5 CHAPTER Applications of modified-nanoporous alumina membrane on potassium sensing 5.1 INTRODUCTION 5.1.1 Potassium ion sensors Potassium monitoring in serum, urine and saliva is very important in biomedical field. Potassium monitoring is also important in environmental water and soil samples [1, 2]. In some cases, potassium concentration gives the information related to the physical condition of patients; for example the heart often stops when potassium in human serum increases to the concentration higher than mmol L-1. Due to the importance of potassium analysis in clinical laboratory, highly selective and highly sensitive potassium sensors are required. Sodium and other ions with the size and properties similar to those of potassium ions often challenges measurement of potassium ions in physiological matrix. Many techniques have been developed to overwhelm 136 this problem. There are different techniques for analyzing potassium ion such as spectrophotometry, chromatography [3], thermogravimetry, inductively coupled plasma-atomic emission spectrometry, flow injection analysis [4] and electroanalytical methods [5-9]. Common electrochemical techniques are potentiometry, amperometry and cyclic voltammetry. The potentiometric method which uses a metal hexacyanoferrate modified electrode as the ion-selective electrode possesses many attractive advantages. Potassium sensors with silver hexacyanoferrate modified electrode measure potassium ions in concentration ranging from 8.0×10-5 to mol L-1 [10]. Other Prussian blue analogues such as nickel [11], cobalt [12], and iron hexacyanoferrate [5] were also employed. These sensors have short response time and good sensitivity. Several reports on use of Prussian blue (PB) for detection of potassium ion include thin film of PB on transparent tin oxide substrate in which K+ ions concentration was determined by in-situ measurement of the absorbance changes of PB film under variable reductive potentials [6]. Other PB based sensors include potentiometric methods which typically require electrochemical or chemical pre-conditioning steps prior to K+ sensing in order 137 to maintain a constant [PB]/[ES] ratio during measurements and have poor reproducibility and long analysis time [6]. In contrast, cyclic voltammetry method offers significant advantages of minimal pre-conditioning steps and fast analysis because constant [PB]/[ES] ratio is achieved at the cyclic voltammetric peak potentials under appropriate conditions. However, PB films subjected to potential cycling conditions exhibit poor stability [10, 13]. It is expected that the highly stable PB nanotubes derived in this work will significantly improve the analytical performance of PB sensors using the cyclic voltammetry detection. In addition, higher loading of PB nanotubes within the membrane electrode compared to conventional PB films can be achieved without compromising film thickness. This will enhance the cyclic voltammetric peak currents which enable accurate determination of the PB redox potentials. 