Topic:Spectroscopy
Spectroscopy is an analytical technique used to measure the interaction between matter and electromagnetic radiation. In equine research, spectroscopy is applied to assess various biological and chemical properties of horses. Techniques such as infrared spectroscopy, nuclear magnetic resonance (NMR) spectroscopy, and mass spectrometry are utilized to analyze tissues, fluids, and other biological samples. These methods provide insights into metabolic processes, nutritional status, and disease states in horses. Spectroscopy aids in the identification and quantification of biomolecules, contributing to the understanding of equine physiology and pathology. This page compiles peer-reviewed research studies and scholarly articles that explore the application, methodology, and findings of spectroscopic techniques in equine science.
Stereochemical structures of synthesized and natural plasmalogalactosylceramides from equine brain. Modified galactosylceramide with a long-chain cyclic acetal at the sugar moiety, plasmalogalactosylceramide, was isolated from equine brain. To identify the isomeric stereostructure of the natural product, the plasmalo derivative was chemically synthesized from galactosylceramide through acetalization. The presence of cyclic acetal linkage, the linked position and length of the acetal chain of the synthesized and natural products were determined by proton nuclear magnetic resonance spectroscopy and fast-atom bombardment-mass spectrometry, as well as gas chromatography-mass spectrometry and gas...
Equine infectious anemia virus transactivator is a homeodomain-type protein. Lentiviral transactivator (Tat) proteins are essential for viral replication. Tat proteins of human immunodeficiency virus type 1 and bovine immunodeficiency virus form complexes with their respective RNA targets (Tat responsive element, TAR), and specific binding of the equine anemia virus (EIAV) Tat protein to a target TAR RNA is suggested by mutational analysis of the TAR RNA. Structural data on equine infectious anemia virus Tat protein reveal a helix-loop-helix-turn-helix limit structure very similar to homeobox domains that are known to bind specifically to DNA. Here we report results of...
Further characterization of equine brain gangliosides: the presence of GM3 having N-glycolyl neuraminic acid in the central nervous system. Equine brain gangliosides were isolated and their structures were characterized, to examine whether equine brain has N-glycolyl neuraminic acid in gangliosides, since other mammals predominantly possess N-acetyl neuraminic acid in brain gangliosides, and equine erythrocytes and organs except the brain have gangliosides exclusively containing N-glycolyl neuraminic acid. The gangliosides purified from the brain were identified by proton NMR spectroscopy and mass spectrometry, as well as GLC, resulting in their identification as GM4, GM3, GM2, GM1, GD1a, GD1b, and GT1b. Of these gangliosides, GM3...
Solvent effects on horse apomyoglobin dynamics. The effects of the solvent conditions (buffer pH 9, 8, or 7 or buffer pH 6.5 alone or mixed with 3.2% ethanol or 6.2% formamide) on the protein dynamics of horse apomyoglobin were investigated through tryptophan fluorescence quenching, spectra, and decay properties. Raising the pH (which induces discontinuous protein conformation changes) increases the structural fluctuations inside the hydrophobic A, G, and H helix core. Mixed solutions containing either 3.2% ethanol or 6.2% formamide (which redistribute water molecules on the protein surface) produce protein dynamics changes in the vicinity ...
Biochemical and conformational characterisation of HSP-3, a stallion seminal plasma protein of the cysteine-rich secretory protein (CRISP) family. HSP-3 is a member of the cysteine-rich secretory protein (CRISP) family from stallion seminal plasma. We report a large-scale purification protocol for native HSP-3. This protein is a non-glycosylated polypeptide chain with a pI of 8-9 and an isotope-averaged molecular mass of 24987 +/- 3 Da. The molecular mass of HSP-3, determined by equilibrium sedimentation, is 26 kDa, showing that the protein exists in solution as a monomer. The concentration of HSP-3 in the seminal plasma of different stallions ranged from 0.3 to 1.3 mg/ml. On average, 0.9-9 million HSP-3 molecules/cell coat the postacros...
