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A one-step analysis method was developed for four types of amino acids using a microfluidic paper-based analytical device fabricated from chromatography filtration paper and laminate films. Aminoacyl-tRNA synthetase was used to detect each amino acid. The obtained laminated paper-based analytical device (LPAD) contained four enzymatic reaction areas. Colorimetric detection was performed based on the molybdenum blue reaction. A model method for the simple, easy, and simultaneous detection of several amino acid concentrations was suggested, in contrast to the conventional methods such as HPLC or LC–MS. The method provided a selective quantification at the ranges of 3.6–100 μM for tryptophan, 10.1–100 μM for glycine, 5.9–100 μM for histidine and 5.6–100 μM for lysine with a detection limit of 1.1 μM, 3.3 μM, 1.9 μM and 1.8 μM, respectively. LPAD fabrication was considerably simple, and the subsequent detection process was easy and required a short period of time (within 15 min).

HFE-7100 is a routine carrier solvent in amino acid-sensitive fingermark detection reagents such as ninhydrin and 1,2-indanedione/zinc chloride (IND/Zn). However, a potential EU ban on hydrofluoroethers may require reformulation of these treatments worldwide. Solstice® PF has shown promise as a replacement for HFE-7100 in the United Kingdom. However, the performance (and hence optimal formulation) of IND/Zn is impacted by differences in climate and substrate composition, necessitating assessments under local conditions for different regions. We present a series of preliminary investigations in an Australian context, using the IND/Zn formulation used by Australian forensic service providers. The general performance of Solstice® PF-based IND/Zn was comparable to that using HFE-7100 on three substrate types, three ageing periods (1, 7 and 30 days) and 5 donors. However, slight differences in colour and luminescence intensity, as well as increased ink diffusion, suggest chemical interactions with other reagent components that may affect stability. Specifically, Solstice® PF-based reagent formed a precipitate within a month of storage, though this did not affect performance over a 4 month period. HFE-7100-based IND/Zn was found to be marginally more effective than Solstice® PF when applied to incidental fingermarks. These results indicate that Solstice® PF is a satisfactory alternative carrier solvent to HFE-7100 in an Australian context, though users should be aware of possible limitations regarding compatibility with other evidence components (particularly inks) and shelf-life.

Ninhydrin was implemented as the primary police method of developing latent fingermarks on paper, cardboard and some other porous surfaces from the late 1960s. Some researchers have used individual amino acids, or mixtures of amino acids, as a method of testing the effectiveness of reagent formulations. It was not however known whether simple mixtures of amino acids could effectively emulate latent fingermarks in reactions with reagents such as ninhydrin. The first part of this study compared the effects of ninhydrin fingermark treatments used internationally in various police laboratories on test targets created by inkjet printing graduated concentrations of a representative mixture of amino acids in a series of blocks on paper. Variations in intensity of development were observed between laboratories which used various formulations and heat and humidity post treatment protocols. In a further trial in 2015 several participants in the International Fingerprint Research Group (IFRG) meeting processed test targets in their own laboratories and submitted them for measurement. Again, variation in developed intensity was observed. The depletion of the activity of ninhydrin solutions during use was also investigated in early evaluations of the test targets. An established fingerprint laboratory then processed a number of samples from a batch of targets to examine batch consistency. This was followed by designing a new test target which enabled comparisons between the developed intensity of printed test target blocks alongside depletion series of split, natural donor fingermarks. A panel of 20 donors provided depletion fingermarks and four ninhydrin formulations and treatment protocols were used. The developed test target blocks were scanned, intensity of development measured, and the results compared with the fingermark development which was evaluated by three assessors using two types of scale. Good correlation between the intensity of the developed test targets and latent fingermark quality and intensity scores was observed with the four ninhydrin treatment protocols, including some which used deliberately downgraded ninhydrin concentrations. This type of evaluation was carried out a second time to investigate modified heat and humidity protocols. The use of such test targets for routine reagent quality control and process verification would appear to be far more accurate and reliable than the use of small numbers of donor fingermarks. It is not clear why the different ninhydrin formulations investigated in the latter part of the work have very different optimum post treatment heating regimes.

Current detection methods for paper-based analytical devices (PADs) rely on spectroscopic and electrochemical properties, which place special requirements on the analyte or need analyte labeling. Here, ion-transmission mass spectrometry (MS) was proposed for coupling with PADs to enable rapid in situ MS analysis of the sample on paper. The sample was analyzed directly on paper via analyte ionization by ions transmitted through the paper, generated by a low-temperature plasma probe. Prior to MS analysis, the sample can be separated by paper electrophoresis or by paper chromatography, among a variety of other features offered by PADs. The versatility of this technique was demonstrated by MS analysis of a paper microarray, a mixture of amino acids, and whole blood doped with drugs on PADs.