5.1.2 Prussian blue PB (Fe4(Fe(CN)6)3·nH2O), a dark blue pigment, was accidentally discovered in 1704 by the Berlin artist Diesbach. PB has received much attention due to their chemical stability, electrochromic reactions, electro- and photocatalytic activity, easy preparation, and low cost. The PB films have 138 been investigated intensively for use in electrochromic displays, fuel cells, solid-state rechargeable batteries [7, 8], and as signal-enhancing devices due to photovoltaic and photoelectrochemical effects. Keggin and Miles was the first researchers who analyzed the crystalline structure of PB based on power diffraction pattern [14]. The PB structure was then determined more precisely by Ludi and co-workers [15]. PB is a mixed valence coordination compound with two redox centers [6, 16] and exhibits an open, zeolitic structure with the iron centers arranged in a cubic lattice with cyanide bridging the iron centers [9]. The zeolitic nature of PB is the cubic cell of 10.2 Å. Alternating iron (II) and iron (III) ions locate on the face of the center cubic lattice. The iron (II) ions are surrounded by carbon atoms while the iron (III) ions are surrounded by nitrogen atoms (Fig. 5.1). Fig. 5.1 Cubic structure of PB. Adapted from reference[17]. 139 5.2 Template-based nano material synthesis In the template-based nano material synthesizes, the templates play role as the framework. Either around or within the template, the typical materials can be grown in situ and shaped into the nanostructure with the morphology dictated by the size and shape of the template scaffold. The template syntheses are relatively simple, reasonably high-throughput, cost-effective procedure and therefore applicable and environmentally acceptable. Many templates have been used in researches by generally they can be divided into “hard” templates and “soft” templates [18]. The soft templates can be associated with gels, micelles, chitin scaffolds. DNA strands, polymer matrices, and reverse micelles are also listed in the soft template. Materials such as nanoporous alumina membranes, track-etched polycarbonate and other nanoporous membranes are considered as the hard templates. Nanoporous alumina membranes are ideal hard templates owing to their high porous density, cylindrical pore structure, and thermal and chemical ability. Nanoporous alumina membranes are used to transfer the nanopore arrangement to other materials. An enormous variety of nanomaterials (listed below) have been successfully fabricated on the basis of nanoporous alumina membranes. The 140 nano materials synthesized using nanoporous alumina membranes can be categorized into the following groups[19]: • Metal nanodots, nanowire, nanorods and nanotubes • Metal oxide nanodots, nanowire and nanotubes • Semiconductor nanodots, nanowires, nanopillars and nanopore arrays. • Polymer, organis and inorganic nanowires and nanotubes. • Carbon nanotubes. • Photonic crystals • Other nanomaterials. In other researches, alumina membranes were simply used as the template. In our research, the platinum-coated alumina membrane system was employed not only as a nano-material synthesis template but also as a sensor. We exploited the electrochemical deposition technique to fabricate Prussian blue nanotubes inside the membrane nanochannels. The platinum-coated layers on both sides of the membrane served as the working and counter/reference electrodes for both electrodeposition of PB nanotubes and sensing of K+ ions. 141 5.2.1 Depositions of Prussian Blue nano structure The PB can be synthesized by chemical and electrochemical methods. By the chemical methods, PB is formed by mixing ferric salt with hexacyanoferrous complex (Fe3++[FeII(CN)6]4-) or ferrous salt with hexacyanoferric complex (Fe2+ + [FeIII(CN)6]3-); while the nano structure PB was developed layer-by-layer using the diffusion-based fabrication method by which nanochannel structure templates were repeatedly dipped into a sequent solutions of Fe3+,[FeII(CN)6]4-, and H2O to