Biomedical subjects

H H Mantsch

Publications and source records attributed to H H Mantsch.

At least 55 records · Page 3Linked to original sources

PubMed1997

Prediction of fetal lung maturity from near-infrared spectra of amniotic fluid.

OBJECTIVE: The aim of this study was to attempt to correlate the quantitative data obtained from the near-infrared (near-IR) spectra of amniotic fluid with the lecithin/sphingomyelin (L/S) ratio determined from thin layer chromatographic analysis on the same samples, in order to establish the feasibility of near-IR spectroscopy as an alternate method for the prediction of fetal lung development. METHODS: Samples of amniotic fluid were obtained by amniocentesis from 52 patients between the 26th and 41st week of pregnancy. About 350 microliters of amniotic fluid was required to record the near-IR spectrum over the entire spectral range between 400 and 2500 nm using a commercial spectrometer. The L/S ratio was determined independently by thin layer chromatography (TLC) for each sample. For correlating the infrared spectra with the TLC-based L/S ratios, a partial least squares analysis was used. RESULTS: The L/S ratios predicted from the near-IR spectra of amniotic fluid were highly correlated and in excellent agreement with those determined by TLC (r = 0.91). CONCLUSION: Near-IR spectroscopy has the potential to become an alternate method to TLC for prediction of fetal pulmonary maturity.

Adult↗

PubMed1997

Infrared spectroscopy: a new frontier in medicine.

As we enter the second half of the nineties, one of the major challenges for biological infrared spectroscopists is to transfer the knowledge we have gained from studies on isolated molecules to the complex world of medicine. That it is possible to meet this challenge is suggested by comparison with the development of other biophysical techniques, such as magnetic resonance spectroscopy and imaging, which have already found their place in medical research and practice. The Spectroscopy Group in Winnipeg is developing and evaluating a variety of new IR techniques for the analysis of body fluids and tissues, both in vitro and in vivo. Herein, we review these methodologies, which comprise both instrumental (imaging and spatially localized IR spectroscopy) and interpretational procedures aimed at optimizing the measurements and their conversion to biodiagnostic information.

Diagnostic Imaging↗

PubMed1996

Thiocyanate levels in human saliva: quantitation by Fourier transform infrared spectroscopy.

A new quantitative method, based on Fourier transform infrared spectroscopy, was developed to evaluate the thiocyanate concentration in human saliva. Saliva samples were collected following a typical protocol and infrared spectra obtained from very small volumes (5 microl) deposited on a barium fluoride substrate. Exogenous potassium thiocyanate was used for calibration of the endogenous thiocyanate. This methodology does not require separation or extraction procedures. Human saliva spectra contain a characteristic marker band, due to thiocyanate, at 2058 cm-1. The integrated area of this band can be used for linear regression analysis and provides a good correlation between band area and thiocyanate concentration. Recovery of thiocyanate added to saliva was 100%. Centrifugation and dialysis experiments demonstrated that thiocyanate in saliva exists as a free or loosely bound ion. Saliva collected in the afternoon from 25 different subjects had a thiocyanate concentration of 0.83 +/- 0.42 (mean +/- SD) mmol/liter. In 4 subjects whose circadian pattern was investigated there was evidence of a higher thiocyanate concentration in saliva samples collected in the morning hours.

Cyanates↗

PubMed1996

Structural properties of UMP-kinase from Escherichia coli: modulation of protein solubility by pH and UTP.

UMP-kinase from Escherichia coli, unlike the analogous enzyme from eukaryotic organisms, is an oligomeric protein subjected to complex regulatory mechanisms in which UTP and GTP act as allosteric effectors. While the enzyme has an unusually low solubility at neutral pH (< or = 0.1 mg of protein/ mL), its solubility increases markedly above pH 8 and below pH 4. Furthermore, the solubility of the bacterial UMP-kinase at neutral pH is greatly enhanced in the presence of Mg-free UTP. Thermal denaturation experiments have demonstrated that UTP also increases the stability of the protein. Fourier-transform infrared spectroscopy and circular dichroism show that the secondary structure of the protein is the same at neutral and at alkaline pH. These data indicate that variations in enzyme solubility must be related to subtle changes in the tertiary and/or quaternary structure which modulate the exposure of hydrophobic surfaces in the protein molecule. A variant of UMP-kinase, obtained by site-directed mutagenesis (Asp159Asn), which is similar to the wild-type enzyme in its stability and kinetic properties, has a much increased water solubility (> 5 mg protein/mL) even at neutral pH. This suggests that salt bridges may be involved in the equilibrium between the soluble and aggregated forms of the wild-type enzyme, and that conformational changes induced upon binding of UTP increase the protein solubility by disrupting these salt bridges.

