X-ray diffraction(20120412)

Chapter 1. The Human Genome1. The Human GenomeLearning outcomesWhen you have read Chapter 1, you should be able to:•Describe the two experiments that led molecular biologists to conclude that genes are made of DNA, and state the limitations of each experiment•Give a detailed description of the structure of a polynucleotide and summarize the chemical differences between DNA and RNA•Discuss the evidence that Watson and Crick used to deduce the double helix structure of DNA and list the key features of this structure•Explain why the DNA double helix has structural flexibility•Describe in outline the content of the human nuclear genome•Draw the structure of an ‘average' human gene•Categorize the human gene catalog into different functional classes•Distinguish between conventional and processed pseudogenes and other types of evolutionary relic•Give specific examples of the repetitive DNA content of the human genome•Give an outline description of the structure and organization of the human mitochondrial genome•Discuss the importance of the Human Genome ProjectLife as we know it is specified by the genomes of the myriad organisms with which we share the planet. Every organism possesses a genome that contains the biological information needed to construct and maintain a living example of that organism. Most genomes, including the human genome and those of all other cellular life forms, are made of DNA (deoxyribonucleic acid) but a few viruses have RNA (ribonucleic acid) genomes. DNA and RNA are polymeric molecules made up of chains of monomeric subunits called nucleotides.The human genome, which is typical of the genomes of all multicellular animals, consists of two distinct parts (Figure 1.1):•The nuclear genome comprises approximately 3 200 000 000 nucleotides of DNA, divided into 24 linear molecules, the shortest 50 000 000 nucleotides in length and the longest 260 000 000 nucleotides, each contained in a different chromosome. These 24 chromosomes consist of 22 autosomes and the two sex chromosomes, X and Y.•The mitochondrial genome is a circular DNA molecule of 16 569 nucleotides, multiple copies of which are located in the energy-generating organelles called mitochondria. Each of the approximately 1013 cells in the adult human body has its own copy or copies of the genome, the only exceptions being those few cell types, such as red blood cells, that lack a nucleus in their fully differentiated state. The vast majority of cells are diploid and so have two copies of each autosome, plus two sex chromosomes, XX for females or XY for males - 46 chromosomes in all. These are called somatic cells, in contrast to sex cells or gametes, which are haploid and have just 23 chromosomes, comprising one of each autosome and one sex chromosome. Both types of cell have about 8000 copies of the mitochondrial genome, 10 or so in each mitochondrion.Figure 1.1. The nuclear and mitochondrial components of the human genome. For more details on the anatomy of the human genome, see Section 1.2.This book is about genomes. It explains what genomes are (Part 1), how they are studied (Part 2), how they function (Part 3), and how they replicate and evolve (Part 4). We begin our journey with our own genome, which is quite naturally the one that interests us the most. Later in this chapter we will examine how the human genome is constructed, some of this information dating from the old days when biologists studied genes rather than genomes, but much of it revealed only since the Human Genome Project was completed in the first year of the new millennium. First, however, we must understand the structure of DNA.1.1. DNADNA was discovered in 1869 by Johann Friedrich Miescher, a Swiss biochemist working in Tübingen, Germany. The first extracts that Miescher made from human white blood cells were crude mixtures of DNA and chromosomal proteins, but the following year he moved to Basel, Switzerland (where the research institute named after him is now located) and prepared a pure sample of nucleic acid from salmon sperm. Miescher's chemical tests showed that DNA is acidic and rich in phosphorus, and also suggested that the individual molecules are very large, although it was not until the 1930s when biophysical techniques were applied to DNA that the huge lengths of the polymeric chains were fully appreciated.Three years before Miescher discovered DNA, Gregor Mendel had published the results of his breeding experiments with pea plants, carried out in the monastery gardens at Brno, a centraleuropean city some 550 km from Tübingen in what is now the Czech Republic. Mendel's paper in the Proceedings of the Society of Natural Sciences in Brno describes his hypothesis that inheritance is controlled by unit factors, the entities that geneticists today call genes. It is very unlikely that Miescher and Mendel were aware of each other's work, and if either of them had happened to read about the other's discoveries then they certainly would not have made any connection between DNA and genes. To make such a connection - to infer that genes are made of DNA - would have been quite illogical in the late 19th century or indeed for many decades afterwards. The precise biological function of DNA was not known, and the supposition that it was a store of cellular phosphorus seemed entirely reasonable at the time. The chemical nature of genes was equally unknown, and indeed was an irrelevance for most geneticists, who in the years immediately after 1900, when Mendel's work was rediscovered, were able to make remarkable advances in understanding heredity without worrying about what genes were actually made of.It was not until the 1930s that scientists began to ask more searching questions about genes. In 1944, Erwin Schrödinger, more famous for the wave equation which still terrifies many biology students taking introductory courses in physical chemistry, published a book entitled What is Life?, which encapsulated a variety of issues that were being discussed not only by geneticists but also by physicists such as Niels Bohr and Max Delbrück. These scientists were the first molecular biologists and the first to suggest that ‘life' could be explained in molecular terms; our current knowledge of how the genome functions stems directly from their pioneering work. The starting point for the new molecular biology was to discover what genes are made of.1.1.1. Genes are made of DNAHow could the molecular nature of the genetic material be determined? Back in 1903, WS Sutton had realized that the inheritance patterns of genes paralleled the behavior of chromosomes during cell division. This observation led to the proposal that genes are located in chromosomes and by the 1930s it was universally accepted that the chromosome theory was correct. Examination of cells by cytochemistry, after staining with dyes that bind specifically to just one type of biochemical, had shown that chromosomes are made of DNA and protein, in roughly equal amounts. Some biologists looked on the combination between the two (‘nucleoprotein') as the genetic material, but others argued differently. From today's perspective it can be difficult to understand why these arguments favored the notion that genes were made, not of DNA, but of protein. The explanation is that, at the time, many biochemists thought that all DNA molecules were the same, which meant that DNA did not have the immense variability that was one of the postulated features of the genetic material. Billions of different genes must exist and for each one to have its own individual activity, the genetic material must be able to take many different forms. If every DNA molecule were identical then DNA could not satisfy this requirement and so genes must be made of protein. This assumption made perfect sense because proteins were known, correctly, to be highly variable polymeric molecules, each one made up of a different combination of 20 chemically distinct amino-acid monomers (Section 3.3.1).The errors that had been made in understanding DNA structure lingered on until the late 1930s. Gradually, however, it was accepted that DNA, like protein, has immense variability. Could DNA therefore be the genetic material? The results of two experiments performed during the middle decades of the 20th century forced biologists to take this possibility seriously.Bacterial genes are made of DNAThe first molecular biologists realized that the most conclusive way to identify the chemical composition of genes would be to purify some and subject them to chemical analysis. But nothing like this had ever been attempted and it was not clear how it could be done. Ironically, the experiment was performed almost unwittingly by a group of scientists who did not look upon themselves as molecular biologists and who were not motivated by a curiosity to know what genes are made of. Instead, their objective was to find a better treatment for one of the most deadly diseases of the early 20th century, pneumonia.Before the discovery of antibiotics, pneumonia was mainly controlled by treating patients in the early stages of the disease with an antiserum prepared by injecting dead cells of the causative bacterium (now called Streptococcus pneumoniae) into an animal. In order to prepare more effective antisera, studies were made of the immunological properties of the bacterium. It was shown that there is a range of different types of S. pneumoniae, each characterized by the mixture of sugars contained in the thick capsule that surrounds the cell and elicits the immunological response (Figure 1.2A). In 1923, Frederick Griffith, a British medical officer, discovered that as well as the virulent strains, there were some types of S. pneumoniae that did not have a capsule and did not cause pneumonia. This discovery was not a huge surprise because other species of pathogenic bacteria were known to have avirulent, unencapsulated forms. However, 5 years later Griffith obtained some results that were totally unexpected (Griffith, 1928). He performed a series of experiments in which he injected mice with various mixtures of bacteria (Figure 1.2B). He showed that, as anticipated, mice injected with virulent S. pneumoniae bacteria developed pneumonia and died, whereas those injected with an avirulent type remained healthy. What he did not anticipate, however, was what would happen when mice were injected with a mixture made up of live avirulent bacteria along with some virulent cells that had been killed by heat treatment. The only live bacteria were the harmless ones, so surely the mice would remain healthy? Not so: the mice died. Griffith carried out biopsies of these dead mice and discovered that their respiratory tracts contained virulent bacteria, which were always of the same immunological type as the strain that had been killed by heat treatment before injection. Somehow the living harmless bacteria had acquired the ability to make the capsule sugars of the dead bacteria. This process - the conversion of living harmless bacteria into virulent cells - was called transformation. Although not recognized as such by Griffith, the transforming principle - the component of the dead cells that conferred on the live cells the ability to make the capsular sugars - was genetic material.Figure 1.2. Griffith's experiments with virulent and avirulent Streptococcus pneumoniae bacteria. (A) Representation of a S. pneumoniae bacterium. A serotype is a bacterial type with distinctive immunological properties, conferred in this case by the combination of sugars present in the capsule. Avirulent types have no capsule. (B) The experiments which showed that a component of heat-killed bacteria can transform living avirulent bacteria into virulent cells. Griffith showed that the avirulent bacteria were always transformed into the same serotype as the dead cells. In other words, the living bacteria acquired the genes specifying synthesis of the capsule of the dead cells.Oswald Avery, together with his colleagues Colin MacLeod and Maclyn McCarty, of Columbia University, New York, set out to determine what the transforming principle was made of. The experiments took a long time to carry out and were not completed until 1944 (Avery et al., 1944). But the results were conclusive: the transforming principle was DNA. The transforming principle behaved in exactly the same way as DNA when subjected to various biophysical tests, it was inactivated by enzymes that degraded DNA, and it was not affected by enzymes that attacked protein or any other type of biochemical (Figure 1.3).Figure 1.3. The transforming principle is DNA. Avery and his colleagues showed that the transforming principle is unaffected by treatment with a protease or a ribonuclease, but is inactivated by treatment with a deoxyribonuclease.Avery's experiments were meticulous but, because of several complicating factors, they did not immediately lead to acceptance of DNA as the genetic material. It was not clear in the minds of all microbiologists that transformation really was a genetic phenomenon, and few geneticists really understood the system well enough to be able to evaluate Avery's work. There was also some doubt about the veracity of the experiments. In particular, there were worries about the specificity of the deoxyribonuclease enzyme that he used to inactivate the transforming principle. This result, a central part of the evidence for the transforming principle being DNA, would be invalid if, as seemed possible, the enzyme contained trace amounts of a contaminating protease and hence was also able to degrade protein. These uncertainties meantthat a second experiment was needed to provide moreinformation on the chemical nature of the geneticmaterial.Virus genes are made of DNAFigure 1.4. Bacteriophages are viruses that infect bacteria. (A) The structure of a head-and-tail bacteriophage such as T2. The DNA genome of the phage is contained in the head part of the protein capsid. (B) The infection cycle. After injection into an Escherichia coli bacterium, the T2phage genome directs synthesis of new phages. For T2, the infection cycle takes about 20 minutes at 37 °C and ends with lysis of the cell and release of 250–300 new phages. This is the lytic infection cycle. Some phages, such as λ, can also follow a lysogenic infection cycle, in which the phage genome becomes inserted into the bacterial chromosome and remains there, in quiescent form, for several generations of the bacterium (Section 4.2.1).The second experiment was carried out by two bona fide molecular biologists, Alfred Hershey and Martha Chase, at Cold Spring Harbor, New York, in 1952 (Hershey and Chase, 1952). Like the work on the transforming principle, the Hershey-Chase experiment was not done specifically to determine the chemical nature of the gene. Hershey and Chase were two of several biologists who were studying the infection cycle of bacteriophages (or ‘phages') - viruses that infect bacteria. Phages are relatively simple structures made of just DNA and protein, with the DNA contained inside the phage, surrounded by a protein capsid (Figure 1.4A). To replicate, a phage must enter a bacterial cell and subvert the bacterial enzymes into expressing the information contained in the phage genes, so that the bacterium synthesizes new phages. Once replication is complete, the new phages leave the bacterium, possibly causing its death as they do so, and move on to infect new cells (Figure 1.4B). The objective of Hershey and Chase's experiment was to determine if the entire phage particle entered the bacterium at the start of the infection cycle, or if part of the phage stayed outside. If only one component of the phage - the DNA or the protein - entered the cell then that component must be the genetic material.Figure 1.5. The Hershey-Chaseexperiment. The bacteriophages werelabeled with 32P and 35S. A few minutesafter infection, the culture was agitated todetach the empty phage capsids from thecell surface. The culture was thencentrifuged and the radioactive content ofthe bacterial pellet determined. This pelletcontained most of 32P-labeled componentof the phages (the DNA) but only 20% ofthe 35S-labeled material (the phageprotein). In a second experiment, Hersheyand Chase showed that new phagesproduced at the end of an interruptedinfection cycle contained less than 1% ofthe protein from the parent phages.Their experimental strategy was based onthe use of radioactive labels, which hadrecently been introduced into biology.DNA contains phosphorus, which is absent from protein, so DNA can be labeled specifically with the radioactive phosphorus isotope, 32P. Protein, on the other hand, contains sulfur, which is absent from DNA, and so protein can be labeled with 35S. Hershey and Chase were not the first to useradiolabeling to try to determine which part of the phage entered the cell, but previous experiments by James Watson, Ole Maaløe and others had been inconclusive because of the difficulty in distinguishing between phage material that was actually inside a bacterium and a component that did not enter the cell but remained attached to the outer cell surface. To get round this difficulty, Hershey and Chase made an important modification to the previous experiments. They infected Escherichia coli bacteria with radiolabeled T2 phages but, rather than allowing the infection process to go to completion, they left the culture for just a few minutes and then agitated it in a blender. The idea was that the blending would detach the phage material from the surface of the bacteria, enabling this component to be separated from the material inside the cells by centrifuging at a speed that collected the relatively heavy bacteria as a pellet at the bottom of the tube but left the detached material in suspension (Figure 1.5). After centrifugation, Hershey and Chase examined the bacterial pellet and found that it contained 70% of the 32P-labeled component of the phages (the DNA) but only 20% of the 35S-labeled material (the phage protein). In a parallel experiment, the bacteria were left for 20 minutes, long enough for the infection cycle to reach completion. With T2 phage, the cycle ends with the bacteria bursting open and releasing new phages into the supernatant. These new phages contained almost half the DNA from the original phages, but less than 1% of the protein.Hershey and Chase's results suggested that DNA was the major component of the infecting phages that entered the bacterial cell and, similarly, was the major, or perhaps only, component to be passed on to the progeny phages. These observations lent support to the view that DNA is the genetic material, but were they conclusive? Not according to Hershey and Chase (1952) who wrote ‘Our experiments show clearly that a physical separation of phage T2 into genetic and non-genetic parts is possible . . . . The chemical identification of the genetic part must wait, however, until some questions . . . have been answered.' Even if the experiment had provided compelling evidence that the genetic material of phages was DNA, it would have been erroneous to extrapolate from these unusual life forms (which some biologists contend are not really ‘living') to cellular organisms. Indeed, we know that some phage genomes are made of RNA. The Hershey-Chase experiment is important, not because of what it tells us, but because it alerted biologists to the fact that DNA might be the genetic material and was therefore worth studying. It was this that influenced Watson and Crick to study DNA and, as we will see below, it was their discovery of the double helix structure, which solved the puzzling question of how genes can replicate, that really convinced the scientific world that genes are made of DNA.1.1.2. The structure of DNAThe names of James Watson and Francis Crick are so closely linked with DNA that it is easy to forget that, when they began their collaboration in Cambridge, England in October 1951, the detailed structure of the DNA polymer was already known. Their contribution was not to determine the structure of DNA per se, but to show that in living cells two DNA chains are intertwined to form the double helix. We will consider the two facets of DNA structure separately. Nucleotides and polynucleotidesDNA is a linear, unbranched polymer in which the monomeric subunits are four chemically distinct nucleotides that can be linked together in any order in chains hundreds, thousands or even millions of units in length. Each nucleotide in a DNA polymer is made up of three components (Figure 1.6):Figure 1.6. The structure of a nucleotide. (A) The general structure of a deoxyribonucleotide, the type of nucleotide found in DNA. (B) The four bases that occur in deoxyribonucleotides.1. 2′-deoxyribose, which is a pentose, a type of sugar composed of five carbon atoms. These five carbons are numbered 1′ (spoken as ‘one-prime'), 2′, etc. The name ‘2′-deoxyribose' indicates that this particular sugar is a derivative of ribose, one in which the hydroxyl (-OH) group attached to the 2′-carbon of ribose has been replaced by a hydrogen (-H) group.2. A nitrogenous base, one of cytosine, thymine (single-ring pyrimidines), adenine or guanine (double-ring purines). The base is attached to the 1′-carbon of the sugar by a β-N-glycosidic bond attached to nitrogen number 1 of the pyrimidine or number 9 of the purine.Figure 1.7. A short DNA polynucleotide showing the structure of the phosphodiester bond. Note that the two ends of the polynucleotide are chemically distinct.Figure 1.8. The polymerization reaction that results in synthesis of a DNA polynucleotide. Synthesis occurs in the 5′→3′ direction, with the new nucleotide being added to the 3′-carbon at the end of the existing polynucleotide. The β- and γ-phosphates of the nucleotide are removed as apyrophosphate molecule.3. A phosphate group, comprising one, two or three linked phosphate units attached to the 5′-carbon of the sugar. The phosphates are designated α, β and γ, with the α-phosphate being the one directly attached to the sugar.A molecule made up of just the sugar and base is called a nucleoside; addition of the phosphates converts this to a nucleotide. Although cells contain nucleotides with one, two or three phosphate groups, only the nucleoside triphosphates act as substrates for DNA synthesis. The full chemical names of the four nucleotides that polymerize to make DNA are:•2′-deoxyadenosine 5′-triphosphate•2′-deoxycytidine 5′-triphosphate•2′-deoxyguanosine 5′-triphosphate•2′-deoxythymidine 5′-triphosphateThe abbreviations of these four nucleotides are dATP, dCTP, dGTP and dTTP, respectively, or, when referring to a DNA sequence, A, C, G and T, respectively.In a polynucleotide, individual nucleotides are linked together by phosphodiester bonds between their 5′- and 3′-carbons (Figure 1.7). From the structure of this linkage we can see that the polymerization reaction (Figure 1.8) involves removal of the two outer phosphates (the β- and γ-phosphates) from one nucleotide and replacement of the hydroxyl group attached to the 3′-carbon of the second nucleotide. Note that the two ends of the polynucleotide are chemically distinct, one having an unreacted triphosphate group attached to the 5′-carbon (the 5′ or 5′-P terminus) and the other having an unreacted hydroxyl attached to the 3′-carbon (the 3′ or 3′-OH terminus). This means that the polynucleotide has a chemical direction, expressed as either 5′→3′(down in Figure 1.8) or 3′→5′ (up in Figure 1.8). An important consequence of the polarity of the phosphodiester bond is that the chemical reaction needed to extend a DNA polymer in the 5′→3′ direction is different to that needed to make a 3′→5′ extension. All natural DNA polymerase enzymes are only able to carry out 5′→3′ synthesis, which adds significant complications to the process by which double-stranded DNA is replicated (Section 13.2). The same limitation applies to RNA polymerases, the enzymes which make RNA copies of DNA molecules (Section 3.2.2).RNAFigure 1.9. The chemical differences betweenDNA and RNA. (A) RNA contains ribonucleotides,in which the sugar is ribose rather than2′-deoxyribose. The difference is that a hydroxylgroup rather than hydrogen atom is attached to the2′-carbon. (B) RNA contains the pyrimidine calleduracil instead of thymine.Although our attention is firmly on DNA, thestructure of RNA is so similar to that of DNA that itmakes sense to introduce it here. RNA is also apolynucleotide but with two differences compared with DNA (Figure 1.9). First, the sugar in an RNA nucleotide is ribose and, second, RNA contains uracil instead of thymine. The four nucleotide substrates for synthesis of RNA are therefore:•adenosine 5′-triphosphate•cytidine 5′-triphosphate•guanosine 5′-triphosphate•uridine 5′-triphosphatewhich are abbreviated to ATP, CTP, GTP and UTP, or A, C, G and U, respectively.As with DNA, RNA polynucleotides contain 3′–5′ phosphodiester bonds, but these phosphodiester bonds are less stable than those in a DNA polynucleotide because of the indirect effect of the hydroxyl group at the 2′-position of the sugar. This may be one reason why the biological functions of RNA do not require the polynucleotide to be more than a few thousand nucleotides in length, at most. There are no RNA counterparts of the million-unit sized DNA molecules found in human chromosomes.1.1.3. The double helixIn the years before 1950, various lines of evidence had shown that cellular DNA molecules are comprised of two or more polynucleotides assembled together in some way. The possibility that unraveling the nature of this assembly might provide insights into how genes work prompted Watson and Crick, among others, to try to solve the structure. According to Watson in his book The Double Helix (see Further Reading), their work was a desperate race against the famous American biochemist, Linus Pauling (Section 3.3.1), who initially proposed an incorrect triple helix model, giving Watson and Crick the time they needed to complete the double helix structure (Watson and Crick, 1953). It is now difficult to separate fact from fiction, especially regarding the part played by Rosalind Franklin, whose X-ray diffraction studies provided the bulk of the experimental data in support of the double helix and who was herself very close to solving the structure. The one thing that is clear is that the double helix, discovered by Watson and Crick on Saturday 7 March 1953, was the single most important breakthrough in biology during the 20th century.The evidence that led to the double helixWatson and Crick used four types of information to deduce the double helix structure: •Biophysical data of various kinds. The water content of DNA fibers was particularly important because it enabled the density of the DNA in a fiber to be estimated. The number of strands in the helix and the spacing between the nucleotides had to be compatible with the fiber density. Pauling's triple helix model was based on an incorrect density measurement which suggested that the DNA molecule was more closely packed than it actually is.•X-ray diffraction patterns(Section 9.1.3), most of which were produced by Rosalind Franklin of Kings College, London, and which revealed the helical nature of the structure and indicated some of the key dimensions within the helix.•The base ratios, which had been discovered by Erwin Chargaff of Columbia University, New York. Chargaff carried out a lengthy series of chromatographic studies of DNA samples from various sources and showed that, although the values are different in different organisms, the amount of adenine is always the same as the amount of thymine,。