wash electrode [20]. The electrokinetic method was also applied in growing the PB nantotube inside the nanoporous alumina membrane [21]. In the electrochemical methods, deposition of PB is carried from the solutions contained a mixture of ferric Fe3+ and ferricyanide [FeIII(CN)6]3-. The electrochemical methods are either spontaneous applying an open-circuit regime or by a reductive electrochemical driving force. The open-circuit deposition is highly depending on the electrode support. The (FeIII [FeIII(CN)6]) is oxidized by the conductive material which form the PB after one-electron reduction [22, 23]. In the published reports, electrochemical growth of PB started from the metal base which coves the entire pore openings on one side of the nano-scale porous membrane template. 142 In this work, we use membrane-based electrode which is porous platinum-coated membrane template with partially covered pore openings described previously in Chapters and 3. In this way, electrochemical growth of PB within the nanochannels starts from the narrow ring of metal coating at the pore edges, producing only PB tubes even at long electrodeposition time. 5.3 EXPERIMENTAL 5.3.1 Reagents and materials Potassium ferricyanide, ferric chloride, potassium chloride, sodium chloride and hydrochloric acid (37%) were purchased from (Sigma-Aldrich, Singapore). Tris buffer (1 M, pH 7) was obtained from 1st Base, Singapore. All chemicals and solvents used were of analytical grade and used as received. All solutions were prepared in Milli-Q ultrapure water (Millipore). 60 μm thick nanoporous alumina membranes with 200 nm nominal pore size were obtained from Whatman (Maidstone, Kent, UK). All membranes were washed with hydrogen peroxide (Scharlau) before use. Subsequently, one side of the membrane was sputter-coated with ca. 50 nm thick porous platinum layer using platinum target (99.99% purity) in JEOL Auto Fine Coater (JFC-1600). 143 5.3.2 Instrumentation Electrochemical measurements were performed with a CHI400 electrochemical workstation (CH instruments). A three-electrode system was employed for the electrodeposition, characterization and sensing application, using the metal layer of the membrane electrode or PB nanotube-modified membrane electrode as working electrode, Ag/AgCl electrode as reference electrode and a platinum wire as the auxiliary electrode was employed for the electrodeposition, characterization and sensing application. The preparation of working electrode is reported in the next section. Scanning electron micrographs (SEM) and energy dispersive X-ray spectra (EDX) of the membrane electrodes were obtained using the FE-SEM JSM-6701F with an EDX analyzer. Samples in SEM analyses must be electrically conductive. In our experiment, the samples such as Prussian blue or bare alumina membrane are not conductive enough, therefore before SEM analyses they were coated with a thin layer of platinum deposited by sputtering method. 5.3.3 Development of PB nanotube membrane electrode Scheme 5.1 illustrates the preparation of PB nanotube membrane electrode. Nanoporous alumina membranes were washed and pre-treated in 35 % 144 hydrogen peroxide; subsequently, one side of the membrane was sputter-coated with ca. 50 nm thick porous platinum layer using platinum target (99.99% purity) in JEOL Auto Fine Coater. The coated membrane then was modified by electrodeposition of PB nanotubes along the nanochannels of the membrane electrode. The electrodeposition was carried out by repeated potential scan from -500 to 600 mV forth and back at the scan rate of 50 mV s-1 in a solution containing ferric chloride, potassium ferricyanide, potassium chloride and hydrochloric acid. Scheme 5.1 Schematics of the construction of PB-nanotube modified membrane electrode from nanoporous alumina membrane template. (a) Sputter coating of alumina membrane template with ca. 50 nm thick conductive porous platinum layer. (b) Electrodeposition of Prussian blue nanotubes along the nanochannels starting from the porous metal layer. 