Allosteric Regulation↗

PubMed1996

Modification of the extracellular matrix following myocardial infarction monitored by FTIR spectroscopy.

Comparison of mid- and near-infrared spectra of control and infarcted rat ventricular tissue reveals the presence of absorptions in infarcted tissue which are highly characteristics of collagen, indicating large scale deposition of type I collagen in the myocardium following infarction. These results demonstrate that IR spectroscopy may be used to rapidly monitor the modifications of the extracellular matrix associated with myocardial infarction.

Animals↗

PubMed1996

Comparative analysis of the amino- and carboxy-terminal domains of calmodulin by Fourier transform infrared spectroscopy.

Fourier transform infrared spectra were obtained for mammalian calmodulin and two of its fragments produced by limited proteolysis with trypsin TR1C (1-77) and TR2C (78-148). Experiments were done in H2O, D2O and D2O/trifluoroethanol (TFE) mixtures. Information about secondary structure was obtained from analysis of the amide I and II bands; while characteristic absorbances for tyrosine, phenylalanine and carboxylate groups were analyzed for changes in tertiary structure. Our data indicate that the secondary and tertiary structure is preserved in the two half molecules of CaM, both in the apo- and Ca(2+)-saturated state. Addition of the structure-inducing solvent TFE causes marked changes only in the apo-TR1C domain. The maximum wavenumber for the amide I band of the two domains of CaM in D2O was markedly different (1642 cm-1 for TR1C versus 1646/1648 cm-1 for Ca2+ and apo-TR2C). This renders the amide I band for the intact protein very broad in comparison to that in other proteins and is indicative of a distribution of alpha-helices with slightly different hydrogen bonding patterns.

Amino Acid Sequence↗

PubMed1996

Quantitation of protein, creatinine, and urea in urine by near-infrared spectroscopy.

OBJECTIVES: To determine the feasibility of near-infrared analysis for quantitating urea, creatinine, and protein in urine. Practical advantages of this method include ease of sample presentation and the absence of reagents or disposables. DESIGN AND METHODS: The near-infrared methods were developed by first measuring the spectra of 123 different urine samples and, using independent clinical analyses, determining the protein, creatinine, and urea levels in each. Calibration models relating near-infrared spectroscopic features to those independently determined concentrations were optimized, and each model then validated using a set of 50 additional samples. RESULTS: Standard errors of calibration were 14.4 mmol/L, 0.66 mmol/L, and 0.20 g/L, and standard errors of prediction 16.6 mmol/L, 0.79 mmol/L, and 0.23 g/L, respectively, for urea, creatinine, and protein. CONCLUSIONS: Near-infrared urea quantitation is as accurate as the reference method, enzymatic (urease) conductivity, used here for calibration. Creatinine analysis is slightly less accurate relative to the reference (Jaffe rate) method; however, these errors can be minimized by careful attention to factors affecting precision. The accuracy of the near-infrared protein analysis cannot approach that of the reference method; nevertheless, the technique is potentially useful for coarse screening and for quantifying protein levels above 0.3 g/L.

Calibration↗

PubMed1996

Study of chronic lymphocytic leukemia cells by FT-IR spectroscopy and cluster analysis.

The peripheral mononuclear cells from 23 normal individuals and the purified B cells from 38 patients with chronic lymphocytic leukemia (CLL) were examined by Fourier transform infrared (FT-IR) spectroscopy. Differences were observed between the CLL and normal cells at the DNA, protein and lipid levels, with CLL cells having greater DNA and lower lipid contents than normal cells. In addition, the spectral character of the CLL and normal cells varied, demonstrating that there were also qualitative differences in the DNA and lipids. Statistical analysis, based on hierarchical clustering, separated normal from CLL cells completely and classified them into two subgroups for normal cells, while the CLL cells could be divided into three subgroups that were distinct from the normal cells. These differences were based on the lipid and DNA content and the overall spectral character of the cells.

B-Lymphocytes↗

PubMed1996

In situ characterization of beta-amyloid in Alzheimer's diseased tissue by synchrotron Fourier transform infrared microspectroscopy.