Thickness effect on electrical properties of Pb(Zr 0.52Ti 0.48)O 3thick films embedded with ZnO nanowhiskers prepared by a hybrid sol –gel routeQ.L.Zhao a ,1,M.S.Cao a ,⁎,1,J.Yuan b ,1,R.Lu a ,D.W.Wang a ,D.Q.Zhang ca School of Materials Science and Engineering,Beijing Institute of Technology,Beijing 100081,Chinab The College of Information Engineering,Central University for Nationalities,Beijing 100081,China cCollege of Chemical Engineering,Qiqihar University,Qiqihar 161006,Chinaa b s t r a c ta r t i c l e i n f o Article history:Received 9October 2009Accepted 14December 2009Available online 23December 2009Keywords:FerroelectricsSol –gel preparation Thick filmsElectrical propertiesSilicon-based lead zirconate titanate thick films embedded with zinc oxide nanowhiskers (ZnO w –PZT)were prepared by a hybrid sol –gel route.ZnO w –PZT films with thickness from 1.5μm to 4μm are perovskite structure and have smooth surface without any cracks.As the thickness increases,the remanent polarization and dielectric constant increase,but the coercive field and tetragonality pared with PZT films,the ZnO w –PZT film has the close tetragonality and electrical properties which are different from those of bulk PZT-based ceramic doped with ZnO powder.The thickness dependences of the ferroelectric and dielectric properties are attributed to the relaxation of internal stress.©2009Elsevier B.V.All rights reserved.1.IntroductionLead zirconate titanate (Pb(Zr 1−x Ti x )O 3)thick films (N 1μm)are functional materials with superior ferroelectric,dielectric and piezoelectric properties and have potential applications in micro-electromechanical systems (MEMS).Sol –gel method is usually used to prepare PZT thick films because of its low sintering temperature,precisely controlling the homogeneity of the com-position,ease of use and low cost to fabricate large area films.Since the pores and cracks appear during annealing,organic polymer solvents [1–3]and non-ferroelectric/ferroelectric powder [4–6]have been added into the PZT sol to improve the electrical and mechanical properties.Recently,zinc oxide (ZnO)was added into piezoelectric ceramics and polymers as an additive to improve the fabrication process,electrical and mechanical properties [7–11].However,there are few studies on the ZnO nanowhiskers (ZnO w )addition in PZT thick films.Furthermore,ZnO nanowire has been proven to be a piezoelectric material [12],so it is expected to provide contributions to the piezoelectric property of PZT thick films.In this work,PZT thick films embedded with ZnO w (ZnO w –PZT)are prepared by a hybrid sol –gelroute,and the effects of thickness on electrical properties are investigated.2.ExperimentalIn brief,0.4mol/L PZT sol was prepared by lead acetate,zirconium nitrate,tetrabutyl titanate and polyvinylpyrrolidone (PVP,4.0×104in average molecular weight)with molar ratios 1.1:0.52:0.48:1.ZnO w were synthesized by combustion oxidation of pure zinc powders without any catalysts or additives [13].Then ZnO w slurry was prepared by dispersing ZnO w in ethanol with a concentration of 0.05mol/L.A PbTiO 3(PT)layer (∼100nm)was first deposited as a seed layer on a Pt/Cr/SiO 2/Si substrate by the sol –gel method.The PZT sol was spin-coated on the substrate with a spinning rate of 1600rpm for 1min and then the wet film was dried at 450°C for 30min.The ZnO w slurry was spin-coated on the PZT film with a spinning rate of 3000rpm for 1min,and dried at 100°C for 3min to evaporate the ethanol.Subsequently,the spin-coating procedure of PZT film was repeated until the desired thickness was achieved.The multilayer film was annealed at 700°C for 5min in air.In order to remove redundant ZnO w ,the thick film was dipped in dilute acetic acid solution for 30s.Finally,another PT layer (∼100nm)was deposited on the top of films to fill the pores and smooth the surface.X-ray diffraction (XRD)analyses were carried out with a Cu K αradiation (X'pert pro MPD,PANalytical).Electron micrographs were obtained by a scanning electron microscope (SEM,Hitachi S-3500N)and an energy dispersive spectrometer (EDS,Oxford Inca).TheMaterials Letters 64(2010)632–635⁎Corresponding author.Tel.:+8601068914062.E-mail addresses:caomaosheng@ (M.S.Cao),yuanjie4000@ (J.Yuan).1These authors contributed equally to thiswork.0167-577X/$–see front matter ©2009Elsevier B.V.All rights reserved.doi:10.1016/j.matlet.2009.12.028Contents lists available at ScienceDirectMaterials Lettersj o u r na l ho m e p a g e :w w w.e l s ev i e r.c o m /l o c a t e /m a t l e tferroelectric and dielectric properties were measured by a ferro-electric test system (Precision Premier II,Radiant Technologies,Inc.)and precision impedance analyzer (Agilent 4294A).3.Results and discussionFig.1shows the XRD of ZnO w ,PZT and ZnO w –PZT thick films with different thicknesses.It is seen that ZnO w is well indexed and exhibits a hexagonal wurtzite structure.All the PZT-based thick films show perovskite structures with the dominant peaks corresponding to (110)and (100).The cell parameters are calculated from XRD results by least-square method.It is indicated that all the films are tetragonal phase,the cell parameter c is smaller,and a is larger than that of bulk PZT (a =4.032,c =4.147[14]),pressive stresses exist in the c direction.With the increasing thickness of the ZnO w –PZT film,c decreases and a increases,which make the tetragonality (c /a )pared with PZT film,ZnO w –PZT film gives the similar cell lattice parameters and tetragonality.In the previous report [7],it has been found that the addition of ZnO increases the tetragonality of PZT-based ceramics due to Zn 2+ion substitution of B-site ions because of its smaller ion radiuses.The discrepancy between the film and ceramic results should be attributed to the different fabrication processes.In this work,the film annealing temperature is 700°C,which is much smaller than that of ceramic (≥900°C),and there is nearly no Zn 2+ion substitution of B-site ions.Therefore,the addition of ZnO w has weak in fluence of the tetragonality of PZT films.The decreases of tetragonality should be caused by theinternal stress relaxation,which would reduce the effects of lattice mismatch between the films and substrates.Fig.2(a)shows the SEM micrograph of ZnO w and it is seen that ZnO w is tetraneedlelike structure,of which one needle length is ∼10μm.Fig.2(b)shows a ZnO w (in dash circle)embedded in 1μm-thick PZT film without any surface treatment.Three bottom needles are buried in the PZT film tightly and no crack appears at the boundary.Fig.2(c)and (d)shows the mapping of Zn and Pb elements,respectively.Due to the larger ZnO w dimension than the film thickness,the uncovered ZnO w would in fluence the surface rough-ness.Thus,dilute acetic acid solution is used to remove redundant ZnO w .Fig.2(e)and (f)shows the surface morphology and cross section of a 2-μm-thick ZnO w –PZT film completed by all processes.It is revealed that the film is of granular structure and crack free.The grain sizes are in the range of sub-micrometers.Fig.3(a)shows the ferroelectric hysteresis loops of the 2-μm-thick PZT and ZnO w –PZT films with an applied maximum voltage of 90V at a frequency of 10Hz.The remanent polarizations P r are 27.4and 24.3μC/cm 2,and both coercive fields E c are 176.5kV/cm.The smaller P r of the ZnO w –PZT film is attributed to the substitution of ZnO w for PZT,where the ferroelectric is replaced by a non-ferroelectric.However,ZnO w seems to have no effect on E c .Fig.3(b)shows the steady-state leakage current –voltage characteristics of the two films.The value of ZnO w –PZT film is larger than that of the PZT film under the same applied voltage.It is consistent with the fact that ZnO is a semiconductor and has less insulation than that of PZT.Therefore,ZnO w in PZT film would lead to more leakage current flowing through them.Fig.3(c)and (d)show that P r increases and E c decreases as the film thickness increases,which are caused by the film relaxation and structure activity increase,i.e.they are helpful for the movement of Ti 4+,Zr 4+,O 2−ions and the reverse of spontaneous polarization [15].The dielectric constants εr and dielectric loss tan δof the 2-μm-thick PZT film and ZnO w –PZT films are measured in a frequency range of 5–1000kHz,as shown in Fig.4.Here εr decreases slightly with increasing frequency;however,tan δhas less dependence on frequency except that of the 3-μm-thick ZnO w –PZT film.In the sol –gel fabrication process,some ZnO w aggregation in PZT thick films could not be avoided and would lead to more leakage current flowing through ZnO w and generate larger conduction loss.The enhancement of εr at low frequency could be attributed to the interfacial polarization,which has small contribution at high frequency [16].The increase of εr with the increasing film thickness is attributed to the effect of interfacial layers attenuated [17].Compared with PZT films,εr and tan δof ZnO w –PZT films are smaller in the measurement frequency range.According to the effective-medium theory,the addition of ZnO w ,an inclusive phase with lower εr ,will reduce εr of PZT films [18].In previous reports,it has been found that the stress in sol –gel derived PZT and PT films decreased with the increased film thickness [15,19].In the present study,the stress relaxation in ZnO w –PZT films reduces the tetragonality and makes non-180°domain wall motions easily,which would result in the increase of P r and decrease of E c [20].Meanwhile,as the film thickness increases,the effect of interfacial layers between PZT and substrate becomes weak and raise the εr of ZnO w –PZT films.4.ConclusionSilicon-based ZnO w –PZT thick films have been prepared via a hybrid sol –gel route using ZnO w suspension and PZT sol.As the film thickness increases,film tetragonality decreases and the film is pared with PZT film,it is suggested that the addition of ZnO w has less effect on the tetragonality of PZT films.ZnO w –PZT films show good ferroelectric and dielectric properties and probably are candidates in MEMSapplications.Fig.1.XRD of (a)ZnO w ,(b)2-μm-thick PZT and ZnO w –PZT films with thicknesses of (c)1.5μm,(d)2μm,(e)3μm,and (f)4μm.The lower figure shows the dependences of cell parameter and tetragonality on film thickness.633Q.L.Zhao et al./Materials Letters 64(2010)632–635634Q.L.Zhao et al./Materials Letters64(2010)632–635Fig.2.SEM micrographs of(a)ZnO w,(b)a ZnO w(in dash circle)embedded in PZTfilm,(c)mapping of Zn element,(d)mapping of Pb element,(e)surface morphology and(f)cross lm.section of the2-μm-thick ZnO w–PZTfiAcknowledgementThis study was supported by the National High-Tech Research and Development Program (863program)of China under Grant No.2007AA03Z103,the National Natural Science Foundation of China under Grant Nos.50742007,50972014and 50872159,the Key Laboratory Foundation of Sonar Technology of China under Grant No.9140C24KF0901.References[1]Kozuka H,Kajimura M,Hirano T,Katayama K.J Sol –Gel Tech 2000;19:205–9.[2]Hu SH,Meng XJ,Wang GS,Sun JL,Li DX.J Cryst Grow 2004;264:307–11.[3]Lin P,Ren W,Wu XQ,Shi P,Yan X,Yao X.J Appl Phys 2007;102:084109.[4]Tanaka K,Kubota T,Sakabe Y.Sens Actuators A 2002;96:179–83.[5]Zhu BP,Wu DW,Zhou QF,Shi J,Shung KK.Appl Phys Lett 2008;93:012905.[6]Duan ZX,Yuan J,Zhao QL,Liu HM,Lin HB,Zhang WT,et al.Chin Phys Lett 2008;25:1472–5.[7]Li H,Yang ZP,Wei LL,Chang YF.Mater Res Bull 2009;44:638–43.[8]Wang GS,Deng Y,Xiang Y,Guo L.Adv Funct Mater 2008;18:2584–92.[9]Dang ZM,Xie D,Yu YF,Xu HP,Hou YD.Mater Chem Phys 2008;109:1–4.[10]Lin HB,Cao MS,Zhao QL,Shi XL,Wang DW,Wang FC.Script Mater 2008;59:780–3.[11]Cao MS,Zhou W,Shi XL,Chen YJ.Appl Phys Lett 2007;91:021912.[12]Wang ZL,Song JH.Science 2006;312:242–6.[13]Shi XL,Cao MS,Zhao YN,Song WL,Rong JL.Sci China Ser E-Tech Sci 2008;51:1433–8.[14]Zhang T,Wasa K,Zhang SY,Chen ZJ,Zhou FM,Zhang ZN,et al.Appl Phys Lett 2009;94:122909.[15]Nagarajan V,Jenkins IG,Alpay SP,Li H,Aggarwal S,Riba LS,et al.J Appl Phys 1999;86:595-02.[16]Long YZ,Li MM,Sui WM,Kong QS,Zhang L.Chin Phys B 2009;18:1221–6.[17]Wang GS,Remiens D,Dogheche E,Herdier R.J Am Ceram Soc 2006;89:3417–20.[18]Cheng YH,Chen XL,Wu K,Wu SN,Chen Y,Meng YM.J Appl Phys 2008;103:034111.[19]Sengupta SS,Park SM,Payne DA,Allen LH.J Appl Phys 1998;83:2291–6.[20]Xu F,Mckinstry T,Ren W,Xu BM,Xie ZL,Hemker KJ.J Appl Phys 2001;89:1336–48.Fig.4.Frequency dependences of dielectric constant (εr )and dielectric loss (tan δ)of ZnO w –PZT and 2-μm-thick PZT films.635Q.L.Zhao et al./Materials Letters 64(2010)632–635。

Preparation,structural and microwave dielectric properties of CaLa 4(Zr x Ti 1Àx )4O 15ceramicsCheng-Hsing Hsu ⇑,Ci-Jie HuangDepartment of Electrical Engineering,National United University,No.1Lien-Da,Kung-Ching Li,Miao-Li 36003,Taiwana r t i c l e i n f o Article history:Received 16August 2013Received in revised form 19October 2013Accepted 22October 2013Available online 31October 2013Keywords:SinteringDielectric propertiesCaLa 4(Zr x Ti 1Àx )4O 15ceramicsa b s t r a c tHexagonal-phase CaLa 4(Zr x Ti 1Àx )4O 15(x =0.01–0.1)with excellent microwave dielectric properties are candidate materials for microwave dielectric resonators.An excess of Zr 4+resulted in the formation of a secondary phase of CaLa 4Ti 5O 17,affecting the microwave dielectric properties of the CaLa 4(Zr x Ti 1Àx )4O 15ceramics.By increasing x from 0.01to 0.05,the dielectric constant and Q Âf value of the specimen could be increased from 45.8to a maximum of 46,and from 44,800GHz to a maximum of 47,500GHz at a sin-tering temperature of 1550°C,respectively.The zero temperature coefficient of the resonant frequency of the CaLa 4(Zr x Ti 1Àx )4O 15ceramics also improved with increasing Zr content.Ó2013Elsevier B.V.All rights reserved.1.IntroductionWith the great development in microwave low-power commu-nication modules,such as WiFi,RFID,and ZigBee communication systems,microwave ceramics with excellent dielectric characteris-tics that can reduce the dimensions of the microwave devices or improve the power-consumption of the microwave low-power communication modules have become quite important.Candidate microwave ceramics for such microwave communication applica-tions must have a high relative permittivity (e r >30),high dielec-tric Q Âf value (Q Âf >5000GHz),and temperature coefficient of resonant frequency close to zero (s f $0ppm/°C)[1,2].Several hex-agonal-phase ceramics (M,La)n Ti n À1O 3n (M =Ca,Sr,Ba;n =5,6),which belong to the cation-deficient perovskite A n B n À1O 3n system,where the M and La ions occupy the A sites and Ti ions occupy the B sites,have been reported to have excellent microwave dielectric properties [3,4].Among these,the CaLa 4Ti 4O 15composition has also been widely applied in microwave communication compo-nents because it has high dielectric constant (e r $41),high quality factor (Q Âf $50,000GHz),and small temperature coefficient of resonant frequency (s f $À25ppm/°C)[5].Several studies have been carried out to improve the microwave dielectric properties,such as the partial replacement of B-site and selection of a compensator in the ceramic systems [6,7].Huang et al.reported that the Mg 2(Ti 0.95Sn 0.05)O 4ceramics have the following values:e r =16.67,Q Âf =2,75,000GHz,and s f =À53.2ppm/°C;and that its dielectric constant and quality factorcan be improved with Sn 4+substitution [8].In addition,Tseng et al.also reported that substitution with Zr effectively improved the microwave dielectric properties of MgTiO 3ceramics,which exhibited the values e r =18.1,Q Âf =3,80,000GHz,and s f =À50ppm/°C [9].As mentioned earlier,it is interesting to study the influence of microwave dielectric properties on the Zr 4+substi-tutions in the B-sites in the CaLa 4Ti 4O 15materials because of the ionic radii of Zr (0.72nm)and Ti (0.605nm)are similar.In this study,the CaLa 4(Zr x Ti 1Àx )4O 15(x =0.01–0.1)ceramics were pre-pared by using conventional solid-state methods,and their micro-wave dielectric properties were investigated as a function of the Zr content.In addition,the correlation between the microstructure and microwave dielectric properties was also investigated.2.Experimental procedureThe CaLa 4(Zr x Ti 1Àx )4O 15(x =0.01–0.1)microwave dielectric ceramics systems were fabricated according to the stoichiometry and were synthesized by conven-tional solid-state methods from individual high-purity oxide powders (>99.9%)of CaCO 3,La 2O 3,ZrO 2,and TiO 2.The powders were ball-mixed in distilled water for 12h in a plastic bottle with agate balls.All wet mixtures were dried and calcined at 1250°C for 4h.The calcined powder with PVA as a binder was pressed into pel-lets with the dimensions of 11mm in diameter.These samples were sintered at temperatures of 1490–1570°C for 4h in air.The bulk densities of the sintered spec-imens were measured using the Archimedes method.X-ray diffraction (XRD,Siemens D5000)data of the bulk samples were collected using Cu K a radiation and a graphite monochromator in the 2h range of 20–60°.The microstructural observations and analysis of the sintered surface were performed using a scanning electron microscope (SEM,Philips XL40FEG,Eindhoven,The Neth-erlands).The densities of the sintered specimens were measured by the liquid Archimedes method.The e r and Q Âf values at microwave frequencies were mea-sured using the Hakki–Coleman dielectric resonator method,as modified and im-proved by Courtney [10,11].The dielectric resonator was positioned between two brass plates to acted as a parallel cavity.The microwave dielectric properties of0925-8388/$-see front matter Ó2013Elsevier B.V.All rights reserved./10.1016/j.jallcom.2013.10.168Corresponding author.Tel.:+88637381401;fax:+88637327887.E-mail address:hsuch@.tw (C.