The file attached to this record is the author's final peer reviewed version. The Publisher's final version can be found by following the DOI link. Enhancement of latent fingermarks on thermal paper poses a number of problems when using traditional methods used for porous substrates due to blackening of the thermal layer as a result of polar solvents present within the reagents and high temperatures oxidising the acid/dye complex. Thus, methods which prevent such reactions are favoured for the development of latent prints on said substrates. A comparative pseudo-operational trial using UV, Hot Print System (HPS), ninhydrin and ThermaNIN was performed on 1000 thermal paper substrates gathered from various sources. The results indicated that the most effective method was an acetone pre-wash followed by ninhydrin. The sequence of HPS-ninhydrin was similarly effective when compared to ninhydrin as a sole technique. ThermaNIN produced fewer marks than ninhydrin but was superior to HPS. Whilst the HPS developed some fingermarks, there was only a very small number of marks uniquely developed by it.

Tropolone reacts with amino acids to form adducts that generate contrast on highly fluorescent paper surfaces upon UV irradiation. Furthermore, the conjugated seven-membered ring of tropolone enables secondary chemical treatments; demonstrated here using two diazonium salts, 4-methoxybenzenediazonium tetrafluoroborate (4MBD) and 4-nitrobenzenediazonium tetrafluoroborate (4NBD). These produce yellow and red dyed fingermarks, respectively. While tropolone treatment is rapid to effect and compatible with ninhydrin in a detection sequence, the methodology proved less effective than current chemical visualisation treatments. Nevertheless, this work unambiguously demonstrates the reactivity of tropolone towards fingermark residues and may inspire future generations of non-benzenoid chemical visualisation treatments.

The duodenal mucosa is exposed to endogenous and exogenous chemicals, including acid, CO2, bile acids and nutrients. Mucosal chemical sensors are necessary to exert physiological responses such as secretion, digestion, absorption, and motility. We propose a mucosal chemosensing system by which luminal chemicals are sensed via mucosal acid sensors and G-protein-coupled receptors. Luminal acid/CO2 sensing consists of ecto- and cytosolic carbonic anhydrases, epithelial ion transporters, and acid sensors expressed on the afferent nerves in the duodenum. Furthermore, a luminal L-glutamate signal is mediated via mucosal L-glutamate receptors, including metabotropic glutamate receptors and taste receptor 1 family heterodimers, with activation of afferent nerves and cyclooxygenase, whereas luminal Ca2+ is differently sensed via the calcium-sensing receptor in the duodenum. Recent studies also show the involvement of enteroendocrine G-protein-coupled receptors in bile acid and fatty acid sensing in the duodenum. These luminal chemosensors help activate mucosal defense mechanisms in or- der to maintain the mucosal integrity and physiological responses. Stimulation of luminal chemosensing in the duodenal mucosa may prevent mucosal injury, affect nutrient metabolism, and modulate sensory nerve activity.

In 1979, Miller and Patel showed that a solution containing two diacetylene monomers, 2,4-hexadiyne-1,6-bis(phenylurethane) (HDDPU) and 2,4-hexadiyne-1,6-bis(p-chlorophenylurethane) (HDDCPU) could be used to develop latent fingermarks on a non-porous surface. In the current work, the same mixture (HDDPU:HDDCPU = 10:1, in acetone solution) was used to develop fingermarks on a wide variety of surfaces, both non-porous and porous, including paper. An airbrush system was optimized for the application of the reagent solution. Once the solution evaporates on a surface, the monomers co-crystallize in different ways, depending upon a number of factors, including the surface residue. " Active" co-crystallization leads (with heat or radiation) to the formation of purple polymer, while " inactive" crystallization results in a non-polymerizable white deposit. Fingermark contrast was achieved as a result of active co-crystallization (giving purple polymer) in either the ridges or the furrows, depending upon the surface and other factors. A general observation (supported by spot tests with linseed oil, salt and amino acid solutions) was that on paper, oily materials are more likely to lead to the formation of the purple polymer, while the presence of water inhibits polymerization. However, these observations are not consistent across all other substrates. It is hypothesized that water disrupts hydrogen bonding between diacetylene molecules, and thus prevents the topochemical polymerization of the diacetylenes, which occurs in the solid state between favourably aligned monomers. An interesting observation was the development of fingermarks deposited on paper that had already been treated with the diacetylene reagent. © 2011 Elsevier Ireland Ltd.