Fourier-transform infrared spectroscopic studies on the coordination of the side-chain COO- groups to Ca2+ in equine lysozyme. Interactions between Ca2+ and the Asp side chains in the Ca2+-binding site of equine lysozyme were investigated by Fourier-transform infrared (FT-IR) spectroscopy. In the spectrum of equine lysozyme, the intensities of the bands at about 1595 cm-1 and 1578 cm-1 in the region of the COO antisymmetric stretches increased upon Ca2+ binding. In the region of the COO- symmetric stretches, the loss of intensity at about 1388 cm-1 and gains of intensities at about 1423 cm-1 and 1403 cm-1 were observed due to Ca2+ binding to equine lysozyme. The spectral changes for equine lysozyme indicate that the C...
A myoglobin variant with a polar substitution in a conserved hydrophobic cluster in the heme binding pocket. Well-ordered internal amino acids can contribute significantly to the stability of proteins. To investigate the importance of the hydrophobic packing interface between helices G and H in the proximal heme pocket of horse heart myoglobin, the highly conserved amino acid, Leu104, was substituted with asparagine, a polar amino acid of similar size. The Leu104Asn mutant protein and its recombinant wild-type horse heart myoglobin counterpart were expressed from synthetic genes in Escherichia coli. Thermal denaturation of these two recombinant myoglobins, as studied by measurement of circular dichro...
Infrared and atomic spectrometry analysis of the mineral composition of a series of equine sabulous material samples and urinary calculi. Atomic spectrometry has been used in 20 samples of equine urinary sabulous deposits in order to detect minor elements accompanying the predominant element, calcium, which is present in the form of calcium carbonate (calcite and/or vaterite). The elements measured have been (besides calcium) magnesium, sodium, potassium, iron, copper and manganese. Phosphates, sulphates and silica are frequently present as minor constituents of equine urinary sabulous deposits and uroliths, but their detection can be difficult by infrared (IR) spectroscopy in the original samples due to overlapping with the ban...
Structural characterisation and comparison of the native and A-states of equine lysozyme. Native state 1H NMR resonance assignments for 125 of the 129 residues of equine lysozyme have enabled measurement of the hydrogen exchange kinetics for over 60 backbone amide and three tryptophan indole hydrogen atoms in the native state. Native holo equine lysozyme hydrogen exchange protection factors are as large as 10(6), the most protected residues being located in elements of secondary structure. High exchange protection in the domain interface correlates with the binding of Ca2+ in this region. Equine lysozyme differs from most non-Ca2+ binding lysozymes in forming a highly populated par...
Multi-element assay of mammary secretions and sera from periparturient mares by inductively coupled argon plasma emission spectroscopy. To document and determine changes in the mineral profiles of sera and mammary secretions from a population of periparturient mares. Methods: 18 clinically normal periparturient Arabian broodmares. Methods: Inductively coupled argon emission spectroscopy was used to measure Ca, Cu, Fe, K, Mg, Mn, Na, P, and Zn concentrations in sera and mammary secretions of periparturient mares. In addition, S was measured in mammary secretions. Results: Serum concentrations of Ca, Cu, Fe, K, Mg, Na, P, and Zn remained constant throughout late pregnancy and the first 7 days of lactation. Compared with values o...
Molten globule state of equine beta-lactoglobulin. The acid-unfolded state of equine beta-lactoglobulin was characterized by means of circular dichroism, nuclear magnetic resonance, analytical gel-filtration chromatography, and analytical centrifugation. The acid-unfolded state of equine beta-lactoglobulin has a substantial secondary structure as shown by the far-ultraviolet circular dichroism spectrum but lacks persistent tertiary packing of the side chains as indicated by the near-ultraviolet circular dichroism and nuclear magnetic resonance spectra. It is nearly as compact as the native conformation as shown by the gel filtration and sedime...