-H.Hsu).dielectric resonators were measured with using a network analyzer.The tempera-ture coefficient of the resonant frequency(s f)was measured using the same tech-nique as that used to measure the quality factor.The test cavity was placed over a thermostat and the temperature range was from+25°C to+80°C.The s f(ppm/°C)was calculated from the change in resonant frequency(D f).s f¼f2Àf121ð1Þwhere f1is the resonant frequency at T1and f2is the resonant frequency at T2. 3.Results and discussionsFigs.1and2show the XRD patterns of the CaLa4(Zr x Ti1Àx)4O15 (x=0.01–0.1)ceramics system sintered at1490–1570°C for4h. All the diffraction peaks in the XRD patterns were similar and can be indexed with a hexagonal structure that belonged to the P 3m space group at various Zr contents and sintering tempera-tures[4].In addition,the CaLa4Ti5O17phase was also detected in the XRD patterns and the peak index of the secondary phase of CaLa4Ti5O17increased with increasing Zr content and sintering temperature.There were small shifts of diffraction peaks toward the lower2h angle with increasing Zr content indicating that the lattice parameters of the CaLa4(Zr x Ti1Àx)4O15(x=0.01–0.1)ceram-ics increased.The microstructural photographs of CaLa4(Zr x Ti1Àx)4O15 (x=0.01–0.1)ceramics are shown in Fig.3.The porosity decreased with increasing x content and sintering temperature,and no pore was observed at a temperature of1550°C for x=0.05owing to grain growth.However,degradation in grain uniformity and en-larged grain growth started to appear for CaLa4(Zr x Ti1Àx)4O15Fig.2.However,very high sintering temperature would cause en-larged grain growth,resulting in a decrease in density.The apparent density and its corresponding theoretical density increased from5.51 (94%TD)to5.66g/cm3(96.6%TD)as the sintering temperature was increased from1490to1550°C for the CaLa4(Zr0.05Ti0.95)4O15ceram-ics when sintered for4h.The results in density may directly affect the dielectric properties.The dielectric constants of the CaLa4(Zr x Ti1Àx)4O15(x=0.01–0.1) ceramic system are shown in Fig.5.The variation in the dielectric constant with various sintering temperatures and x contents was consistent with that of the relative density,implying that the dielectric constant was mainly controlled by the density of the specimen.The e r values of x=0.05were measured from45.8to 46with increasing sintering temperature,and thereafter it decreased.In addition,an increase in the e r value with increasing sintering temperature was in agreement with that of the density in the composition range of0.01–0.1.On the other hand,an increase in e r value occured and was attributed to the secondary phases of CaLa4Ti5O17(53.7)with high dielectric constant; however,a decreased e r value was detected at higher Zr content (x>0.05)due to enlarged grain growth[3].At a sintering temper-ature of1550°C for4h,a e r value of46was obtained for CaLa4(Zr0.05Ti0.95)4O15.Fig.6shows the QÂf value of the CaLa4(Zr x Ti1Àx)4O15 (x=0.01–0.1)ceramic system,and it exhibits the same trend with the density.Ahn et al.reported that the sintering temper-ature affects the ordering of B-sites[12].On the other hand,the microwave dielectric loss is not only caused by the lattice vibra-tional modes,but also by the pores,grain morphology,and sec-Fig.1.XRD patterns of the CaLa4(Zr0.05Ti0.95)4O15ceramics with various sintering temperatures.46 C.-H.Hsu,C.-J.Huang/Journal of Alloys and Compounds587(2014)45–49Fig.2.XRD patterns of the CaLa4(Zr x Ti1Àx)4O15(x=0.01–0.1)ceramics sintered at1550°C.SEM micrographs of the CaLa4(Zr x Ti1Àx)4O15ceramics with various x contents and sintering temperatures:(a)x=0.05,1490°C;(b)x=0.05,1510°C;(c) (d)x=0.01,1550°C;(e)x=0.03,1550°C;(f)x=0.05,1550°C;(g)x=0.07,1550°C;(h)x=0.1,1550°C;(i)x=0.05,1570°C.The s f values of the CaLa4(Zr x Ti1Àx)4O15(x=0.01–0.1)ceramic system was shown in Fig.7.The temperature coefficient of reso-nant frequency was related to the microstructure,composition, and secondary phase of the material.The s f values of the CaLa4(Zr x-Ti1Àx)4O15(x=0.01–0.1)ceramic system varied fromÀ11.3to À9.5ppm/°C at a sintering temperature of1550°C.The variation in the s f value of CaLa4(Zr x Ti1Àx)4O15(x=0.01–0.1)ceramics can be explained by extrinsic and intrinsic factors.Extrinsic factor is believed to the positive s f value of CaLa4Ti5O17(+20ppm/°C),and the s f value can be tuned by the secondary phases.3On the other hand,the intrinsic factor for the variation in the s f value could be reasonably that the distortion of microstructure,crystal lattice,de-fect,and phase transition,which could be responsible for the var-iation in the s f value[14].Significant changes were observed in the s f value with different Zr contents,which implies that the s f value was sensitive to the Zr content.4.ConclusionIn the present study,microwave dielectric properties and sin-tering behavior of Zr substituted for Ti to form CaLa4(Zr x Ti1Àx)4O15 (x=0.01–0.1)solid solution ceramic system were investigated.The CaLa4Ti5O17was detected as the secondary phases at various sin-tering temperatures and Zr contents.The dielectric constant and temperature coefficient of the resonant frequency of CaLa4(Zr x Ti1-Àx)4O15ceramics,ranging from45.8to46and fromÀ11.3to À9.5ppm/°C,were slightly improved,when compared with those of pure CaLa4Ti4O15ceramics.The QÂf value also remained an equivalent level of near50,000GHz.The present results indicated that the CaLa4(Zr x Ti1Àx)4O15ceramics have some better advantages with respect to microwave dielectric component applications than pure CaLa4Ti4O15ceramics.A dielectric constant(e r)of46,a high QÂf value of47,500GHz,and a s f ofÀ9.5ppm/°C were developed for CaLa4(Zr0.05Ti0.95)4O15ceramics sintered at1550°C for4h.AcknowledgmentThis work was sponsored by the National Science Council of the Republic of China under Grant NSC101-2622-E-239-004-CC3.Table1EDS analysis result of CaLa4Ti5O17ceramics presented in Fig.2(i).Spot Atom(%)Ca K La L Zr L Ti K O KA 4.1718.330.9116.1360.4648 C.-H.Hsu,C.-J.Huang/Journal of Alloys and Compounds587(2014)45–49References[1]I.M.Reaney,D.Iddles,J.Am.Ceram.Soc.89(2006)2063–2072.[2]J.Chameswary,S.Thomas,M.T.Sebastian,J.Am.Ceram.Soc.93(2010)1863–1865.[3]I.N.Jawahar,N.I.Santha,M.T.Sebastian,J.Mater.Res.17(2002)3084–3089.[4]J.Pei,Z.X.Yue,F.Zhao,Z.L.Gui,L.T.Li,J.Alloys Comp.459(2008)390–394.[5]Y.Tohdo,K.Kakimoto,H.Ohsato,H.Yamada,T.Okawa,J.Eur.Ceram.Soc.26(2006)2033–2043.[6]C.F.Tseng,C.H.Hsu,J.Am.Ceram.Soc.92(2009)1149–1152.[7]C.H.Hsu,H.A.Ho,Mater.Lett.64(2010)396–398.[8]C.L.Huang,J.Y.Chen,J.Am.Ceram.Soc.92(2009)2237–2241.[9]C.F.Tseng,J.Alloys Comp.509(2011)9447–9450.[10]W.Hakki,P.D.Coleman,IEEE Trans.Microwave Theory Technol.8(1960)402–410.[11]W.E.Courtney,IEEE Trans.Microwave Theory Technol.18(1970)476–485.[12]C.W.Ahn,S.N.Nahm,Y.S.Lim,W.Choi,H.M.Park,H.J.Lee,Jpn.J.Appl.Phys.41(2002)5277–5280.[13]B.D.Silverman,Phys.Rev.125(1962)1921–1930.[14]X.Y.Chen,S.X.Bai,W.J.Zhang,J.Alloys Comp.541(2012)132–136.C.-H.Hsu,C.-J.Huang/Journal of Alloys and Compounds587(2014)45–4949。

JOURNAL OF RARE EARTHS, Vol. 30, No. 3, Mar. 2012, P. 202Foundation item: Project supported by National Basic Research Program of China (2007CB935502, 2010CB327704), National Natural Science F oundation ofChina (NSFC 50872131, 60977013, 20921002)Corresponding author: JIA Peiyun (E-mail: jiapeiyun@; Tel.: +86-451-51688228) DOI: 10.1016/S1002-0721(12)60023-4Molten salt synthesis and luminescent properties of YVO 4:Eu nanocrystalline phosphorsWANG Fang (⥟ 㢇)1, LIU Chenglu ( 䴆)1, ZHOU Zhiqiang ( )1, JIA Peiyun (䌒Խѥ)1, LIN Jun ( )2(1. School of Sciences, Northeast Forestry University, Harbin 150040, China; 2. State Key Laboratory of Rare Earth Resource Utilization, Changchun Institute of Applied Chemistry, Chinese Academy of Sciences, Changchun 130022, China) Received 20 July 2011; revised 21 August 2011Abstract: YVO 4:Eu nanocrystalline phosphors were successfully prepared at 400 ºC in equal moles of NaNO 3 and KNO 3 molten salts. NaOH concentration and annealing temperature played important roles in phase purity and crystallinity of the nanocrystallines, and the optimum NaOH concentration and annealing temperature were 6:40 and 400 ºC, respectively. The nanocrystallines were well crystallized with a cubic morphology in an average grain size of 18 nm. Upon excitation of the vanadate groups at 314 nm, YVO 4:Eu nanocrystallines exhibited the characteristic emission of Eu 3+, which indicated that there was an energy transfer from vanadate groups to Eu 3+. Moreover, the influence of superficial effect, especially the dangling bonds on the structure and luminescent properties of the nanocrystallines was discussed in detail. Keywords:nanocrystalline; phosphor; luminescence; molten salt; YVO 4; rare earthsAs an excellent luminescent material, YVO 4:Eu has been used as a red phosphor of cathode ray tube television since it was discovered in 1964[1]. Recently, YVO 4:Eu nanophos-phors have attracted great attention again due to their poten-tial application in biological fluorescence labeling and gas sensing [2,3]. A good many strategies based on wet chemistry method have been used to synthesize YVO 4:Eu nanophos-phors [4–9]. However, in these methods, organic dispersing agents were utilized to prevent the agglomeration of the nanophosphors. The organic compounds adsorbed, chemically or physically, on the surface of the nanophosphors prevent the research of interfacial effect on the luminescence properties and some potential applications of the nanophosphors.Molten salts have been widely used in heat transfer me-dium, non-ferrous metal synthesis, fuel cell and molten salt reactor. F ree from organic dispersing agents, molten salts were developed to prepare nanocrystallines owing to their cost-effectiveness and environmental friendliness. Liu and his co-workers prepared a series of single-crystalline com-plex oxide nanocrystallines of various structural and mor-phology in molten NaOH, KOH or their mixtures [10]. In this paper, we developed a single-step molten salt method to synthesize YVO 4:Eu nanocrystallines in equal moles of NaNO 3 and KNO 3 molten salts, by which single-phase, well crystallized YVO 4:Eu nanocrystallines were successfully prepared.1 Experimental1.1 Synthesis of YVO 4:Eu nanocrystallinesNaNO 3, KNO 3, NH 4VO 3, NaOH (analytical reagent, Aladdin Reagent (China) Co., Ltd.), Y 2O 3 and Eu 2O 3 (99.99%, Shanghai Yuelong New Materials Co., Ltd.) were used as the starting materials. In a typical synthesis of Y 0.95Eu 0.05VO 4 nanocrystallines, 107.3 mg Y 2O 3 and 8.8 mg Eu 2O 3 were dissolved in dilute HNO 3 solution under stirring, the pH of the solution was adjusted to between 5.0 and 6.0 with NaOH dilute solution, then the solution was dried on water bath at 80 ºC for 6 h to obtain Y(NO 3)3 and Eu(NO 3)3 mixtures. As-prepared mixtures were added with 117.0 mg NH 4VO 3, 240.0 mg NaOH, 1699.8 mg NaNO 3 and 2022.0 mg KNO 3, and fully ground in an agate mortar with suitable amount of ethanol. Subsequently, the powders were trans-ferred into a corundum crucible, heated from room tempera-ture to 400 ºC, kept for 12 h and cooled down to room tem-perature naturally. F inally, the samples were fully washed with deionized water until the nitrate group could not be de-tected and dried in a vacuum drying oven at 150 ºC for 2 h for subsequent characterization. For convenience, the NaOH concentration is defined as the molar ratio of NaOH to the sum of NaNO 3 and KNO 3 for synthesis of 1 mmol YVO 4:Eu nanocrystallines. In the above case, the NaOH concentration was 6:40.1.2 CharacterizationX-ray diffraction (XRD) measurements were performed on a Rigaku-Dmax 2500 diffractometer using Cu KĮ radia-tion (Ȝ=0.15405 nm). The morphology and grain size of theWANG Fang et al.,Molten salt synthesis and luminescent properties of YVO 4:Eu nanocrystalline phosphors 203samples were inspected on a JEOL 2010 transmission elec-tron microscope (TEM) operating at 200 kV. The excitation and emission spectra were taken on a Hitachi F-7000 spec-trofluorimeter equipped with a 150 W xenon lamp as the ex-citation source. All the measurements were performed at room temperature.2Results and discussion2.1 X RD analysisF ig. 1 displays the XRD patterns of Y 0.95Eu 0.05VO 4 pre-pared at different NaOH concentrations and temperatures in equal moles of NaNO 3 and KNO 3 molten salts. Fig. 1(1) to Fig. 1(5) provide the patterns of Y 0.95Eu 0.05VO 4 prepared at 250 ºC and NaOH concentration from 0:40 to 8:40. It can be found from these patterns that the optimum NaOH concen-tration for preparing Y 0.95Eu 0.05VO 4 at 250 ºC is 