We report the first evidence of the structure of beta-amyloid protein as it exists in situ within a slice of human Alzheimer's diseased brain tissue. Using a Fourier transform infrared microspectroscopic technique, areas of interest can be selected for spectral measurements with regions of potential contamination masked. In so doing, it is possible to obtain infrared spectra only of beta-amyloid and not the surrounding grey matter within which it lies. However, to obtain spectra of high-quality signal-to-noise ratio using a conventional infrared source, we were limited to aperture sizes between 24 microns x 24 microns to 50 microns x 50 microns. Markedly improved high-quality spectra were acquired with infrared radiation provided by a synchrotron light source (National Synchrotron Light Source, Brookhaven National Laboratories), using aperture sizes as small as 12 microns x 12 microns. This allowed spectroscopic mapping of brain tissue regions containing amyloid. We observe that in situ protein of grey matter exist predominantly in an alpha-helical and/or unordered conformation, whereas within amyloid deposits a beta-sheet structure predominates. The hydrogen bonding strength of the beta-structure found in situ is different from that reported in the literature for isolated/chemically synthesized beta-amyloid peptides.

Alzheimer Disease↗

PubMed1996

Turn conformations in peptides containing the -Xaa-Ser- sequence.

The conformations of the protected dipeptides Boc-L-Pro-L-Ser-NHMe, Boc-L-Pro-D-Ser-NHMe, Boc-L-Val-L-Ser-NHMe and Boc-L-Val-D-Ser-NHMe have been explored through interpretation of their infrared spectra in CH2Cl2, DMSO and D2O solution. In CH2Cl2 solution the formation of a ten-membered ring (beta-turn) for each compound is signaled by characteristic shifts in both the urethane C = O and the terminal NH stretching frequencies. For each peptide, differences in the amide I absorption patterns for LL and LD isomers are consistent with the formation of type I and type II beta-turns respectively in CH2Cl2 solution. The amide I absorptions suggest substantial disruption of intramolecular hydrogen bonding in DMSO, and no intermolecular hydrogen bonding whatsoever in aqueous solution. In CH2Cl2 solution the OH stretching vibration is consistent with the formation of a hydrogen bond to the C = O of the serine group; however, two additional absorptions at frequencies characteristic of "free' OH groups also appear in all spectra. Implications regarding the serine in stabilizing the beta-turn are discussed.

Absorption↗

PubMed1996

Infrared spectroscopy of dystrophic mdx mouse muscle tissue distinguishes among treatment groups.

Four groups of mdx mice (deflazacort, high dose of 1.5 mg/kg and low dose of 0.75 mg/kg; prednisone, 1.0 mg/kg; and a placebo) were examined in a double-blind protocol. The experiments tested the hypothesis that infrared spectroscopy can distinguish among gastrocnemius muscle tissues derived from dystrophic animals (n = 22) from different treatment groups and from control muscle tissue (n = 23). Results showed that muscle, inflamed muscle, and tendon can be distinguished on the basis of their infrared absorption patterns. Distinctions among the spectra of the four treatment groups were sought with automated pattern-recognition methods. These classification methods, based either on spectral regions (900-1,500 cm-1) or on principal-component analysis, were in close agreement, assigning 15 or 16, respectively, of 22 mdx spectra to the correct treatment group. Both trials cleanly separated the high-dose deflazacort from the placebo group of muscles, whereas the prednisone and low-dose deflazacort groups were persistently confused in these classifications. Changes in the histology of muscle inflammation paralleled the spectral-classification results. Thus the proposed method, combining infrared spectroscopy with pattern-recognition algorithms, can distinguish treatment effects on muscle tissue. Specific spectral features characteristic of tissue type, disease progression, and treatment effects are not yet elucidated.

Algorithms↗

PubMed1995

Ribonuclease A revisited: infrared spectroscopic evidence for lack of native-like secondary structures in the thermally denatured state.