1H-NMR study of inter-segmental hydrogen bonds in sperm whale and horse apomyoglobins. NMR signals for HisB5 N(delta)H and HisEF5 N(epsilon)H protons of sperm whale and horse apomyoglobins were assigned and compared with the corresponding signals of the holoproteins in terms of pH and temperature dependence behaviors of their shifts and line widths in order to gain insight into structural difference between the apoproteins and the holoproteins. Since these protons are involved in internal hydrogen bonds at the interfaces between the B helix and the GH corner and between the EF corner and the H helix, local structures of the interfaces in these proteins have been inferred from th...
Heart rate variability in the horse by ambulatory monitoring. Using a microprocessor controlled Ambulatory Monitoring System (AMS) developed by one of us (LvD), we have been studying the changes in and control of heart rate in the resting horse. The system provides us with InterBeat Intervals (IBI in milliseconds), motion sensing, and a time domain measure (mean successive differences: MSD) of heart rate variability for periods up to 72 hours. Thoracic impedance is also available but parameters for the equine chest are not currently available. The system is completely noninvasive, small, and carried on a surcingle worn by the subject. The equine subject ...
Myoglobin oxygen dissociation by multiwavelength spectroscopy. Multiwavelength optical spectroscopy was used to determine the oxygen-binding characteristics for equine myoglobin. Oxygen-binding relationships as a function of oxygen tension were determined for temperatures of 10, 25, 35, 37, and 40 degrees C, at pH 7.0. In addition, dissociation curves were determined at 37 degrees C for pH 6.5, 7.0, and 7.5. Equilibration was achieved with a myoglobin solution, at the desired temperature and pH, and 16 oxygen-nitrogen gas mixtures of known oxygen fraction. Correction for the inevitable presence of metmyoglobin was made by using a three-component least squ...
Local electrostatic potentials in pyridoxal phosphate labelled horse heart cytochrome c. The present work shows the application of an optical label pyridoxal phosphate (PLP) for the experimental determination of local electrostatic potentials in singly substituted cytochromes c modified by pyridoxal phosphate at Lys 79 (PLP-Lys-79-cyt.c) or at Lys 86 (PLP-Lys-86-cyt.c). PLP has also been used to calculate the pKa values of all ionizable groups and the electrostatic potentials in the modified proteins and to analyse their properties. The experimental pKa values for the pyridine nitrogen and phenolic hydroxyl of the bound label were obtained from pH-dependent absorbance and fluoresc...
The structure of the keratan sulphate chains attached to fibromodulin isolated from articular cartilage. Fibromodulin has been isolated from bovine and equine articular cartilage and the attached keratan sulphate chains subjected to digestion by keratanase II. The oligosaccharides generated have been reduced and subsequently isolated by strong anion-exchange chromatography. Their structures have been determined by high-field 1H-NMR spectroscopy and high-pH anion-exchange chromatography. Both alpha(2-6)- and alpha(2-3)-linked N-acetylneuraminic acid have been found in the capping oligosaccharides, and, fucose which is alpha(1-3)-linked to N-acetylglucosamine has been found as a branch in both repe...
Glyceryl trinitrate enhances nitric oxide mediated perfusion within the equine hoof. Laminitis, a microvascular disease of the equine hoof leads to severe lameness. Exogenous iv 1-arginine and transdermal nitric oxide donors, such as GTN, applied to the pasterns improve lameness during acute laminitis. Near Infrared spectroscopy in an earlier study showed haemostasis and ischaemia in the hoof during acute laminitis, both were alleviated by 1-arginine. Quantitative NIRS in the present study shows that transdermal GTN increases blood flow in the equine hoof. It is concluded that glyceryl trinitrate enhances nitric oxide mediated perfusion within the equine hoof in normal and chr...
Molecular diffusion into horse spleen ferritin: a nitroxide radical spin probe study. Electron paramagnetic resonance spectroscopy and gel permeation chromatography were employed to study the molecular diffusion of a number of small nitroxide spin probes (approximately 7-9 A diameter) into the central cavity of the iron-storage protein ferritin. Charge and polarity of these radicals play a critical role in the diffusion process. The negatively charged radical 4-carboxy-2,2,6,6-tetramethylpiperidine-N-oxyl (4-carboxy-TEMPO) does not penetrate the cavity whereas the positively charged 4-amino-TEMPO and 3-(aminomethyl)-proxyl radical and polar 4-hydroxy-TEMPO radical do. Unlike th...