6:40. This optimum NaOH concentration is maybe determined by the chemical properties of the reaction precursors. As we know, a high NaOH concentration can effectively transfer NH 4VO 3 into NaVO 4 which is, of course, in favor of the formation of YVO 4:Eu; on the other hand, NaOH can also transform Y 3+ (or Eu 3+) into Y(OH)3(or Eu(OH)3) which is disadvanta-geous to the formation of YVO 4:Eu. So a mediate NaOH concentration (6:40) is beneficial to the formation of YVO 4:Eu. At NaOH concentration of 6:40, patterns of Y 0.95Eu 0.05VO 4 prepared at different annealing temperatures (Fig. 1(4) and Fig. 1(6) to Fig. 1(9)) indicate that the inten-sity of the patterns increases with the elevation of annealing temperature, reaches a maximum at 400 ºC, and then de-creases. Obviously, the optimum annealing temperature is 400 ºC. The pattern of the sample prepared at 400 ºC and NaOH concentration of 6:40 matches well with JCPDS NO. 13-0741 and have the highest intensity among all the patterns. Therefore, the samples characterized subsequently were allsynthesized at 400 ºC and NaOH concentration of 6:40.Fig. 1 XRD patterns of Y 0.95Eu 0.05VO 4 nanocrystallines prepared at250 ºC and NaOH concentration of 0:40 (1), 2:40 (2), 4:40 (3), 6:40 (4), 8:40 (5) and at 300 ºC (6), 350 ºC (7), 400 ºC (8), 450 ºC (9) and NaOH concentration of 6:40; and JCPDS NO. 17-0341 (* indicates the diffraction peaks of unknown phases)The crystallite size of the Y 0.95Eu 0.05VO 4 was calculated based on Scherer’s formula according to the diffraction peak of (200) plane [11] and the crystallite size is about 18 nm, which will be further verified by the TEM micrographs shown in section 2.2.The cell parameters of Y 0.95Eu 0.05VO 4 nanocrystallines (a =0.7117 nm, c =0.6280 nm and cell volume=0.3181 nm 3) calculated by the program Jade 5.0 are compared with that of JCPDS No. 17-0341 (a =0.712 nm, c =0.629 nm and cell vol-ume=0.3189 nm 3). It can be found that the cell volume of Y 0.95Eu 0.05VO 4 nanocrystallines is smaller than that of JCPDS No. 17-0341. However, the radius of Eu 3+ (0.112 nm) is slightly larger than that of Y 3+ (0.108 nm), so the substitu-tion of Eu 3+ for Y 3+ should enlarge the cell volume [12]. This inconsistency is caused by the high surface/volume ratio of the nanocrystallines. It is well known that the decrease of grain size will increase the surface/volume ratio and the dan-gling bonds on the surface simultaneously. In Y 0.95Eu 0.05VO 4 nanocrystallines, the original VíOíY(Eu)íOíV in bulk will be broken into VíOíƑ (Ƒ indicates the surface of Y 0.95Eu 0.05VO 4 nanocrystallines) and OíY(Eu)íƑ on the surface. For VíOíƑ dangling bonds, the absence of absorp-tion from Y 3+(Eu 3+) will lead to the contraction of VíO bond length; for OíY(Eu)íƑ ones, the decrease of coordinate number (CN) of Y(Eu)3+ will as well result in the contraction of YíO bond length [13]. Since the enlargement of Eu 3+ can-not compensate the bond length contraction of VíOíƑ and OíY(Eu)íƑ dangling bonds, the cell volume will decrease naturally.2.2 TEM analysisTEM was used to characterize the size, morphology and structure of the nanocrystallines. Fig. 2(a) provides a repre-sentative TEM micrograph of Y 0.95Eu 0.05VO 4 nanocrystalli-nes. It can be found that the well-dispersed nanocrystallines with a cubic morphology are uniform in size and have an average grain size of about 18 nm. This result is consistent with the crystallite size calculated by Scherer’s formula. High resolution (HR) TEM (F ig. 2(b)) characterization re-veals the highly crystalline nature of the nanocrystallines. Typical HRTEM micrograph of Y 0.95Eu 0.05VO 4 nanocrystal-lines exhibits an interplanar spacing of 0.35 nm, which cor-responds to the (200) planes of zircon-type YVO 4 (JCPDSNo. 17-0341). It coincides with the results of XRD.F ig. 2 TEM micrograph (a) and HRTEM micrograph (b) ofY 0.95Eu 0.05VO 4 nanocrystallines prepared at 400 ºC and NaOH concentration of 6:40204 JOURNAL OF RARE EARTHS, Vol. 30, No. 3, Mar. 20122.3 Photoluminescence (PL) analysisF ig. 3 shows the excitation and emission spectra of Y 0.95Eu 0.05VO 4 nanocrystallines. The excitation spectrum (Fig. 3(a)) consists of a strong excitation band with a maxi-mum at 314 nm and several weak lines at 364, 384, 397, 418 and 467 nm. The strong excitation band is attributed to the charge transfer transition in VO 43– groups; the weak lines originate from the f-f transitions within 4f 6 configuration of Eu 3+. Upon excitation of the vanadate group at 314 nm, the emission spectrum (F ig. 3(b)) exhibits the characteristic emission of Eu 3+ at 540, 596, 621, 653, 700 and 706 nm, dominated by the 5D 0ĺ7F 2 transition at 621 nm. The domi-nation of 5D 0ĺ7F 2 emission is in accordance with the sym-metry properties of sites that Eu 3+ adopts (D 2d )[14]. The emis-sion from the vanadate group can be neglected compared with the emission of Eu 3+, which indicates that the energy transfer from the vanadate group to Eu 3+ ions is very effi-cient [15]. The luminescent properties of YVO 4:Eu nanocrys-tallines are compared with their bulk in the same composi-tion as shown in Ref. [16]. In the excitation spectrum of Y 0.95Eu 0.05VO 4 nanocrystallines, the location of V–O CTB shows a evident blue shift compared with the bulk Y 0.95Eu 0.05VO 4. F rom the viewpoint of coordination field theory, the V–O CTB is ascribed to the transition from the bonding 1A 1 (1T 2) to antibonding 1E (1T 2). The contraction of dangling VíOíƑ bonds will strengthen the V–O bond and, thus, enlarge the energy gap between bonding 1A 1(1T 2) and antibonding 1E(1T 2) in VO 43í group, which will make the V–O CTB shift to high energy. The emission spectrum of YVO 4:Eu nanocrystallines is similar with their bulk coun-terparts. This is because the f-f transition within 4f 6 configu-ration is hardly influenced by the coordination environment.F ig. 3 Excitation (a) (Ȝem =621 nm) and emission (b) spectra (Ȝex =314 nm) of Y 0.95Eu 0.05VO 4 nanocrystallines annealed at 400 ºC and NaOH concentration of 6:403ConclusionsYVO 4:Eu nanocrystallines were successfully synthesized at 400 ºC and NaOH concentration of 6:40 in equal moles of NaNO 3 and KNO 3 molten salts. Excitation into V–O CTB, YVO 4:Eu 3+ nanocrystallines showed the characteristic emis-sion of Eu 3+ ions, which indicated that there existed an energy transfer from the V–O CTB to Eu 3+. The high surface/volume ratio of the YVO 4:Eu 3+ nanocrystallines resulted in the de-crease of cell volume and the blue shift of V–O CTB.Acknowledgement: We also thank financial support of the Post-graduate Sci-tech Innovation Program of Northeast F orestry Uni-versity (Gram10).References:[1] Levine A K, Palilla F C. A new, highly efficient red-emitting cathodoluminescenent phosphor (YVO 4:Eu) for color televi-sion. Appl. Phys. Lett., 1964, 5: 118.[2] Kang J, Zhang X Y, Sun L D, Zhang X X. Bioconjugation of functionalized fluorescent YVO 4:Eu nanocrystals with BSA for immunoassay. Talanta , 2007, 71: 1186.[3] Zhou Q, Zhang L C, Fan H Y, Wu L, Lv Y. An ethanol gas sensor using energy transfer cataluminescence on nanosized YVO 4:Eu 3+ surface. Sens. Actuators B , 2010, 144: 192.[4] Riwotzki K, Haase M. Wet-chemical synthesis of doped colloidal nanoparticles: YVO 4:Ln (Ln=Eu, Sm, Dy). J. Phys. Chem. B , 1998, 102: 10129.[5] Huignard A, Buissette V, Laurent G, Gacoin T, Boilot J P. Synthesis and characterizations of YVO 4:Eu colloids. Chem. Mater., 2002, 14: 2264.[6] Kaowphong S, Nakashima K, Petrykin V, Thongtem S, Kakihana M. Methanol-water system for solvothermal synthesis of YVO 4:Eu with high photoluminescent intensity. J. Am. Ceram. Soc., 2009, 92(S1): S16.[7] He F, Yang P P, Niu N, Wang W X, Gai S L, Wang D, Lin J. Hydrothermal synthesis and luminescent properties of YVO 4:Ln 3+ (Ln=Eu, Dy, and Sm) microspheres. J. Colloid. Interf. Sci., 2010, 343: 71.[8] Wu X C, Tao Y R, Mao C J, Liu D J, Mao Y Q. In situ hydro-thermal synthesis of YVO 4 nanorods and microtubes using (NH 4)0.5V 2O 5 nanowires templates. J. Cryst. Growth., 2006, 290: 207.[9] Liu Y, Ma J F, Dai C H, Song Z W, Sun Y, Fang J R, Gao C, Zhao J G. Low-temperature synthesis of YVO 4 nanoparticles and their photocatalytic activity. J. Am. Ceram. Soc., 2009, 92: 2791. [10] Liu H, Hu C G, Wang Z L. Composite-hydroxide-mediatedapproach for the synthesis of nanostructures of complex func-tional-oxides. Nano Lett., 2006, 6: 1535.[11] Glas J E, Omnell K. Studies on the ultrastructure of dentalenamel: 1. Size and shape of the apatite crystallites as deduced from X-ray diffraction data. J. Ultra. Res., 1960, 3: 334.[12] Shannon R D, Revised effective ionic radii and systematicstudies of interatomic distances in halides and chalcogenides. Acta Cryst. A , 1976, 32: 751.[13] Sun C Q. Size dependence of nanostructures: impact of bondorder deficiency. Prog. Solid State Chem., 2007, 35: 1.[14] Yu M, Lin J, Wang Z, Fu J, Wang S, Zhang H J, Han J C.abrication, patterning, and optical properties of nano- crystalline YVO 4:A(A=Eu 3+, Dy 3+, Sm 3+, Er 3+) phosphors films via sol-gel soft lithography. Chem. Mater., 2002, 14: 2224. [15] Hsu C, Powell R C. Energy transfer in europium doped yttriumvanadate crystals. J. Lumin., 1975, 10: 273.[16] Rambabu U, Amalnerkar D P, Kale B B, Buddhudu S.F luorescence spectra of Eu 3+-doped LnVO 4 (Ln=La and Y) powder phosphors. Mater. Res. Bull., 2000, 35: 929.。

Short CommunicationEnhanced liquid phase catalytic hydrodechlorination of 2,4-dichlorophenol over mesoporous carbon supported Pd catalystsYun Shao,Zhaoyi Xu,Haiqin Wan ⁎,Yuqiu Wan,Huan Chen,Shourong Zheng ⁎,Dongqiang ZhuState Key Laboratory of Pollution Control and Resource Reuse,School of the Environment,Nanjing University,Nanjing 210093,PR Chinaa b s t r a c ta r t i c l e i n f o Article history:Received 9March 2011Received in revised form 6May 2011Accepted 9May 2011Available online 17May 2011Keywords:Catalytic hydrodechlorination of 2,4-dichlorophenol Supported Pd catalyst Activated carbon Mesoporous carbonSupported Pd catalysts on ordered mesoporous carbon (OMC)and activated carbon (AC)were prepared and their catalytic behavior for the liquid phase catalytic hydrodechlorination (HDC)of 2,4-dichlorophenol was investigated.In comparison with Pd/AC,Pd particles of Pd/OMC were effectively con fined in the mesopores of OMC,resulting in high Pd dispersion and Pd 2+content.Accordingly,Pd/OMC exhibited higher catalytic activity than Pd/AC.Moreover,increasing catalyst reduction temperature lowered the catalytic activities and favored stepwise HDC of 2,4-dichlorophenol.©2011Elsevier B.V.All rights reserved.1.IntroductionChlorinated phenols are important raw materials and intermediates in chemical industry.However,the wide use of chlorinated phenols has resulted in a large amount of chlorinated phenols-bearing wastewater,from which the effective removal of chlorinated phenols is legislatively obligatory due to their strong toxicity and low biodegradability [1].Liquid phase catalytic hydrodechlorination (HDC)over supported noble metal catalysts provides a nondestructive approach for the detoxi fica-tion of chlorinated phenols and recovery of valuable chemicals [2].Supported Pd,Pt,and Rh catalysts are generally used in the liquid phase HDC,where supported Pd catalyst is the most active one [3].Additionally,activated carbon (AC)is usually used as catalyst support due to its large surface areas and high stability under acidic and alkaline conditions.However,AC primarily consists of irregular shaped micropore with a wide pore size distribution,incapable of hosting nano-scale metal particle.Such shortcomings can be avoided by using ordered mesoporous carbon (OMC)as the support.Because OMC is composed of ordered mesopore with a narrow pore distribution,nano-scale metal particle can be con fined in the mesopores [4].Hence,it is reasonable to hypothesize that OMC supported Pd catalyst may exhibit different catalytic behavior from AC supported Pd catalyst.However,few studies have been conducted on the catalytic behavior of OMC supported Pd catalyst for the catalytic HDC of chlorinated phenols thus far.In this study,OMC and AC supported Pd catalysts were prepared and the liquid phase catalytic HDC of 2,4-dichlorophenol (2,4-DCP)was investigated.The results showed that Pd/OMC displayed superior catalytic activity to Pd/AC,highlighting the potential of using Pd/OMC as a more effective catalyst for the catalytic HDC of chlorinated phenol.2.Experimental 2.1.Catalyst preparationOrdered mesoporous silica SBA-15was prepared using a triblock copolymer,EO 20PO 70EO 20(Pluronic P123,Aldrich),as the structure directing agent and tetraethoxysilane (TEOS,98%,Shanghai Chemical Co.)as the silica source [5,6].OMC was synthesized using calcined SBA-15as the hard template [6,7].Commercial AC (Filtrasorb-300,Calgon Carbon Co.)was included for comparison purpose.To avoid possible intraparticle diffusion,the supports were ground to pass through a 400-mesh sieve (b 37μm)prior to catalyst preparation [8].Supported Pd catalysts were prepared using the incipient wetness impregnation method with H 2PdCl 4as the Pd precursor.The Pd loading amount was preset to be about 5wt.%,which was generally adopted for AC supported catalysts [8,9].The samples were dried at 80°C under vacuum for 4h,and then reduced in a H 2flow (40mL/min)at 200,300or 400°C for 2h.The resultant catalyst was referred to as Pd/OMC-X or Pd/AC-X respectively,where X is the reduction temperature.2.2.Catalyst characterizationX-ray diffraction (XRD)patterns of the samples were obtained from a Rigaku D/max-RA powder diffraction-meter.Transmission electronCatalysis Communications 12(2011)1405–1409⁎Corresponding authors.Tel.:+862589680373;fax:+862583707304.E-mail addresses:wanhq@ (H.Wan),srzheng@ (S.Zheng).1566-7367/$–see front matter ©2011Elsevier B.V.All rights reserved.doi:10.1016/j.catcom.2011.05.007Contents lists available at ScienceDirectCatalysis Communicationsj o u r n a l h o m e p a g e :w ww.e l s ev i e r.c o m /l o c a t e /c a tc o mmicroscopy(TEM)images of the catalysts were obtained from a JEM-2100transmission electron microscope.Brunauer–Emmett–Teller (BET)specific surface areas of the supports and catalysts were measured using the nitrogen adsorption method on a Micromeritics ASAP2020 instrument.X-ray photoelectron spectroscopy(XPS)was performed on a Thermo ESCALAB250instrument equipped with a monochromatized Al Kαexcitation source(hν=1486.6eV).The C1s peak(284.6eV)was used for the calibration of binding energy values.The Pd contents of Pd/ OMC and Pd/AC were determined to be5.60and5.82wt.