLXLAEXVIDQMFP-UHFFFAOYSA-N 0.000 description 2
- NBIIXXVUZAFLBC-UHFFFAOYSA-N Phosphoric acid Chemical compound OP(O)(O)=O NBIIXXVUZAFLBC-UHFFFAOYSA-N 0.000 description 2
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- QMIDDQFZXOSYIL-UHFFFAOYSA-K [Na+].[Na+].[Na+].[O-]P([O-])([O-])=O.OCC(O)CO Chemical compound [Na+].[Na+].[Na+].[O-]P([O-])([O-])=O.OCC(O)CO QMIDDQFZXOSYIL-UHFFFAOYSA-K 0.000 description 1
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- 229910001385 heavy metal Inorganic materials 0.000 description 1
- BHEPBYXIRTUNPN-UHFFFAOYSA-N hydridophosphorus(.) (triplet) Chemical compound [PH] BHEPBYXIRTUNPN-UHFFFAOYSA-N 0.000 description 1
- 238000007689 inspection Methods 0.000 description 1
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- 238000005551 mechanical alloying Methods 0.000 description 1
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- 229910052976 metal sulfide Inorganic materials 0.000 description 1
- MEFBJEMVZONFCJ-UHFFFAOYSA-N molybdate Chemical compound [O-][Mo]([O-])(=O)=O MEFBJEMVZONFCJ-UHFFFAOYSA-N 0.000 description 1
- 239000002077 nanosphere Substances 0.000 description 1
- 229910000510 noble metal Inorganic materials 0.000 description 1
- 230000001590 oxidative effect Effects 0.000 description 1
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- 239000011734 sodium Substances 0.000 description 1
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- 229960002901 sodium glycerophosphate Drugs 0.000 description 1
- REULQIKBNNDNDX-UHFFFAOYSA-M sodium;2,3-dihydroxypropyl hydrogen phosphate Chemical compound [Na+].OCC(O)COP(O)([O-])=O REULQIKBNNDNDX-UHFFFAOYSA-M 0.000 description 1