In this paper, we present our preliminary studies into naphthoquinones as novel reagents for the detection of latent fingermarks on paper. Latent fingermarks deposited on paper substrates were treated with solutions of selected naphthoquinones in ethyl acetate/HFE-7100, with subsequent heating. The selected compounds were 1,4-dihydroxy-2-naphthoic acid, 1,2-naphthoquinone-4-sulfonate, 2-methoxy-1,4-naphthoquinone and 2-methyl-1,4-naphthoquinone. All of the tested compounds yielded purple-brown visible fingermarks, which also exhibited photoluminescence when illuminated with a high intensity filtered light source at 555 nm and viewed through red goggles. Indirect heat using an oven at 150 °C for 1 h was found to be superior to direct heat with an iron, which while providing faster development lead to increased levels of background colouration. Luminescence spectrophotometry revealed differences in photoluminescence characteristics for fingermarks developed with the different naphthoquinones, with excitation over the range 530–590 nm. Luminescence spectrophotometry of developed lysine, glycine and serine spots on paper was used to confirm that the naphthoquinones were reacting with amino acids in the latent fingermark.

BMP2 signaling and RUNX2 regulatory pathways converge for transcriptional control of bone formation in vivo. SMAD proteins are recruited to RUNX2 regulatory complexes via an overlapping nuclear matrix targeting signal/Smad interacting domain sequence (391–432) in Runx2. To establish the contribution of RUNX2-SMAD interaction to osteoblastogenesis, we characterized a number of point mutants. Only a triple mutation of amino acids 426–428 (HTY-AAA) results in loss of RUNX2 interactions with either BMP2- or TGF-β- responsive SMADs and fails to integrate the BMP2/TGF-β signal on target gene promoters. In a Runx2 null cell reconstitution assay, the HTY mutant did not activate the program of osteoblast differentiation (alkaline phosphatase, collagen type 1, osteopontin, bone sialoprotein and osteocalcin) in response to BMP2 signaling. Thus, subnuclear targeting function and formation of a RUNX2-SMAD osteogenic complex are functionally inseparable. Taken together, these studies provide direct evidence that RUNX2 is essential for execution and completion of BMP2 signaling for osteoblast differentiation.

Peritubular dentin (PTD) is a hypermineralized phase within the dentinal tubules in some vertebrate teeth as an interface between the intertubular dentin (ITD) and the cell processes. Our aim has been to understand the composition, structure and role of PTD as a mineralized tissue. We have utilized the technique of time of flight secondary ion mass spectrometry (TOF-SIMS) to map the distribution of positive and negative inorganic ions as well as organic components in the fully mineralized, intact PTD structure in bovine tooth cross-sections, and correlated these with scanning electron microscopy (SEM) in standard and backscatter modes. In recent work, we developed a procedure to freeze fracture the teeth and separate PTD from the less dense ITD by the use of aqueous sodium phosphotungstate step density gradients, after degrading the ITD collagen with NaOCl. Here, PTD-containing fragments were characterized by SEM and TOF-SIMS surface structure analysis. The TOF-SIMS data show that the isolated PTD does not contain collagen, but its surface is rich in glutamic acid-containing protein(s). The TOF-SIMS spectra also indicated that the intact PTD fragments contain phospholipids, and chemical analyses showed phosphatidylserine, phosphatidylinositol and phosphatidylcholine as the principal lipid components. In SEM sections, untreated PTD shows as a smooth collar around the tubule, but after digestion with ethylenediamine to remove all organic components, the porous nature of the mineral phase of small, thin platy apatite crystals becomes evident. Thus, the organic matrix of PTD appears to be a proteolipid-phospholipid complex.

Effect of paper mill effluents on Phaseolus aureus Roxb was studied. The effluent significantly inhibited germination of root and shoot length. The bio-chemical injury does not appear spontaneously but with the increase in effluent treatment there is reduction in observed biochemical parameters (chlorophyll, protein, amino acid, nuclic acids and carbohydrate) which are negatively correlated. The shoots of the seedlings were found to be resistant; whereas roots of the seedlings were susceptible to paper mill effluent treatment.