%,respectively, by X-rayfluorescence(ARL-9800).2.3.Liquid phase catalytic HDC of2,4-DCPCatalyst activity for the liquid phase catalytic HDC of2,4-DCP was evaluated using the batch reaction approach at20°C under atmospheric pressure of hydrogen.Catalysis tests were conducted in a500-mL of four-neckedflask reactor with a sample port,a pH-stat,H2inlet and a condenser.The reaction temperature was stabilized at20±0.5°C with a water-bath(SDC-6,scientz Co.,China).Briefly,50mg of catalyst was added into the reactor containing200mL of3.5mmol L−12,4-DCP aqueous solution.In order to eliminate HCl poisoning,the pH of2,4-DCP solution was pre-adjusted to12.0[10].After purging with a N2flow (250mL min−1)under stirring(1200rpm)for30min,the N2flow was switched into a H2flow(250mL min−1)during the reaction process. Samples were taken at different time intervals and catalyst particles were removed byfiltration.Thefiltrate was neutralized using2.0mol L−1HCl and the concentration of reactant,intermediate and product in the filtrate was analyzed by a high performance liquid chromatography with an ultraviolet(UV)detector at270nm and a4.6×150mm HC-C18 column(Agilent).The mobile phase consisted of60%CH3CN and40% water(v:v).The results of two separate runs of the HDC of2,4-DCP showed high data reproducibility(see Fig.1S,supporting information).3.Results and discussion3.1.Catalyst characterizationThe small-angle XRD patterns of the samples are compared in Fig.2S,supporting information.For OMC and Pd/OMC catalysts, diffraction peaks with2θat1.01°,1.76°and2.02°,indexed as(100), (110)and(200)diffractions,were observed,reflecting the presence of typical hexagonal mesostructured phase[5,11].N2adsorption results are shown in Fig.3S,supporting information and resultant parameters are listed in Table1.For AC,N2adsorption amount mainly increased at low relative pressure(p/p0),characteristic of the microporous nature of AC.In contrast,typical capillary condensation was observed within a relative pressure range of0.40–0.65for OMC and Pd/OMC,reflecting the presence of mesopores.The pore size distribution profile of AC showed that AC was mainly composed of micropores dominated by pore sizes smaller than1.5nm,while OMC exhibited a narrow pore size distribution with the most probable pore diameter centered at4.2nm.Accordingly,the Pd/OMC catalysts had the most probable pore diameters around4.3nm,similar to that of OMC.TEM images and histograms of Pd particle size distributions of the catalysts are compared in Fig.1.Ordered mesostructure and Pd particles were clearly identified in Pd/OMC.Moreover,much smaller Pd particles and narrower Pd particle distributions of Pd/OMC were observed as compared to Pd/AC(see Fig.1b),likely due to the effective confinement of Pd particles in OMC pores.For Pd/AC and Pd/OMC increasing reduction temperature led to broadened Pd particle size distribution toward larger particle size.The average Pd particle sizes of the catalysts were further quantified based on a surface area weighted diameter[10]:s=∑n i d3i=∑n i d2i:ð1ÞWhere n i is the number of counted Pd particles with diameter of d i and the total number of particles∑n iðÞis larger than150.As shown in Table1,the average Pd particle sizes of Pd/OMC and Pd/AC increased with the reduction temperature,indicative of particle aggregation due to the high mobility of metal particle on carbona-ceous material surface[12].At given reduction temperature,the average Pd particle size of Pd/AC was larger than Pd/OMC,confirming the confinement effect of mesopore.The surface atomic ratios of Pd/C and O/C based on XPS analysis are summarized in Table2.At similar reduction temperature,Pd/AC had higher surface atomic contents of Pd and O than Pd/OMC.Given smaller Pd particles,the lower surface Pd contents of Pd/OMC than Pd/ AC suggest that Pd particles are predominantly located in the mesopores of OMC,but on the external surface of AC.It is understandable that due to its microporous nature AC is incapable of hosting nano-scale Pd particles in the micropores.Alternatively, OMC consists of ordered mesopore with the most probable diameter of4.2nm,resulting in effective confinement of Pd particles in the mesopore.It should be emphasized that high surface oxygen containing functionality of carbonaceous support favors Pd dispersion [13].Given lower oxygen containing functionality of OMC the higher Pd dispersion of Pd/OMC than Pd/AC again confirms the confinement effect of OMC,wherein oxygen containing functionality of OMC is fully accessible,but oxygen containing functionality on external surface of AC is available for nano-scale Pd particles.XPS spectra of the catalysts in the Pd3d5/2region are compiled in Fig.2.For Pd/OMC and Pd/AC,the binding energies of Pd3d5/2varied with catalyst reduction temperature,implying the coexistence of metallic and oxidized Pd.To quantify the contents of metallic and oxidized Pd,the XPS profiles were deconvoluted and thefitting parameters are listed in Table2.For Pd/OMC increasing reduction temperature lowered Pd2+contents.Similar trend but more marked decrease of Pd2+content with the reduction temperature was observed for Pd/AC.Upon H2reduction treatment,Pd2+is partially retained due to the stabilization of Pd2+by trace chlorine fromTable1Properties of the supports and catalysts reduced at200,300and400°C.Sample BET surface area(m2g−1)Micropore volume(cm3g−1)Mesoporous volume(cm3g−1)Pore volume(cm3g−1)Pore diameter(nm)Average Pd particle size(nm)OMC9910.1 1.2 1.3 4.2–AC9810.40.10.5––Pd/AC-200––––– 3.8 Pd/AC-300––––– 4.7 Pd/AC-400––––– 5.4 Pd/OMC-2009300.1 1.1 1.2 4.4 2.4 Pd/OMC-3008640.1 1.0 1.1 4.3 2.6 Pd/OMC-4008660.1 1.0 1.1 4.3 3.5 1406Y.Shao et al./Catalysis Communications12(2011)1405–1409palladium precursor and strong interaction between Pd 2+and oxygen containing groups from support surface [14,15].Despite of the lower oxygen content of OMC the oxygen containing functionality of OMC isfully accessible.Moreover,stronger metal-support interaction can be expected for Pd/OMC due to its smaller Pd particle size [16],resulting in more effective stabilization of Pd 2+and higher Pd 2+content in Pd/OMC as compared to Pd/AC.3.2.Liquid phase catalytic HDC of 2,4-DCPPrior to catalyst activity evaluation,it is necessary to exclude the in fluence of mass-transfer limitation.Hence the initial catalytic activities of Pd/OMC-300and Pd/AC-300with varied catalyst dosages were compared,where the initial activity was calculated based on a first-order rate law at 2,4-DCP conversion lower than 25%.The results are presented in Fig.4S,supporting information.For both catalysts,initial HDC rates were proportional to the mass of catalyst,while the initial activities remained nearly constant after catalyst massR e l a t i v e a m o u n t (%)Particle size (nm)Pd/OMC-300Pd/OMC-400Pd/OMC-200Pd/AC-200Pd/AC-300Pd/AC-400Particle size (nm)R e l a t i v e a m o u n t (%)baFig.1.(a)TEM images and (b)histograms of Pd particle size distributions of the catalysts.Table 2Binding energies of surface palladium species and surface atomic ratios.SampleBinding energy (eV)Pd 2+/(Pd 2++Pd 0)Pd/C (×10−3)O/C(×10−2)Pd 0Pd 2+Pd/AC-200335.7337.50.457.969.05Pd/AC-300335.4337.30.317.078.05Pd/AC-400335.5337.10.22 5.937.19Pd/OMC-200335.6336.90.48 4.52 4.71Pd/OMC-300335.6336.80.46 4.40 4.66Pd/OMC-400335.7337.10.364.104.341407Y.Shao et al./Catalysis Communications 12(2011)1405–1409normalization,indicative of the absence of mass transfer limitation under the tested reaction conditions [8,17].The liquid phase catalytic HDC of 2,4-DCP over the catalysts is compiled in Fig.3.Only 2-chlorophenol was identi fied as the partially dechlorinated product,and cyclohexanone from further hydrogena-tion of phenol was not detected.At given reduction temperature Pd/OMC exhibited higher catalytic activity than Pd/AC.For example,after reaction for 30min 2,4-DCP was removed by 75%for Pd/AC-200and by 92%for Pd/OMC-200.Accordingly,2-CP was completely reduced in 90min for Pd/OMC-200,but in 150min for Pd/AC-200.The higher catalytic activity of Pd/OMC is tentatively attributed to its higher Pd dispersion as compared to Pd/AC.Furthermore,increasing reduction temperature resulted in declined catalytic activities of Pd/OMC and Pd/AC;while more marked decrease was observed on Pd/AC.For example,increasing reduction temperature from 200to 400°C led to decreased 2,4-DCP removal from 99to 73%for Pd/OMC,but from 95to 16%for Pd/AC within 60min.Such marked difference cannot be explained in terms of Pd particle size because Pd/OMC displays similar Pd particle aggregation to Pd/AC upon H 2reduction treatment.Gomez-Sainero et al.[14]studied the catalytic HDC of CCl 4over Pd/346344342340338336334332I n t e n s i t yBinding Energy (eV)Pd/OMC-200Pd/OMC-300Pd/OMC-400346344342340338336334332I n t e n s i t yBinding Energy (eV)Pd/AC-200Pd/AC-300Pd/AC-400Fig.2.XPS spectra of the catalysts.C o n c e n t r a t i o n (m m o l L -1)Time (min)C o n c e n t r a t i o n (m m o l L -1)Time (min)Fig.3.Liquid phase catalytic HDC of 2,4-DCP over the catalysts.(♦)2,4-DCP,(■)2-CP and (▲)phenol.Lines represent the fitting curves using Eqs.(2)–(4).1408Y.Shao et al./Catalysis Communications 12(2011)1405–1409AC and concluded that the coexistence of Pd 2+and Pd 0was essential for the effective HDC of CCl 4,wherein Pd 0acted as the active site for H 2activation and Pd 2+served as the active site for the activation of C \Cl bond via abstraction of nucleophilic chloride anion and formation of highly reactive +CCl 3ion.Hence,at similar reduction temperature the higher catalytic activity of Pd/OMC can be attributed to its higher Pd 2+content.Accordingly,more markedly decreased Pd 2+content and catalytic activity of Pd/AC was observed with catalyst reduction temperature.The results imply that the catalytic activity of the catalysts can be further optimized by adjusting Pd 2+/Pd 0ratio.Notably,the catalytic activities of the catalysts correlate to both Pd particle size and Pd 2+/Pd 0ratio.Undoubtedly,more research should be warranted in the future to determine their relative importance on catalyst activity.The variation of catalyst structural properties may in fluence the reaction mechanism.The liquid phase catalytic HDC of 2,4-DCP could be implemented via stepwise and concerted pathways (Scheme 1)[8].The rate constants could be further quanti fied by fitting the kinetic data using following equations [18]:C 2;4−DCP =C 02;4−DCP exp −k 1+k 3ðÞt ðÞð2ÞC 2−CP=k 1C 02;4−DCP k 2−k 1−k 3exp −k 1+k 3ðÞt ðÞ−exp −k 2t ðÞðÞð3ÞC Phenol=k 1k 2C o2;4−DCP 2−k 1−k 3−113exp −k 1+k 3ðÞt ðÞ−1ðÞ+12exp −k 2t ðÞ−1ðÞ−k 3C 02;4−DCP13exp −k 1+k 3ðÞt ðÞ−1ðÞð4ÞWhereC 02,4-DCPis the initial 2,4-DCP concentration,C 2,4-DCP ,C 2-CP and C Phenol are the concentrations of 2,4-DCP,2-CP and phenol at reaction time t ,k 1,k 2and k 3are the rate constants of 2,4-DCP to 2-CP,2-CP to phenol,and 2,4-DCP to phenol,respectively.The resultant fitting parameters are listed in Table 3.At similar reduction temperature Pd/OMC had higher rate constants (k 1,k 2and k 3)than Pd/AC,and the rate constants of Pd/OMC and Pd/AC decreased with catalyst reduction temperature,again con firming the positive relation-ship of catalytic activity with Pd 2+content.It is interesting to note that for Pd/OMC and Pd/AC lower catalyst reduction temperature results in higher k 3/k 1ratio.Such trend could be explained in terms of Pd 2+content,wherein high Pd 2+content favors simultaneous activation of ortho-and para-substituted Cl,giving rise to preferential HDC of 2,4-DCP via a concerted pathway.Additionally,at given reduction temperature higher k 3/k 1ratio was observed for Pd/AC than for Pd/OMC.This is probably because when compared with Pd/OMC Pd/AC has larger Pd particle,which provides more planar Pd surface and thus facilitates simultaneous access of Pd 2+sites by ortho-and para-substituted Cl of 2,4-DCP.To verify possible catalyst deactivation,catalyst reuse was conducted and the results are compared in Fig.5S,supporting information.For Pd/OMC-300and Pd/AC-300,gradual deactivation was observed with catalyst recycle.XPS analysis indicated that Pd 2+/Pd 0ratios of the used catalysts remained identical to those of the fresh catalysts.Additionally,Pd content analysis indicated that after one cycle Pd content decreased by 13.0%for Pd/AC-300and 7.7%for Pd/OMC-300.In comparison withPd/AC-300,substantially slower deactivation was shown for Pd/OMC,likely due to the suppressed Pd leaching by the con finement effect.4.ConclusionsIn the present study,the catalytic behaviors of Pd/OMC and Pd/AC for 2,4-DCP HDC were compared.For the supported catalysts,Pd particles are effectively con fined in the mesopores of OMC,but dominantly located on the external surface of AC,resulting in higher Pd dispersion and Pd 2+content of Pd/OMC than Pd/AC.Accordingly,at given catalyst reduction temperature Pd/OMC displays higher catalytic activity than Pd/AC.For both catalysts,increasing catalyst reduction temperature lowers the catalytic activity and favors the stepwise HDC of 2,4-DCP.However,in comparison with Pd/AC the con finement effect of OMC stabilizes Pd 2+,leading to less marked decrease in the catalytic activity of Pd/OMC at elevated reduction temperature.AcknowledgementsThe financial support from the Natural Science Foundation of China (no.20877039)and Program of New Century Excellent Talents in University (NECT-08-0277)are gratefully acknowledged.We are indebted to the Modern Analytical Center,Nanjing University for the material characterization.Appendix A.Supplementary dataSupplementary data to this article can be found online at 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This article appeared in a journal published by Elsevier.The attached copy is furnished to the author for internal non-commercial research and education use,including for instruction at the authors institutionand sharing with colleagues.Other uses,including reproduction and distribution,or selling or licensing copies,or posting to personal,institutional or third partywebsites are prohibited.In most cases authors are permitted to post their version of thearticle(e.g.in Word or Tex form)to their personal website orinstitutional repository.Authors requiring further informationregarding Elsevier’s archiving and manuscript policies areencouraged to visit:/copyrightJ O U R N A L O F T H E M E C H A N I C A L B E H A V I O R O F B I O M E D I C A L M A T E R I A L S3(2010)454–463available atjournal