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Abstract
本发明公开了假交替单胞菌(Pseudoalteromonas sp.)DSBS在镉磷的nM级不饱和高盐废水中制备纳米材料中的应用;所述假交替单胞菌DSBS已于2013年12月12日保藏于中国典型培养物保藏中心,地址:武汉市武昌珞珈山,保藏号:CCTCC M2013652。本发明还公开了一种镉‑磷‑硫纳米材料,由假交替单胞菌DSBS在镉磷的nM级不饱和高盐废水中生物合成。本发明的菌株能够在不同盐度、镉磷不饱和体系中合成纳米颗粒,并且实现水体中的镉和磷的去除。底物利用率高,出水无镉磷污染。
Description
技术领域
本发明涉及假交替单胞菌在制备纳米材料中的应用。
背景技术
纳米材料有独特的物理特征和化学性质,可用于传感器、催化、半导体、药物输送、污染物降解和环境修复等领域,近年来受到学术和工程界的广泛关注。
目前,金属纳米材料的合成主要有物理、化学和生物方法。物理方法包括蒸发冷凝、物理粉碎、机械合金化,但其产出率低、能耗高、成本大。化学方法包括化学沉淀、水热合成、模板法等。但其合成的纳米颗粒不易控制大小、形状和结构;需要高温高压、特定pH、高浓度的过饱和溶液体系等严苛条件;反应底物的利用率底;会产生很多污染物质,如化学前驱、有毒溶剂和有害副产物等。
相比较而言,生物合成纳米材料更加环境友好,合成条件温和。细菌可以还原形成贵金属纳米颗粒如Ti纳米球,合成非金属纳米材料如单质Se,氧化金属离子形成纳米氧化物如UO2,产生硫负离子形成金属硫化物晶体。但目前,对于细菌合成金属磷酸盐纳米材料的报道较少,特别是有生物毒性的金属盐,如镉与磷结合的纳米材料。
镉的许多纳米材料具有重要的应用价值。Cd(OH)2可以作为重要的能量材料应用于太阳能电池,CdS、CdSe可作为电极添加物,CdS纳米颗粒和纳米薄膜具有很高的光电转换效率和光催化性能,是重要的半导体材料,CdSeS可用于生物荧光标记,多种含镉纳米颗粒都可作为水处理吸附剂。生物合成镉的纳米材料将会有很好的工程前景。
生物合成纳米材料技术虽然环境友好,但在培养条件方面往往要求有较高浓度的金属离子或阴离子作为反应底物。据报道,Serratia sp.在胞外聚合物中能形成钙-磷纳米颗粒,要求钙离子浓度1mM,磷源为单一甘油磷酸钠并且浓度高达5mM。在这种mM级的阳离子浓度中,阴离子只要达到0.84μM就会产生自发的化学沉淀。这意味着只要有0.017%的甘油磷酸钠被水解而释放正磷酸根,钙-磷化学沉淀就会发生,这使得生物合成纳米颗粒非常容易。但同时会导致反应底物的利用率低,反应完全后溶液仍有μM级的不饱和底物难以继续沉淀,不仅造成原材料的浪费,还造成潜在环境危害。因此,在不饱和金属阳离子和阴离子的溶液中合成纳米材料的技术具有重要意义。
在许多废水中,都含有可合成镉磷纳米颗粒的原材料。如化工废水中镉浓度有2.4-250mg/L,含磷6-8mg/L。由于海水中固有的磷镉离子,海水利用废水中也存在一定浓度的镉和磷。这些废水中镉磷都是不饱和体系,同时存在镉磷去除的需要。目前,废水中镉磷的生物处理手段主要依赖吸附和厌氧除磷,过程复杂,需要控制曝气条件和污泥龄。在高盐的海水利用废水处理方面,主要采用驯化耐盐活性污泥方法,但耗时长、运行费用高,化学沉淀为主的高效沉淀生物过滤工艺,会消耗大量化学药剂并产生化学污泥。如果将废水处理和生物合成纳米材料相结合,提出一种既可以在不同盐度废水中去除镉磷,又可以利用废水中的镉磷合成纳米材料的生物技术,将会更好的实现生物合成纳米材料的优势,完成废水处理和资源的高效回收。
发明内容
针对上述现有技术,本发明的目的是提供一种假交替单胞菌在制备纳米材料中的应用。