This study concerned the anaerobic treatment of five different industrial wastewaters with a diverse and complex chemical composition. The kinetics of biotransformation of this wastewater at different chemical oxygen demand (COD) were studied in a batch reactor. Wastewater from an amino acid producing industry (Fermex) and from a tank that received several types of wastewaters (collector) contained 0.83 g l(-1) and 0.085 g l(-1) sulfate, respectively. During the study period of 20 days, methane formation was observed in all types of wastewaters. Studies on COD biodegradation showed the reaction velocity was higher for Fermex wastewater and lower for collector wastewater, with values of 0.0022 h(-1) and 0.0011 h(-1), respectively. A lower methanogenic activity of 0.163 g CH4 day(-1) g(-1) volatile suspended solids (VSS) and 0.20 g CH4 day(-1) g(-1) VSS, respectively, was observed for paper producing and brewery wastewater. Adapted granular sludge showed the best biodegradation of COD during the 20-day period. The sulfate-reducing activity in pharmaceutical and collector wastewater was studied. A positive effect of sulfate-reducing activity on methanogenic activity was noted for both types of wastewaters, both of which contained sulfate ions. All reactions of methane generation for the tested industrial wastewaters were first-order. The results of this study suggest that the tested wastewaters are amenable to anaerobic treatment.

The purpose of this study was to evaluate photodegradabilities of the following new low-nitrogen chelating agents: N-bis[(carboxymethoxy)ethyl]glycine (compound 1), N-bis[(1,2-dicarboxyethoxy)ethyl]glycine (compound 2) and N-bis[(1,2-dicarboxyethoxy)ethyl]aspartic acid (compound 3). At first photodegradation of these chelating agents as uncomplexed Na-compound 1–3 and Cu(II) complexes were tested, both in lake and distilled water, by exposing them to near-UV region radiation at the range of 315–400 nm. Uncomplexed Na-compounds 2 and 3 were selected to sunlight exposure experiments carried out in lake and distilled water. Compound 3 was also tested in sunlight as Cu and Ca complexes in both solutions. Photodegradation of Na6-compound 3 in distilled water was studied by exposing it to radiation at the wavelength of 253.7 nm. Photodegradation products were analysed by means of GC-MS (gas chromatography with mass selective detector). The results demonstrated that compound 1 was quite photostable even as Cu complex while compounds 2 and 3 were found to be photodegradable. Over 90% reduction of compound 3 was achieved during one week and 80% reduction of compound 2 in two weeks' time when they were added as Na salt to lake water and exposed to sunlight. Compound 3 as Cu complex degraded totally in the sunlight in less than one week. In the case of compound 3, the degradation rate decreased depending on the counter cation in the order Cu > Na ⩾ Ca. The study demonstrated that photodegradation of Na6-compound 3 does not result in total mineralization of the compound. A photodegradation pathway for Na6-compound 3 is proposed.

1. We used spinal microdialysis in awake rats to investigate whether the repeated withdrawal with naloxone during continuous spinal infusion of morphine would lead to a progressively greater spinal glutamate release and a more pronounced intrathecal tolerance. 2. Rats received lumbar intrathecal (IT) infusion of morphine (IT-M: 20 nmol microl(-1) h(-1)) or saline (IT-S: 1 microl h(-1)) continuously for 3 days. Both groups were further subdivided to receive intraperitoneal (i.p.) injection of naloxone (IP-N: 0.6 mg kg(-1)) or saline (IP-S: 3 ml kg(-1)) every 24 h after the beginning of IT infusion. Daily thermal escape latencies, withdrawal signs, the resting basal release of spinal amino acids before IP injection and the release immediately after the injection (evoked) were measured. 3. Rats receiving IT morphine showed a maximum increase in thermal escape latency on day 1, after which this value declined, with the fastest decline observed in IT morphine + IP naloxone group. On day 1, no significant difference was observed among groups in the resting basal release of amino acids. Rats in IT morphine + i.p. naloxone group displayed a progressive increase in this value. The release was not significantly altered in other groups. 4. For the IT-M + IP-N group, basal resting dialysate concentrations of Glu, Asp and Tau rose steadily over the 3-day infusion interval. No change in basal resting release was noted for any other treatment. 5. Evoked release (after i.p. naloxone) in IT-M animals displayed a progressive increase over the three repeated exposures. Evoked release did not change significantly in other treatment groups. 6. The degree of precipitated withdrawal significantly correlated with the increase in glutamate acutely evoked by i.p. injection. 7. The present results show that periodic transient withdrawal of spinal opiate agonist activity leads to a progressive increase in glutamate outflow and withdrawal signs, in a manner consistent with an enhanced development of spinal tolerance.