homepage:/locate/jmbbmResearch paperStructure and mechanical properties of pincers of lobster (Procambarus clarkii )and crab (Eriocheir Sinensis )Fei Zhou a ,∗,Zhiwei Wu a ,Meiling Wang a ,Kangmin Chen ba Nanjing University of Aeronautics and Astronautics,Nanjing,210016,Chinab Center of Analysis,Jiangsu University,Zhenjiang,212013,ChinaA R T I C L E I N F O Article history:Received 25August 2009Received in revised form 19April 2010Accepted 3May 2010Published online 11May 2010Keywords:PincerMechanical properties Structure BiomimeticA B S T R A C TThe structure and mechanical properties of the pincer exoskeletons (cuticles)of lobster (Procambarus clarkii )and crab (Eriocheir Sinensis )were investigated,respectively.The microstructures and inorganic materials of the pincer exoskeletons were observed and determined using a scanning electron microscope and X-ray diffraction.The mechanical properties of the pincer exoskeletons were evaluated by nano-indentation tests and tensile tests under different conditions.The results showed that the inorganic materials in the lobster claw exoskeleton exhibited an amorphous structure,while those in the crab claw exoskeleton were crystallized as calcium carbonate with a calcite crystal structure and were stable at the temperature below 250◦C.The surfaces of the pincers were biologically unsmooth.Many concave valleys with setae near the tip and many convex domes far off the tip were observed on the surface of the lobster pincer,while many micro-spines were seen on the surface of crab pincer.The microstructures of the pincer exoskeleton exhibited highly mineralized chitin–protein fibers arranged in a twisted plywood structure.The surface hardness and elastic modulus of the crab claws were 0.33GPa and 8.18GPa,respectively,higher than those of the lobster claw (0.27and 5.44GPa).The transition in the mechanical properties and structure between the exocuticle and the endocuticle was discontinuity.The tensile strength of the crab pincer was two times higher than that of the lobster pincer,and the dry specimen was fractured more easily than the fresh specimen.c2010Elsevier Ltd.All rights reserved.1.IntroductionLobster and crab all belong to arthropods,which are coveredwith an exoskeleton.The exoskeleton of arthropods consists mainly of fibrous chitin-based tissue with a high degree of mineralization such as calcium carbonate to give mechanical rigidity.It is obvious that the exoskeleton materials possess ∗Corresponding author.Tel.:+862584893083;fax:+862584893083.E-mail addresses:fzhou@ ,zhoufei88cn@ (F .Zhou).many excellent functions to support the body,resist mechan-ical loads and provide environmental protection and resis-tance to desiccation.For lobster and crab,there are two large claws in front of their heads,(called pincers),which are used to hold and crack prey or to burrow a hole.Thus,the pin-cers could have better mechanical properties in comparison to other parts.Previously,some scientists have paid more at-tention to the mechanical properties of pincers from lobster1751-6161/$-see front matter c2010Elsevier Ltd.All rights reserved.doi:10.1016/j.jmbbm.2010.05.001J O U R N A L O F T H E M E C H A N I C A L B E H A V I O R O F B I O M E D I C A L M A T E R I A L S3(2010)454–463455Fig.1–P.clarkii(a)and E.Sinensis(b)in freshwater lake.and crab.For example,Raabe and co-workers(Raabe et al., 2005a,b;Sachs et al.,2006a,b;Romano et al.,2007;Sachs et al., 2008)have studied the structure and mechanical properties of the cuticle of American lobster(Homarus americanus),and indicated that the exocuticle was characterized by a veryfine woven structure of thefibrous chitin–protein matrix(called a‘twisted plywood or Bouligand pattern structure,which is formed by the helicoidal stacking sequence of thefibrous chitin–protein layer.).The hardness and stiffness increased from the endocuticle(inner layer)to the exocuticle(outer layer)(Sachs et al.,2006a).Furthermore,the tensile property of the lobster pincers was influenced by moisture.The dry samples failed catastrophically in a brittle manner in the lin-ear elastic region,whereas the natural wet samples showed a pronounced plastic region and stepwise crack propagation (Sachs et al.,2006b).Recently,Chen et al.have studied the structure and mechanical properties of the exoskeleton(cu-ticle)of the sheep crab(Loxorhynchus grandis),and reported that the crab exoskeleton was a three-dimensional composite comprising brittle chitin–protein bundles arranged in a Bouli-gand pattern(the X–Y plane)and ductile pore canal tubules in the direction normal to the surface(the Z direction).The ten-sile strength of wet samples was higher than that of dry sam-ples(Chen et al.,2008).It is evident that the above-mentioned lobsters and crabs were from the ocean and were the largest and heaviest.But the shape and weight of lobsters and crabs from the freshwater lake were smaller and lighter.Further-more,the lobster and crab in the freshwater lake all cling to the muddy ground and burrow living holes,and then have lighter,more elastic and harder pincers.Their pincers could give us some interesting ideas towards the design or produc-tion of novel structural materials.However,the structure and mechanical properties of the pincer exoskeleton of lobster and crab from freshwater lake have not yet been investigated. Thus,the aim of this work is to study the structure and me-chanical properties of the pincer exoskeletons of lobster(Pro-cambarus clarkii)and crab(Eriocheir Sinensis),and then discuss their differences.2.Experimental proceduresThe living lobsters(P.clarkii)and crabs(E.Sinensis)(Fig.1) from a freshwater lake were bought in a local food market in Nanjing,China.They were in the intermolt stage,in which the exoskeletons were completely developed and mineralized. The specimens used in here were taken from the claws of the freshwater lobster and crab.The differential scanning calorimetric(DSC)measurements of pincers(13–17mg)were carried out by a CDR-4P Thermal Analyzer(SBIF,Shanghai, PR China)with an accuracy of±0.2K.The non-isothermal experiments were performed at the heating rates of 20K min−1usingα-Al2O3powder as a reference material.The inorganic materials of dry samples and the heat-treatment samples at250◦C for30min were determined using D8-Advance X-ray diffraction(XRD)(Bruker,Germany)with Cu Kαradiation(λ=0.15404nm).A continuous scan mode was used to collect2θdata from10to100◦at the sampling pitch of 0.02and the scan rate of2◦/min.The X-ray tube voltage and current were set at40kV and40mA,respectively.The microstructures were characterized using afield emis-sion scanning electron microscope(SEM)(JEOL-JSM-7001F) equipped with EDS(Inca Energy350,Oxford,UK).The spec-imens for SEM observation were taken from the fresh claws of lobster and crab,and dehydrated in ethanol–water solution with different ethanol concentrations(75%–100%).In order to observe the surface microstructure of pincers,the pincer sur-face was cleaned and prepared via removing micro-hair from pincer surface.For the cross-sectional sample,a rectangular piece was obtained and then fractured along the longitudinal direction.The fractured sample was mounted into a sample holder made of aluminum alloy,and dried in air for5min, then sputter-coated with10nm gold.Samples were observed with the secondary electron(SE)mode at a15kV accelerating voltage.The variations of hardness and elastic modulus for the pincers with distance along the transverse(Fig.2)and cross-sectional direction were investigated,respectively.The specimens used for surface hardness measurement along the transverse direction were taken from the fresh pincers of the adult lobster and crab.The transverse distance between each indent near tip was6±1mm(Fig.2).T wo kinds of cross-sectional specimens for nanoindentation were taken from the large part of air-dried pincers.The specimen size was about5mm×5mm×1mm(length,width and thickness respectively).The specimens were mounted in epoxy resin with their cross-sectional area revealed,then ground and finely polished(Fig.3).To obtain the influence of temperature on the mechanical properties of pincers,the pincers were heated at different temperatures for30min.The hardness456J O U R N A L O F T H E M E C H A N I C A L B E H A V I O R O F B I O M E D I C A L M A T E R I A L S3(2010)454–463Fig.2–Specimens for nanoindentation from the lobster’s and crab’s pincers at different positions.and elastic modulus of the pincers of lobster(P.clarkii)and crab(E.Sinensis)were measured using a nano-indenter(SA2, MTS,USA)with a Berkovich diamond tip.Load–unload curves were performed at a maximum load of10mN,and the maximum depth of indentation was800nm.The tests were repeated10times at different places to get the accurate value of hardness and elastic modulus.The projected contact area A c was determined from the tip-shaped polynomial function of the contact depth at the maximum indentation load.Thereduced elastic modulus is given by1/E r=(1−ν2i )/E i+(1−ν2s)/E s where E i,E s andνi,νs are the elastic modulus andPoisson’s ratio for the indenter and the substrate materials,respectively.For a diamond indenter material,E i is1050GPaandνi is0.1.The Oliver–Pharr method(Oliver and Pharr,1992)is used to calculate the hardness and reduced modulus valuesfrom loading–unloading curves.Because the limitation of sample size and curvature,the tensile specimens were taken from the different segments ofthe chelipeds(the lobster chela and the crab merus)(Fig.4(a)).After muscle was removed,the specimens were dissectedinto rectangles approximately20mm in length,5mm inwidth and1.5mm in thickness.The rectangle sample wasfixed in the standard tensile specimen mould(Fig.4(b)),and then the remnants around the mould was removed viagrounding,so the standard tensile specimen with a dog-bone shape was acquired(Fig.4(c)).As far as the lobster andcrab were concerned,forty pieces were cut from four kindsof pincers to prepare two equal sets of samples,one in adry condition and one wet.T wenty samples were air-dried,while others were kept in fresh water to prevent desiccationduring preparation.Because the exoskeleton would becomecompletely demineralized after several days,the tensile testswere carried out within24h after the specimens wereobtained.The tensile tester had two symmetric grips;thedown grip wasfixed while the upper grip was movable andattached to a micro-tensile tester,which was equipped with a100N load cell.The upper grip speed was0.05mm/s,equal toa strain rate of1.25×10−2s−1.As is known,the tensilestressFig.3–Cross-sectional specimen for nanoindentation:(a)model of epoxy resin;(b)pincer in epoxy resin block.a b cFig.4–Preparation of tensile specimens for lobster and crab’s pincers:(a)Schematic representation of sampling position for tensile test;(b)the mould for samples preparation:the rectangle samples for pincers werefixed in the mould and the remnants around the mould were removed via grounding,so the standard tensile specimen with a dog-bone shape was acquired;(c)Schematic representation of tensile test:The standard tensile specimens were mounted in two symmetric grips and then were hauled.J O U R N A L O F T H E M E C H A N I C A L B E H A V I O R O F B I O M E D I C A L M A T E R I A L S3(2010)454–463457a bc dFig.5–X-ray diffraction patterns for lobster(a,b)and crab(c,d)pincers.σ=F/A0and strainε=∆L/L,where F is tensile force,A0is theoriginal cross-section area of sample,∆L is the prolongationamount,L is the standard length of sample.Thus,the tensilestress–strain curves were obtained according to the tensileforce–prolongation curves.3.Results and discussion3.1.X-ray diffraction and DSC analysis of pincersFig.5shows the X-ray diffraction patterns of pincers atdifferent temperatures.It is clear that the mineral phasesconsisted of calcium carbonate and amorphous minerals.Incomparison to the X-ray diffraction patterns of the specimensat room temperature,those at250◦C changed a little.Thisindicated that the mineral phase structure was stable whenthe temperature was below250◦C.Raabe et al.also reportedthat the heat-treatment had a little influence on the overallstructure(Raabe et al.,2005a).As seen in Fig.5(a)and(b),thediffraction intensity of calcites was weak.This indicated thatthe mineral phases in lobster pincers could be mainly calcitewith a nanocrystalline structure dispersed in the amorphousmatrix.But in Fig.5(c)and(d),the diffraction intensityof calcites was strong.This showed that there were manycalcites in the crab pincers.In fact,the exoskeleton of thepincer was mineralized with calcium carbonate in the form ofsmall crystallites,calcite and amorphous calcium carbonate,deposited within the chitin–protein matrix(Tong and Yao,1997).This pointed out that the mineral content in the crab(E.Sinensis)pincer was higher than that in the lobster(P.clarkii)pincer.Thus,the calcium carbonate separated crystals out inthe form of calcite.When the temperature was beyond250◦C,as seen in Fig.6,there were three exothermic peaks at313,412and524◦C for the lobster pincer,while two exothermicpeaks located at336and548◦C for the crab pincer wereobserved.Romano et al.indicated that water and proteinswere removed from the cuticle at a temperature between50and270◦C,the decomposition of the biopolymer occurredat370◦C,and the chitin degraded between280and about600◦C(Romano et al.,2007).This indicated that thefirstexothermic peaks at313and336◦C were attributed to thecombustion of the organic matrix of pincer.With an increasein heat temperature,the biopolymer in the lobster pincer wasdecomposited and burned to form the second exothermicpeak at412◦C.If the temperature was beyond500◦C,thechitin degraded completely,and then combusted at524and548◦C to generate the exothermic peak.As seen in Fig.6,it is obvious that the exothermic peak temperature of thecrab pincer was higher than that of the lobster pincer.Thisindicated that the structural stability of the crab pincer wasbetter than that of the lobster pincer.458J O U R N A L O F T H E M E C H A N I C A L B E H A V I O R O F B I O M E D I C A L M A T E R I A L S3(2010)454–463Fig.6–DSC curves for pinchers.3.2.Microstructure of pincers3.2.1.Surface microstructure of pincersFig.7displays the surface microstructure of the pincers. There were many concave valleys with setae(rigid hairs)close to the tip(Fig.7(a)),while the convex domes with setae were observed far off the tip(Fig.7(b)).As seen in Fig.7(c),there were many spines on the seta surface.The setae at different regions