本发明是基于发明人的前期研究,以保藏在中国典型培养物保藏中心的假交替单胞菌菌株(保藏编号:CCTCC M2013652)作为研究的基础,进行了一系列的技术扩展研究。发现该菌株不仅能够高效去除水体中的镉和磷,还能够在μM级和nM级镉磷不饱和的低盐和高盐废水中生长,在胞外形成直径10-60nm的镉-磷-硫纳米颗粒。
为实现上述目的,本发明采用下述技术方案:
根据本发明的第一方面,提供假交替单胞菌(Pseudoalteromonas sp.)DSBS在制备纳米材料中的应用。
上述假交替单胞菌(Pseudoalteromonas sp.)DSBS已于2013年12月12日保藏于中国典型培养物保藏中心,地址:武汉市武昌珞珈山,保藏号:CCTCC M2013652。记载在申请人的另一项专利申请“一株高效去除废水中镉和磷的假交替单胞菌及其应用”(CN103740623B)中。
根据本发明的第二方面,提供一种镉-磷-硫纳米材料,由假交替单胞菌(Pseudoalteromonas sp.)DSBS在含镉和磷的水体中生物合成。
所述含镉和磷的水体中,镉和磷的浓度为μM级至nM级,是镉磷的不饱和体系。
根据本发明的第三方面,提供上述镉-磷-硫纳米材料的制备方法,包括假交替单胞菌(Pseudoalteromonas sp.)DSBS菌体富集的步骤,以及将富集的菌体在含镉和磷的水体中进行生物合成的步骤。
上述制备方法中,所述菌体富集的步骤包括:将假交替单胞菌(Pseudoalteromonas sp.)DSBS接种于液体LB培养基中,180-220rpm,20-30℃,培养16-24h;优选的,200rpm,25℃,培养20h。
所述液体LB培养基的配方为:蛋白胨10g/L,酵母粉3g/L,陈海水配制,海水盐度为3.5%。
上述制备方法中,所述生物合成的步骤包括:将富集的菌体接种于含镉和磷的水体中,180-220rpm,15-30℃,培养42-54h;优选的,200rpm,25℃,培养48h。
所述富集的菌体的接种量为8-12%(v/v);优选的,10%(v/v)。
上述生物合成的步骤,还包括:将培养后的菌体用去离子水离心洗涤,优选的,3000rpm离心。
本发明中,所述含镉和磷的水体可以为低盐废水或高盐废水。
上文中,所述高盐废水是指总盐质量分数(即盐度)在1%及以上的水体,低盐废水是指总盐质量分数(即盐度)在1%以下的水体。在本发明的一个具体实施方式中,所述低盐废水的盐度为0.35%;在本发明的一个具体实施方式中,所述高盐废水的盐度为3.5%。
上述镉-磷-硫纳米材料在去除水体中污染物中的应用,或者在制备半导体功能材料,或者在制备传感器、太阳能电池的窗口材料、光催化剂、发光二极管中的应用也属于本发明的保护范围。
所述水体中的污染物包括磷、氟、重金属和放射性废物。
本发明的有益效果:
(1)本发明所述菌株能够在完成废水磷镉去除的同时生物合成镉-磷-硫纳米颗粒。出水无磷镉污染,并且纳米材料的合成实现了资源的高效回收利用。
(2)本发明所述菌株合成纳米颗粒的环境条件温和。溶解氧、pH和温度等培养条件完全模拟好氧污水处理过程,操作简单,成本较低。
(3)本发明所述菌株能够在磷镉不饱和溶液中合成纳米颗粒。从含镉废水中μM级的镉浓度,到海水中nM级的镉浓度,该菌株皆可合成纳米颗粒,在金属离子的浓度要求上很宽泛,适应性强,节约原料,降低了成本。
(4)本发明所述菌株能够在低盐和高盐条件下合成纳米颗粒,盐度适应性强。
(5)本发明所述菌株在废水中合成的纳米材料具有多种形貌和粒径。纳米颗粒有三种形态:均匀分散在菌体表面、球形聚集和成堆聚集。μM级镉浓度纳米材料粒径25-60nm,nM级则在10nm左右。
(6)本发明所述菌株合成的纳米颗粒为夹杂胞外多糖的镉-磷-硫纳米颗粒,镉在胞外的沉淀降低了其对细菌的毒性,提高菌体抗镉性能。