5.3.4 Voltammetry of PB nanotubes The PB nanotube-modified membrane electrodes were investigated using 145 ��同一个源流。第七届委请中国导演前来执导，或许一定程度上曾参考九鲤 洞原有的戏簿版本，但终究因为传统和传承的不同而出现差异。从各个戏簿的 分簿结构而言，九鲤洞目连戏戏簿对于情节的发展，有它特定的结构方式，并 不恪守郑之珍本的三团圆结局。郑本第一卷从〈元旦上寿〉到〈母子团圆〉、 第二卷从〈寿母劝善〉到〈见佛团圆〉、第三卷从〈师友讲道〉到〈见佛团 圆〉。以九鲤洞第七届本为例，戏簿第一本从〈相春〉到〈罗卜经商〉、第二 本从〈佛母开荤〉到〈闺房剪发〉、第三本从〈佛母回煞〉到〈超度团圆〉。 九鲤洞目连戏戏簿这样的分簿结构，在目连戏研究史上究竟具有甚么意义？ 在郑之珍本中，第一卷已经包含了九鲤洞第二本的〈刘氏开荤〉情节，第 二卷包含九鲤洞第三本的〈刘氏回煞〉、〈过黑松林〉、〈过寒冰池〉、〈过 火焰山〉、〈过烂沙河〉、〈擒沙和尚〉等的情节，而第三卷又包含〈曹府元 宵〉、〈曹氏清明〉、〈公子回家〉、〈见女托媒〉、〈求婚逼嫁〉、〈曹氏 剪发〉、〈曹氏逃难〉、〈曹氏到庵〉、〈曹公见女〉、〈曹氏赴会〉等大量 关于曹女的情节。由此看来，当九鲤洞目连戏采取第二本从〈佛母开荤〉、第 三本从〈刘氏回煞〉的开场结构时，显然得删节大量郑本中的情节，才有可能 完成三本在情节分布上的平衡。 从以上两个版本的比较，我们可以很快看出九鲤洞目连戏在情节上的删 节。第一是九鲤洞目连戏在曹女情节上的大量删节。九鲤洞目连戏的曹女情 节，开始于第一本第三出至第五出，第二本或第三本只以两出交待曹女情节的 发展，相对于郑本上卷第十三出〈修斋荐父〉和第三卷大量的曹女情节，九鲤 洞目连戏显然“清减”许多。在结构上，第三本曹女情节的大量删减，有助于 28 同注（13）。 80 九鲤洞目连戏在第三本上专注于地狱情节的摹写。第二，是九鲤洞目连戏减少 观音和西游记的情节，包括郑本中〈观音生日〉、〈化强从善〉、〈观音渡 阨〉、〈遣将擒将〉、〈过寒冰池〉、〈过火焰山〉、〈过烂沙河〉、〈擒沙 和尚〉等出目。相较于郑之珍本和泉州目连戏而言，九鲤洞目连戏中显然观音 的戏份减少很多，尤其《西游记》诸多展现观音救苦本色的情节，九鲤洞目连 戏多不保留，而着重保留目连变文中，世尊对于目连情节发展上的推动。第 三，是九鲤洞目连戏删去十友情节、〈罗卜辞官〉等细节，显然九鲤洞目连戏 认为这些细节对整体情节推动没有甚么影响。 从上述九鲤洞目连戏在情节删节上的讨论，我们发现所删节的情节都出现 在郑本的第二卷和第三卷。删节了这些情节，配合上一节的“布施”观念的发 挥，也可以发现“上布施”、“下布施”和“普施”是三本分簿结构背后的思 想观念，或人物德行修持的重心。第一本着重于傅相行善的“上布施”的描 摹，以及刘贾为恶的彰显。第二本开始于刘氏造恶的开端，间中还描写刘氏以 恶掩恶的恶劣行径，最终落至被拘拿入地狱的下场。中间穿插罗卜归家后继承 父志行善的善举，也就是“下布施”的描摹。第三本完全着力于描写刘氏落入 地狱后的场景，以及目连如何透过一切努力，将母亲救出地狱的情节。最后再 以“普施回向”结束整场目连戏的演出。 综上所说，本文从九鲤洞过去目连戏演员对于目连戏的简单描摹，得出九 鲤洞目连戏戏簿在分簿上的结构原则，再度呈现戏簿发挥布施观念的情节重 心。全剧就是在布施主轴的贯串下，突显出九鲤洞逢甲大普度中的表演重心。 这一布施的主轴，完全符合九鲤洞主教卢士元对“人生在世则营于名利，未免 奔走四方异路，游历于万里夷邦，苟受风霜疫气而失身他邦，魂随蝶化，无所 依附，诚可悲也”的感叹，29进而希望救度这些众生的悲愿。 其次，就结构的意义而言，九鲤洞目连戏戏簿的分簿结构，清楚地呈现出 三天目连戏演出的情节重心。戏簿第一本从〈相春〉到〈罗卜经商〉，着重傅 相为善布施的善举，终于可以上登天界，享受天道的果报，是为“上布施”。 不过，第一本以〈罗卜经商〉作为小收煞，不仅没有小团圆的情景，反倒是离 29 附录4．1．2．：南方纠察院部暨琼瑶法教主教卢士元示谕牓文。 81 别的景象，明显不符合南戏的结果原则。本文认为以〈罗卜经商〉作为小收 煞，实际上是为了让第二本从〈佛母开荤〉开始。原因在于〈佛母开荤〉是九 鲤洞目连戏� [...]... to water Fig 5. 3 Scanning electron micrograph of nanoporous alumina membrane template (A) before and (B) after sputtering of ~50 nm thick metal (Pt) layer ( 450 tilted surface views); (C) 450 tilted surface view of PB nanotubes after removal of the metal layer and partial dissolution of alumina template; (D) cross-section of a PB-nanotube modified membrane electrode showing the penetration of PB into... of PB nanotubes in membrane electrode Fig 5. 2 shows cyclic voltammograms of Pt-coated alumina membrane electrode in a solution containing 2 .5 mM FeCl3, 2 .5 mM K3Fe(CN)6, 0.1 M KCl, 0.1 M HCl at the scan rate of 50 mV s-1 After electrodeposition of PB, the dark grey electromembrane acquired the typical intense blue color of PB The electrodeposition reaction of PB is given in Eqn 5. 1 [22] 4Fe3+ + 3e-... ratio of PB nanotubes, rapid access of ions to the nanotubes can be achieved Thus, PB nanotube sensor exhibit excellent analytical performance for K+ analysis when combined with cyclic voltammetry methods at moderate scan rate of 50 mV s-1 154 5. 