Surface plasmon resonance studies of complex formation between cytochrome c and bovine cytochrome c oxidase incorporated into a supported planar lipid bilayer. II. Binding of cytochrome c to oxidase-containing cardiolipin/phosphatidylcholine membranes. Complex formation between horse heart cytochrome c (cyt c) and bovine cytochrome c oxidase (cco) incorporated into a supported planar egg phosphatidylcholine membrane containing varying amounts of cardiolipin (CL) (0-20 mol%) has been studied under low (10 mM) and medium (160 mM) ionic strength conditions by surface plasmon resonance (SPR) spectroscopy. Both specific and nonspecific modes of cyt c binding are observed. The dissociation constant of the specific interaction between cyt c and cco increases from approximately 6.5 microM at low ionic strength to 18 microM at medium ionic strength, ...
Influence of glycerol on the structure and stability of ferric horse heart myoglobin: a SAXS and circular dichroism study. The influence of glycerol on the structural properties of Fe(III)-horse heart myoglobin has been investigated by absorbance, CD and SR-SAXS spectroscopy. The results obtained indicate that both the tertiary and the secondary (alpha-helix) conformations of the protein are influenced by glycerol; in particular, an increase of approx. 8% in helical content was observed. Further, analysis of both the acid- and guanidine-induced denaturation transitions points to a glycerol-induced decreased stability of the tertiary structure; conversely, the alpha-helix conformation is found to be stabilized by t...
Looking for residues involved in the muscle acylphosphatase catalytic mechanism and structural stabilization: role of Asn41, Thr42, and Thr46. Asn41, Thr42, and Thr46 are invariant residues in both muscle and erythrocyte acylphosphatases isolated so far. Horse muscle acylphosphatase solution structure suggests their close spatial relationship to Arg23, the main substrate binding site. The catalytic and structural role of such residues, as well as their influence on muscle acylphosphatase stability, was investigated by preparing several gene mutants (Thr42Ala, Thr46Ala, Asn41Ala, Asn41Ser, and Asn41Gln) by oligonucleotide-directed mutagenesis. The mutated genes were cloned and expressed in Escherichia coli, and the mutant enzymes were...
Minor constituents of sabulous material in equine urine. The secondary constituents accompanying calcite and vaterite (crystalline forms of calcium carbonate) in the sabulous deposits from 140 vesical samples and one renal sample of equine urine were studied by infrared spectroscopy (IR), scanning electron microscopy (SEM) and energy dispersive X-ray analysis (EDX). Apatitic calcium phosphate, present in 12 per cent of the samples, generally appeared in the form of spherulites with smooth and rough surfaces. Calcium sulphate, clearly detected by IR in 12.7 per cent of the samples, did not have a characteristic structure under SEM, although EDX detec...
Phospholipid composition of erythrocyte membranes and plasma of mammalian blood including Australian marsupials; quantitative 31P NMR analysis using detergent. The phospholipid classes of erythrocyte membranes and plasma from several domestic animals and marsupials were quantified by 31P NMR using detergents. Washed erythrocyte samples were thoroughly haemolysed by tip-sonication and dissolved in sodium cholate; plasma samples were dissolved in Triton X-100. The species studied were: common wombat (Vombatus ursinus), black-striped wallaby (Macropus dorsalis), bandicoot (Isoodon macrocarpus), Eastern grey kangaroo (Macropus giganteus), Tammar wallaby (Macropus eugenii), sheep (Ovis aries), goat (Capra hircus), cattle (Bos taurus), horse (Equus caballu...