附图说明
图1:本发明的菌株在低盐废水中形成纳米颗粒的原子力显微镜图像(A)、扫描电镜图像(B、C、D)和能谱分析(E);
图2:本发明的菌株在高盐废水中的磷去除情况;
图3:本发明的菌株在高盐废水中形成纳米颗粒的原子力显微镜图像(A)、扫描电镜图像(B)和能谱分析(C)。
具体实施方式
下述实施例中所涉及的仪器、试剂、材料等,若无特别说明,均为现有技术中已有的常规仪器、试剂、材料等,可通过正规商业途径获得。下述实施例中所涉及的实验方法,检测方法等,若无特别说明,均为现有技术中已有的常规实验方法,检测方法等。
实施例1:假交替单胞菌(Pseudoalteromonas sp.)DSBS在低盐废水中制备纳米材料
(1)将假交替单胞菌(Pseudoalteromonas sp.)DSBS按0.9%(v/v)接种于液体LB培养基中,在25℃、200rpm恒温摇床中培养20h,得到富集的菌体。
(2)将富集的菌悬液按10%(v/v)接种于低盐废水中,在25℃、200rpm恒温摇床中培养48h,即得到含有镉-磷-硫纳米颗粒的菌悬液。
(3)将含有镉-磷-硫纳米颗粒的菌悬液用去离子水离心洗涤。离心速度为3000rpm,采用低速离心防止菌体表面颗粒物洗脱,制备得到纳米材料。
所述的液体LB培养基组分如下:
蛋白胨10g/L,酵母粉3g/L,陈海水配制,海水盐度为3.5%。
所述的低盐废水组分如下:
一水合葡萄糖5.06g/L,NaAC 1.5g/L,NaCl 3.5g/L,NH4Cl 2.6g/L,MgSO4·7H2O2.4g/L,总镉(Cd(NO3)2·4H2O)和总磷(K2HPO4)浓度分别为8mg/L和9mg/L,初始pH 7.2,盐度0.35%。为模拟低盐废水环境,其中镉和磷浓度为一般工业含镉废水中的浓度,在该pH和离子浓度下,不足以形成无机化学镉沉淀,属于镉磷的μM级不饱和低盐环境。
为进一步表征菌体及其形成的纳米颗粒,使用原子力显微镜和扫描电镜观察去除磷镉后的菌体。原子力显微镜观察结果如图1中A所示,菌体表面均匀聚集着纳米颗粒物质,少数颗粒脱落分散在四周,颗粒直径为25-100nm。图中部分菌体有鞭毛,部分没有,可能是在培养或样品预处理过程中脱落。
扫描电镜观察菌体的预处理过程中,磷酸缓冲液洗涤的离心速度为3000rpm,采用低速离心防止菌体表面颗粒物洗脱,其他离心速度为6000rpm,结果如图1中B、C、D所示。图1B中,直径在25-60nm的较小颗粒聚集成直径为100nm的均匀球形,粘附在菌体表面。图1C中,较小颗粒聚集在胞外多糖的纤维上。图1D中,较小颗粒成堆聚集在菌体之间。这说明细菌合成的纳米颗粒有三种不同的形态:均匀分散在菌体表面的直径25-60nm的较小颗粒,粘附在菌体表面的直径100nm的较小颗粒球形聚集体,聚集体脱离细菌成堆粘附在菌体之间的多糖纤维上。同时对图中点位的能谱分析表明,颗粒主要含有C、O、Cd、P、S元素。这说明该纳米颗粒是夹杂多糖的镉-磷-硫纳米颗粒。其中Na是析出的可溶性离子,Al是样品台元素,均不是纳米颗粒中的元素。
因此,该菌在磷镉浓度不饱和的低盐废水中可实现磷镉的同时去除,并在胞外形成具有三种形态的夹杂多糖的镉-磷-硫纳米颗粒,直径为25-60nm。
实施例2:假交替单胞菌(Pseudoalteromonas sp.)DSBS在高盐废水中制备纳米材料
(1)将假交替单胞菌(Pseudoalteromonas sp.)DSBS按0.9%(v/v)接种于液体LB培养基中,在25℃、200rpm恒温摇床中培养20h,得到富集的菌体。
(2)将富集的菌体3000rpm离心10min,弃去上清液,再用高盐废水重悬至原体积,得到洗涤一次的菌体。