1. delta-(l-alpha-Aminoadipyl)-l-cysteine, the corresponding d- and dl-alpha-aminoadipyl isomers, delta-(dl-alpha-amino[6-(14)C]adipyl)-l-cysteine and gamma- and alpha-l-glutamyl-l-cysteine were synthesized. 2. The behaviour of delta-(l-aminoadipyl)-l-cysteine and the corresponding d- and dl-alpha-aminoadipyl isomers was studied in the presence of suspensions of intact mycelium of a Cephalosporium sp., suspensions treated ultrasonically and extracts obtained by grinding with alumina. 3. With intact mycelium the l-alpha-aminoadipyl isomer was removed more rapidly from the extracellular fluid than the corresponding d-isomer. 4. Addition of delta-(dl-alpha-amino[6-(14)C]adipyl)-l-cysteine to suspensions of intact mycelium led to the labelling of extracellular and intracellular penicillin N and cephalosporin C, but also to extensive hydrolysis of the dipeptide. 5. Broken-cell systems hydrolysed delta-(l-alpha-aminoadipyl)-l-cysteine and the corresponding d-alpha-aminoadipyl isomer, but the former was hydrolysed more readily than the latter. 6. gamma- and alpha-l-Glutamyl-l-cysteine were also hydrolysed but delta-(l-alpha-aminoadipyl)-l-cysteinyl-l-valine was not. 7. Only part of the enzyme activity in broken-cell systems responsible for the hydrolysis of delta-(alpha-aminoadipyl)-l-cysteine was present in the supernatant obtained on centrifugation at 20000g. 8. Possible implications of these findings are discussed.

1 A new diagonal electrophoretic technique for determining the order of the lysine-containing tryptic peptides of a protein is described. The protein is converted into its trifluoracetyl derivative, digested enzymatically (or chemically), and the resulting peptides separated by paper electrophoresis. The paper is then treated with ammonia vapour, which re-exposes the ɛ-amino groups of the lysine residues, and submitted to a second electrophoresis at right angles to the first direction. Peptides containing lysine residues, together with the N-terminal peptide of the protein, are found to lie off a diagonal formed by all other peptides, whence they may be readily purified. A study of these peptides enables the order of the lysine-containing tryptic peptides in the protein to be deduced. 2 The technique has been successfully tested with insulin. 3 When the method was applied to porcine pepsin, the four tryptic peptides isolated were easily ordered and the carboxyl-terminal sequence of the protein shown to be Arg-Gln-Tyr-Tyr-Thr-Val-Phe-Asp-Arg-Ala-Asn-Asn-Lys-Val-Gly-Leu-Ala-Pro-Val-Ala. The three basic amino acid residues in the molecule are thus found clustering towards the C-terminus of the polypeptide chain. 4 A common ancestral gene for porcine pepsin and bovine (calf) rennin is suggested by the close homology between the C-terminal sequences of the two proteins.

The partial amino acid sequence of two large peptides is described. These were prepared from the N-terminal half of the heavy chain of immunoglobulin G from pooled normal rabbit serum by tryptic digestion after the ∈-amino groups of the lysine residues had been blocked with S-ethyl trifluorothioacetate. These peptides are believed to account for about 145 residues of fragment C-1, the N-terminal section of rabbit immunoglobulin G heavy chain prepared by cyanogen bromide cleavage. The evidence from the present paper and the preceding paper (Cebra, Givol & Porter, 1968) suggests that it may be possible to deduce a predominant amino acid sequence for most, if not all, of this section of the molecule.

Four different 1-aminocyclohexanes bearing a tethered thioxanthone group in the 2-position were prepared. The synthesis commenced with the respective N-protected β-amino acids, the carboxyl group of which was employed for the introduction of the thioxanthone moiety. After construction of the thioxanthone and protecting group removal, the conversion of the amino group into the respective thiourea was accomplished by treatment with N-3,5-bis(trifluoromethyl)phenyl isothiocyanate and yielded the title compounds in which the thioxanthone resides in different spatial positions relative to the thiourea motif. Overall yields varied between 20–35%.

Thialysine is oxidatively deaminated by snake venom L-aminoacid oxidase at alkaline pH. The oxygen consumption curves show a characteristic diphasic course: the quick uptake of half a mole of oxygen per mole of substrate, in aggreement with a typical oxidative deamination, is followed by a slow extra oxygen consumption. The first product of the reaction is the corresponding alpha-oxo-epsilon-amino acid, which spontaneously cyclizes to the internal Schiff base 5-6-dihydro-delta 3,1,4-thiazin-3-carboxylic acid (TZCA). This latter has been identified by its UV absorption spectrum, by some chemical reactions, by paper chromatography, and by the production of cystamine and glyoxylic acid after prolonged oxidation of thialysine followed by acid hydrolysis. The possibility of an alpha-beta elimination reaction giving rise to cysteamine from thialysine, coupled to the oxidative deamination, has been excluded.

THE high resolving powers of columns of cross-linked sulphonated polystyrene resins when applied to the separation of mixtures of amino-acids has been demonstrated by the classic work of Moore and Stein1 and afterwards by other workers2,3.