Nitric oxide donors as treatment for grass induced acute laminitis in ponies. The potential for participation of the arginine-nitric oxide system in the aetiology of acute equine laminitis has been assessed. Nitric oxide (NO), produced by the action of NO synthase (NOS) on its substrate l-arginine, relaxes vascular smooth muscle to cause vasodilation. An attenuated normal vasodilatory tone may characterise the pathogenesis of acute equine laminitis. An intravenous infusion of 10% l-arginine in 0.9% saline caused vasodilatation in the hoof of a normal pony and immediate reperfusion of laminal tissues in an acutely laminitic pony, detected noninvasively by near infrared s...
Factors affecting the ultrasonic properties of equine digital flexor tendons. The velocity, attenuation and apparent backscattering coefficient of 6-11-MHz ultrasound were measured in three orthogonal directions in equine deep digital flexor (DDF) and superficial digital flexor (SDF) tendons at 0 degree C. Ultrasonic measurements were examined for correlation with tendon water, collagen, DNA and glycosaminoglycans contents, determined by chemical analyses and with structure observed by scanning electron microscopy. The SDF tendon contained more water, more DNA (i.e., more cells), less collagen and less glycosaminoglycans and exhibited lower velocities and attenuations t...
Neutron anatomy. The familiar extremes of crystalline material are single-crystals and random powders. In between these two extremes are polycrystalline aggregates, not randomly arranged but possessing some preferred orientation and this is the form taken by constructional materials, be they steel girders or the bones of a human or animal skeleton. The details of the preferred orientation determine the ability of the material to withstand stress in any direction. In the case of bone the crucial factor is the orientation of the c-axes of the mineral content-the crystals of the hexagonal hydroxyapatite- and this...
Near infrared spectroscopy of pedal haemodynamics and oxygenation in normal and laminitic horses. The present study applies near infrared spectroscopy (NIRS) to the haemodynamics of the pedal circulation in normal and laminitic horses. NIRS is a noninvasive technique which uses changes in light absorption at 4 wavelengths to provide information on the changes in cytochrome aa3 (cyt aa3) reduction-oxidation (redox) status, and changes in the tissue concentration of oxyhaemoglobin (O2Hb), deoxyhaemoglobin (HHb) and therefore total haemoglobin (tHb). Other studies have shown NIRS to be sensitive to changes in tissue oxygenation and perfusion in human cerebral and limb circulation. In this stu...
Variable-temperature study of the heme-reorientation process in equine myoglobin. The redistribution of the initially-formed myoglobin heme-insertion isomers from the initially formed 50/50 mixture to the equilibrium ratio of 90/10 has long been assumed to occur by one of two mechanisms, both of which require the rupture of the heme iron-protein bond (La Mar, G.N., Toi, H. and Krishnamoorthi, K. (1984) J. Am. Chem. Soc. 106, 6395-6401). In this study we compared the use of nuclear magnetic resonance and optical spectroscopic techniques as methods for studying the reorientation of heme within myoglobin. We found that kinetics determinations of the heme insertion isomer redis...
Beta-thiopropionyl cytochromes c modified at lysyl residues: preparation and characterization of the monosubstituted horse cytochromes c. beta-Thiopropionyl derivatives of horse cytochrome c singly modified at each of 18 different lysine epsilon-amino groups have been prepared using sulfosuccinimidyl-2-(biotinamido)ethyl-1,3-dithiopropionate and purified to homogeneity by high-pressure liquid chromatography. These derivatives were characterized by determination of: (i) the location of the modification; (ii) reduction potentials; (iii) visible and NMR spectra: and by (iv) measurement of electron transfer activity with cytochrome-c oxidase. No significant changes in structure were indicated, except for the ferric forms of the deri...
[Heart rate fluctuations in the horse at rest: (2) Biological variation factors related to behavioural profile]. Variability of heart rate was analysed within a sample of 30 horses in order to determine the influence of factors (animal, age, sex and breed) associated with different behavioural patterns. The heart rate fluctuations were analysed by computing the power spectral density of a series of heartbeat intervals. A generalized linear model was used to compare the spectra and to reveal the effect of each factor: animal age, sex and breed. The scale and shape of spectral densities were significantly different between animals. This individual variability was partly explained by the sex and age effects...