(3)将洗涤后的菌悬液按0.2‰(v/v)接种于三组高盐废水中,在25℃、200rpm恒温摇床中培养48h,即得到含有镉-磷-硫纳米颗粒的菌悬液。
(4)将含有镉-磷-硫纳米颗粒的菌悬液用去离子水洗涤,离心速度为3000rpm,采用低速离心防止菌体表面颗粒物洗脱,制备得到纳米材料。
根据本发明优选的,所述的液体LB培养基组分如下:
蛋白胨10g/L,酵母粉3g/L,陈海水配制,海水盐度为3.5%。
所述的高盐废水组分如下:
NaAC 0.82g/L,NH4Cl 0.11g/L,海水素33.33g/L,初始pH 7.2,盐度3.5%。三组合成海水的总镉(Cd(NO3)2·4H2O)和总磷(K2HPO4)浓度有差异,分别为A组0.1756×10-3mg/L和0.1548mg/L,B组5.671×10-3mg/L和5mg/L,C组10.21×10-3mg/L和9mg/L。为模拟高盐废水环境,其中镉和磷浓度为海水中的浓度,属于镉的nM级不饱和高盐环境。
在三组废水中,用钼酸盐分光光度法测定上清液中总磷的浓度变化,结果如图2所示。在高盐环境中,该菌对9mg/L和5mg/L的磷分别有46.24%和72.48%的去除率。
为进一步表征菌体及其形成的纳米颗粒,同样使用原子力显微镜和扫描电镜观察菌体。原子力显微镜结果如图3中A所示,菌体表面均匀分散着纳米颗粒物质,直径均在10nm以下。合成海水中磷镉的浓度在nM级,更低的镉浓度导致了纳米颗粒的粒径更小。另外,此视野中菌体无鞭毛,可能由于断裂而未观察到。
扫描电镜结果如图3中B所示,菌体间有直径10nm的球形颗粒,并粘附在纤维状的胞外多糖上。同时对图中点位的能谱分析表明,颗粒同样主要含有C、O、Cd、P、S元素。但由于培养体系中镉、磷浓度低,颗粒中Cd、P、S的比例较图1E中结果更低。
因此,该菌在磷镉浓度不饱和的高盐废水中也可在胞外形成夹杂多糖的镉-磷-硫纳米颗粒,直径为10nm。
Claims (5)
1.假交替单胞菌(Pseudoalteromonas sp.)DSBS在镉磷的nM级不饱和高盐废水中制备纳米材料的应用;所述假交替单胞菌DSBS已于2013年12月12日保藏于中国典型培养物保藏中心,地址:武汉市武昌珞珈山,保藏号:CCTCCM2013652,所述高盐废水是指总盐质量分数在1%及以上的水体。
2.一种镉-磷-硫纳米材料的制备方法,其特征在于,所述镉-磷-硫纳米材料由假交替单胞菌(Pseudoalteromonas sp.)DSBS在镉磷的nM级不饱和高盐废水中生物合成;
包括假交替单胞菌DSBS菌体富集的步骤,将富集的菌体在镉磷的nM级不饱和高盐废水中进行生物合成的步骤,以及将培养后的菌体用去离子水洗涤3000rpm离心的步骤;
所述假交替单胞菌DSBS已于2013年12月12日保藏于中国典型培养物保藏中心,地址:武汉市武昌珞珈山,保藏号:CCTCCM2013652;
所述高盐废水是指总盐质量分数在1%及以上的水体。
3.如权利要求2所述的制备方法,其特征在于,所述菌体富集的步骤包括:将假交替单胞菌DSBS接种于液体LB培养基中,180-220rpm,20-30℃,培养16-24h,所述液体LB培养基的配方为:蛋白胨10g/L,酵母粉3g/L,陈海水配制,海水盐度为3.5%。
4.如权利要求2所述的制备方法,其特征在于,所述生物合成的步骤包括:将富集的菌体接种于所述镉磷的nM级不饱和高盐废水中,180-220rpm,15-30℃,培养42-54h。
5.如权利要求4所述的制备方法,其特征在于,在所述菌体富集的步骤和所述生物合成的步骤之间,将富集的菌体3000rpm离心10min,弃去上清液,再用高盐废水重悬至原体积,得到洗涤一次的菌体;在所述生物合成的步骤中富集的菌体的接种量为0.02%v/v。
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