displayed various size and growth directions.It is evident from Fig.7(d)that the microstructure on the crab pincer’s surface was different from the lobster surface.Many micro-stings in a cluster were distributed on the crab pincer surface with a certain sequence(Fig.7(d)).The biologically non-smooth surface is always observed in nature.According to the classification of the biologically non-smooth surfaces (Cong et al.,1992),the pincer surface of lobster(P.clarkii)and crab(E.Sinensis)is rough.It is known that the non-smooth surface for soil animals has the anti-adhesive ability to soil. Thus,the lobster(P.clarkii)and the crab(E.Sinensis)are fond of digging holes as their means of living in a lake.3.2.2.Cross-section microstructure of pincersThe cross-section microstructure of the pincers of lobster (P.clarkii)and crab(E.Sinensis)is shown in Fig.8.It is clear that the lamellar structure of the pincer is observed,and the dense layer and the loose layer were distributed alternatively (Fig.8(a)and(c)).This indicated that the difference between exocuticle and endocuticle was easily distinguishable as a change in the stacking space of the twisted plywood layer.As seen in Fig.8(b)and(d),the crab’s and lobster’s pincers show the similar structures.The fracture surface of the twisted plywood layer was covered with many pore canals(pc),which were parallel to the cuticle,and there were many pore canal tubes(pct)on the surface of pore canals.However,thefiber layer thickness of the lobster pincer was thinner than that of the crab pincer.Furthermore,the clusterfibers formed the dense stack layer along the horizontal direction,anda bc dFig.7–Surface microstructure of pincers:There were many concave hollows and setae near the tip of the lobster’s pincers (a),whereas the convex dome with setae was observed away from the tip of the lobster’s pincers(b);there were many spines on the seta surface according to enlarged image(c);Many microspines in a cluster on the crab’s pincer surface with a certain sequence were showed by SEM,and the number of each sting is different(d).J O U R N A L O F T H E M E C H A N I C A L B E H A V I O R O F B I O M E D I C A L M A T E R I A L S3(2010)454–463459 a bc deFig.8–Cross-section microstructure of pincers for lobster(a,b)and crab(c,d,e).thesefibers rotated around the normal axis of the cuticle to form the twisted plywood structure via stacking helicoidally from one plane to another by180◦C about its normal direction,which was frequently observed in natural skeletal or protective extracellular tissues(Fig.8(e)).According to the EDS analysis of the pincers(Fig.9),calcium,oxygen,carbon, and phosphorus were main components of the cuticles,while magnesium and sodium were present in a minor amount, in good accord with Boßelmann et al.(2007).This indicated that the main mineral materials were calcite and amorphous calcium phosphate(ACP).The concentration of calcium in the crab pincer was higher than that of the lobster pincer. In fact,the mineral elements content in the pincer varied with the species of arthropods and with the location in the exoskeleton.But in here,the lobsters and crabs were from the same freshwater lake.This showed that the mineral content of crab pincer was higher than that of lobster.3.3.Mechanical properties of pincers3.3.1.Hardness and elastic modulus of the pincersFig.10shows the variation of hardness(Fig.10(a))and elas-tic modulus(Fig.10(b))with transverse distance from tip to the fourth joint(corresponding to Fig.2).The highest hard-ness was obtained at the situation near the tip.The mean val-ues of hardness and elastic modulus for the crab pincer were 0.33GPa and8.18GPa,respectively,all higher than those of the lobster pincer(0.27GPa,5.44GPa).As seen in Fig.11,the cross-sectional transition distance for the mechanical prop-erties was200µm.When the cross-sectional distance from the surface was lower than200µm,the hardness and elas-tic modulus for the lobster’s and the crab’s pincers varied in the range of0.14–0.30GPa and6–17GPa,0.8–1.1GPa and 22–30GPa,respectively.If the cross-sectional distance from the surface was beyond200µm,the hardness and the elastic460J O U R N A L O F T H E M E C H A N I C A L B E H A V I O R O F B I O M E D I C A L M A T E R I A L S3(2010)454–463Fig.9–EDS analyses of the lobster’s (a)and crab’s (b)cuticles.a bFig.10–Hardness and elastic modulus of pincers for lobster and crab as shown in Fig.2.modulus of the lobster pincer decreased rapidly and varied in the range of 0.004–0.05GPa and 0.2–1.3GPa,while those for the crab pincer fluctuated in the range of 0.14–0.7GPa and 8–22GPa.In comparison with the lobster pincer,the transi-tion in mechanical properties of the crab pincer was gradual.Furthermore,the results from Fig.11showed that the hard-ness value in the exocuticle was higher than that in the en-docuticle,and the change in mechanical properties between two layers was large.This means that lots of the pore canals in the lobster endocuticle result in an arti ficially lower hard-ness and reduced elastic modulus,and then the mechanical properties show a strong discontinuity across the interface between the exocuticle and the endocuticle.The discontinu-ity in mechanical properties along the pincer’s cross-sectional direction was analogous to the results from American lob-ster (H.americanus)(Raabe et al.,2005a ,b )and sheep crab (L.grandis)(Chen et al.,2008).It is clear that the hardness and elastic modulus in the normal and in-plane direction were different.The results in Fig.8(a)and (c)showed that the exo-cuticle (outer layer)was dense,while the endocuticle (inner layer)was loose.When the hardness and elastic modulus were measured in the in-plane direction,the mechanical properties of the epicuticle and exocuticle would be dis-played.When the hardness and elastic modulus were mea-sured in the normal direction,the variation of mechanical properties from the exocuticle to the endocuticle would be displayed.The transition in structure between the exocuticle and endocuticle was abrupt,which induced the transition in mechanical properties along the cross-sectional direction.The in fluence of heat temperature on the hardness of the pincers is shown in Fig.12.With an increase in temperature,the hardness of the lobster pincer first decreased from 0.33to 0.28GPa at 150◦C,and then increased to 0.51GPa obviously,while that of the crab pincer fluctuated slightly in the range of 0.41–0.42MPa at a temperature less than 150◦C,and then increased to 0.69GPa rapidly.Actually,water and proteins were removed from the cuticle at a temperature between 50and 270◦C (Romano et al.,2007).This indicated that the increase of hardness for the pincers was due to the removal of water and proteins from the pincer.J O U R N A L O F T H E M E C H A N I C A L B E H A V I O R O F B I O M E D I C A L M A T E R I A L S3(2010)454–463461(a) Lobster pincers(b) Lobster pincers (c) Crab pincers(d) Crab pincersFig.11–Hardness and elastic modulus of lobster and crab pincers along the cross-sectional direction of thepincer.Fig.12–Influence of temperature on the hardness of lobster and crab pincers.3.3.2.Tensile strengthT ypical tensile stress–strain curves under dry and fresh con-ditions are shown in Fig.13.It is clear that the tensile stress vs.strain curves show non-linear relationships before frac-ture.The fresh specimen of the crab pincer had an ultimate tensile strength of89.42MPa and a fracture strain of10%,Fig.13–Stress–strain curves of lobster’s and crab’s pinchers in both dry and fresh states.while the dry samples fractured at64.67MPa and6.5%.For the lobster pincer,the tensile strength and fracture strain for the fresh samples were41.07MPa and3.5%,while those for the dry samples were38.29MPa and2.83%,respectively. This indicated that the crab’s pincer possessed excellent462J O U R N A L O F T H E M E C H A N I C A L B E H A V I O R O F B I O M E D I C A L M A T E R I A L S3(2010)454–463tensile properties.As seen in Sachs et al.(2006b)and Chen et al.(2008),the dry samples for American lobster Sachs et al. (2006b)and sheep crab(Chen et al.,2008)displayed a linear stress–strain response.But for wet samples before fracture, the American lobster showed an extended non-linear range of elastic–plastic deformation(Sachs et al.,2006b)while the sheep crab still displayed the linear elastic deformation(Chen et al.,2008).The fracture strain of sheep crab was higher than that of American lobster.This indicated that the mechanical properties of pincers were influenced by the microstructure of pincers exoskeletons.In comparison with the results of Sachs et al.(2006b)and Chen et al.(2008),the present results were different from them.In Sachs et al.(2006b)and Chen et al.(2008),the stress–strain behaviors reflected only the me-chanical properties of the endocuticle because the epicuticle and exocuticle were removed during the sample preparation. But here,the stress–strain behaviors reflected the mechani-cal properties of whole exoskeletons.Although the exocuti-cle and the endocuticle fractured at low strain,the epicuticle was still stretching and deforming.Thus,the fracture strains in here all were higher than those in Sachs et al.(2006b) and Chen et al.(2008).This means that the strain values on the stress–strain graphs might not reflect the real stretching strain of the exocuticle and the endocuticle,but the stress val-ues were still credible.3.4.DiscussionAs seen in Figs.5and11–13,it is evident that the mineral phases in lobster pincers could be mainly calcite with a nanocrystalline structure dispersed in the amorphous matrix, while those in the crab pincers were crystallized as calcium carbonate with a calcite crystal structure,and the mechanical properties of the crab pincer were better than those of the lobster pincer.Raabe et al.(2005a),Sachs et al.(2006a) and Chen et al.(2008)reported that the hardness of sheep crab pincer was947±74MPa at100µm depths from the surface,higher than those of the American lobster claw (130–270MPa in exocuticle and30–55MPa in endocuticle) (Raabe et al.,2005a;Sachs et al.,2006a),which was related to the higher mineral content in sheep crab with an ash content of63.6%±4.3%of dry weight(Hayes and Armstrong, 1961).This indicated that the mineral content in the crab pincer was higher than that of the lobster pincer.As seen in Table1,the thickness values of twisted plywood layers in the exocuticle and the endocuticle for crab merus were all lower than those of lobster chela.This indicated that the exoskeleton of crab was denser and thinner than that of lobster.So the mechanical properties of the crab pincer were better than those of the lobster pincer.This indicates that the difference in mechanical properties and structure may be partly due to the different anatomical locations rather than the different species.As seen in Fig.13,it is clear that the fracture strength of the fresh samples was higher than that of dry samples.This indicated that the removal of water from biological materials would play an important role in their mechanical property.Since water is known to act as a plasticizer in biological materials(Vincent,2002),the organic materials would lose their elastic characteristics as water was removed from the pincers.Furthermore,the failure of inorganic minerals occurred at low strains,followed by the fracture of the chitin–protein matrix(Hepburn et al.,1975). Thus,the tensile strength of dry specimens was less than that of fresh specimens.4.ConclusionsThe structure and mechanical properties of the pincers of lobster(P.clarkii)and crab(E.Sinensis)were investigated in detail.The main results could be summarized as:(1)The mineral phases in lobster pincers could be mainlycalcite with a nanocrystalline structure dispersed in the amorphous matrix,while those of the crab pincers were crystallized as calcium carbonate with a calcite crystal structure,which induced the structural stability of crab pincer better than that of lobster pincer.(2)The mechanical properties of the crab(E.Sinensis)pincerswere better than those of the lobster(P.clarkii)pincers, which was due to the twisted plywood layers in the exocuticle,and the endocuticles for crab pincer were all denser than those of lobster pincer.AcknowledgementsThis work was supported by the National Natural Science Foundation of China(NNSFC)(No.50635030).We would like to acknowledge NNSFC forfinancial support.R E F E R E N C E SBoßelmann,F.,Romano,P.,Fabritius,H.,Raabe,D.,Epple,M., 2007.The composition of the exoskeleton of two crustacea: the American lobster Homarus americanus and the edible crab Cancer pagurus.Thermochim.Acta463,65–68.Chen,P.Y.,Lin, A.Y.M.,McKittrick,J.,Meyers,M.A.,2008.Structure and mechanical properties of crab exoskeletons.Acta Biomater.4,587–596.Cong,Q.,Ren,L.,Wu,L.,1992.Taxonomic research on geometric non-smooth animal surface shapes.Trans.Chin.Soc.Agricul.Engin.2(8),7–13.。

python读取⽂件csv，先按第4列排序，再按第5列的数值排序插⼊代码：from operator import itemgetterinput_file = open("PDBhaemoglobinReport.csv")output_file = open("PDBhaemoglobinSorted.csv","w")table = []header = input_file.readline() #读取并弹出第⼀⾏for line in input_file:col = line.split(',') #每⾏分隔为列表，好处理列格式col[3] = float(col[3][1:-1])col[4] = int(col[4][1:-2]) #各⾏没有先strip 末位是\ntable.append(col) #嵌套列表table[[8,8][*,*],...]table_sorted = sorted(table, key=itemgetter(3, 4))#先后按列索引3,4排序output_file.write(header + '\t')for row in table_sorted: #遍历读取排序后的嵌套列表row = [str(x) for x in row] #转换为字符串格式，好写⼊⽂本output_file.write("\t".join(row) + '\n')input_file.close()output_file.close()附：PDBhaemoglobinReportcsv内容PDB ID,Chain ID,Exp. Method,Resolution,Chain Length"1A4F","A","X-RAY DIFFRACTION","2.00","141""1C7C","A","X-RAY DIFFRACTION","1.80","283""1CG5","A","X-RAY DIFFRACTION","1.60","141""1FAW","A","X-RAY DIFFRACTION","3.09","141""1HDA","A","X-RAY DIFFRACTION","2.20","141""1IRD","A","X-RAY DIFFRACTION","1.25","141""1KFR","A","X-RAY DIFFRACTION","1.85","147""1QPW","A","X-RAY DIFFRACTION","1.80","141""1SPG","A","X-RAY DIFFRACTION","1.95","144""1UX8","A","X-RAY DIFFRACTION","2.15","132"PDBhaemoglobinSorted.csv ：PDB ID,Chain ID,Exp. Method,Resolution,Chain Length"2W72" "A" "X-RAY DIFFRACTION" 1.07 141"1IRD" "A" "X-RAY DIFFRACTION" 1.25 141"2H8F" "A" "X-RAY DIFFRACTION" 1.3 143"2WY4" "A" "X-RAY DIFFRACTION" 1.35 140"2D5X" "A" "X-RAY DIFFRACTION" 1.45 141"4ESA" "A" "X-RAY DIFFRACTION" 1.45 143"3VRG" "A" "X-RAY DIFFRACTION" 1.5 141"1CG5" "A" "X-RAY DIFFRACTION" 1.6 141"4HBI" "A" "X-RAY DIFFRACTION" 1.6 146"1QPW" "A" "X-RAY DIFFRACTION" 1.8 141。