4.3 Performance of PB nanotube K+ selective sensor Fabrication of fast response and stable potassium ion selective sensor are useful for measurement of physiological... SEM/EDX spectrum of PB in the nanochannels showing the presence of Fe and K; (E) surface view of PB nanotubes after dissolution of alumina template 149 5. 4.2 Cyclic voltammetry of PB nanotubes 0.4 0 .5 M KCl 0.3 5 ×10-4 M 0.1 -0.1 0.3 E mp (V) vs Ag/AgCl Current (mA) 0.2 -0.2 -0.3 -0.4 -0 .5 -0.1 Slope = 58 0.2 V 0.1 0 -10.0 0.1 0.3 -8.0 0 .5 -6.0 -4.0 Log [K+] -2.0 0.0 0.7 E (V) vs Ag/AgCl Fig 5. 4 Cyclic voltammograms... because of the high surface-to-volume ratio of nanotubes For example, 5 μm length , 25 nm thick wall PB nanotubes grown within the membrane electrode (0.66 cm2 geometric area; 200 nm pore diameter; 1.2 x 109 pores cm-2) gives surface-to-volume ratio of 3.4 x 1 05, compared to the ratio of 1 x 104 for a conventional PB film grown on planar electrode of the same geometric area The width and height of one... Cross-section profile of the membrane electrode after 15 electrodeposition potential cycles reveals growth of PB nanotubes starting from the metal layer (Fig 5. 3 D) SEM/EDX spectrum of the deposited PB given in Fig 5. 3 D shows the presence of K and Fe In the control sample of the alumina template without PB, only Al and O were observed Removal of the metal layer and partial removal of the alumina template... HCl Scan rate 50 mV s-1 Fig 5. 3 A and B show the scanning electron micrographs of the tilted surface view of the nanoporous membrane and the metal-coated membrane 147 Under the optimal condition, the metal layer partially covers the pore and reduces the nominal pore size from 200 nm to 150 nm Thus, a ring-shaped metal base of ca 25 nm ring thickness is available for the electrodeposition of PB nanotubes... pollution [ 35] To evaluate the sensing performance of the PB nanotube sensor, we measure its stability, reproducibility, analysis time, ion selectivity, linear range and limit of detection 3.0 Current (mA) 1st 50 th 100th 1.0 -1.0 -3.0 -0.1 0.1 0.3 0 .5 E (V) vs Ag/AgCl Fig 5. 6 Cyclic voltammograms of a PB-nanotube membrane electrode at 1, 50 and 100 repeated cycles showing the high stability of the PB... 0.8 Fig 5. 7 Selectivity of PB nanotube sensor for K+ ions and interferences from other cations Experimental conditions are same as in the case of Fig 5. 4 Cyclic voltammogram of the PB membrane electrode in 1 M pH 7 Tris buffer containing (A) blank, 0. 05 M NaCl solution and 0. 05 M KCl; (B) 0. 05 M KCl in the presence of additional cation Mg2+, Ca2+ or Na+ at 0 M, 0.017 M, 0. 05 M, 0.17 M (corresponding... mole fraction of 1, 0. 75, 0 .5 and 0. 25 respectively) Mid-peak potential remains constant in different concentrations of these other cations An additional advantage of PB nanotube sensor is the presence of non-Nernstian linear region 5. 0×10-8 – 7.0×10-4 M in the plot of Emp versus Log[K+] (Fig 5. 8) This non-Nernstian relation widens the linear sensing range considerably and lowers the limit of detection . scan rate of 50 mV s -1 . 154 5. 4.3 Performance of PB nanotube K + selective sensor. Fabrication of fast response and stable potassium ion selective sensor are useful for measurement of physiological. nm to 150 nm. Thus, a ring-shaped metal base of ca. 25 nm ring thickness is available for the electrodeposition of PB nanotubes. Cross-section profile of the membrane electrode after 15 electrodeposition. both sides of the membrane served as the working and counter/reference electrodes for both electrodeposition of PB nanotubes and sensing of K + ions. 141 5. 2.1 Depositions of Prussian