To address a number of conflicting reports in the literature, we undertook an infrared spectroscopic study to test for the presence of native-like secondary structures in thermally denatured ribonuclease A. Ribonuclease A does not aggregate at high temperatures, and the infrared spectrum shows a completely featureless amide I band contour. Using 13C-labeled urea, we were also able to obtain the infrared spectrum of the chemically denatured protein, which is practically identical with that of the heat-denatured protein. To the best of our knowledge, this is the first study that uses 13C-labeled urea as a chemical denaturant which circumvents the problem encountered with the strong absorption of urea in the conformation-sensitive amide I region of proteins; it opens up the possibility of investigating protein folding/unfolding processes in the presence of high concentrations of chemical denaturants. From an analysis of the amide I region of the infrared spectra of thermally and chemically denatured RNase A, it was concluded that heat-denatured ribonuclease A does not contain any significant amount of authentic hydrogen-bonded secondary structures. Furthermore, a comparison of the infrared spectra of ribonuclease A with those of ribonuclease T1 demonstrates that in spite of major differences between their native structures there are practically no differences between their heat-denatured states. This would not be expected if there were residual native-like secondary structures in the thermally denatured state of one or both of these proteins.

Carbon Isotopes↗

PubMed1995

Fourier-transform infrared spectroscopic analysis of rabbit lung surfactant: subfraction-associated phospholipid and protein profiles.

Surfactant obtained from bronchoalveolar lavage (BAL) can be separated into subfractions based on sedimentation characteristics. It has been suggested that the 10,000 x g, 60,000 x g and 100,000 x g subfractions isolated by this approach represent stages of surfactant extracellular processing. These three subfractions have been reported to differ in their morphology, composition and ability to lower surface tension. We wished to determine if infrared spectroscopy, which may be applied as a non-invasive technique could potentially prove useful for characterization and quantification of bronchoalveolar lavage (BAL) protein and phospholipid, and if this approach could detect differences in intermediate surfactant processing stages. Subfractions were collected from adult rabbit lungs by BAL and differential centrifugation and analyzed by Fourier transform infrared (FT-IR) spectroscopy. Biochemical assay of phospholipid and protein showed differences between subfractions that correlated well with the phospholipid/protein ratios obtained from FT-IR spectra (r = 0.939; r2 = 0.882). The subfraction sedimenting at 100,000 x g (P100) exhibited spectral shifts in the Amide I band, suggesting that the protein secondary structure was different compared to other fractions. Spectra obtained after separation of lipids and protein components showed an apparent disordering of protein secondary structure but little or no effect on the structure or mobility of phospholipids. These results support the idea that subfractions represent various processing stages of surfactant. In addition, they show that results from FT-IR analyses correlate significantly with traditional biochemical assay methods which may prove of clinical use.(ABSTRACT TRUNCATED AT 250 WORDS)

Amides↗

PubMed1995

FT-IR spectroscopy of methylmercury-exposed mouse lung.

Infrared spectroscopy which has traditionally been utilized by chemists and physicists for characterization and identification of the structural properties of chemical compounds is now becoming more relevant as a biodiagnostic tool. Recent reports suggest that arthritis and Alzheimer's disease can be diagnosed by using this technique. Changes associated with these diseases diagnosable with this technique are generally overt. In this study we have used 'Fourier transform infrared spectroscopy' (FT-IR) to analyze subtle changes in composition and structure of lipids and proteins in lung tissue, bronchoalveolar lavage and purified lamellar body fraction of mice exposed to methylmercury. Infrared measurements were made in attenuated total reflection mode using the Split Pea (Harrick Scientific Corporation, USA). Mice were treated with 4 doses of methylmercuric chloride (15 mg/kg body weight/dose), and control animals received an equivalent volume of physiological saline. Comparison of the control and experimental spectra revealed alterations in the intensities and frequencies of vibrational modes of lipids following methylmercury exposure. Results indicate that FT-IR spectral analyses may be a valuable tool for detecting subtle variations in biological components associated with drug exposure to lungs and, in particular may be very useful for assessing changes in bronchoalveolar lavage.

Animals↗

PubMed1995

Protein folding intermediates with rapidly exchangeable amide protons contain authentic hydrogen-bonded secondary structures.

Recent studies on protein folding intermediates by pulsed amide proton exchange and by far-ultraviolet circular dichroism have shown important discrepancies between the secondary structure contents estimated by these two methods at early folding stages. To solve these apparent discrepancies, structural studies have been performed on the isolated, 101 residue long, C-terminal proteolytic domain (F2) of the Escherichia coli tryptophan synthase beta chain, which had previously been reported to behave as an early folding intermediate [Chaffotte, A. F., Cadieux, C., Guillou, Y., & Goldberg, M. E. (1992) Biochemistry 31, 4303-4308]. The secondary structure of F2 has been investigated by far-UV circular dichroism (CD), Fourier transform infrared (FTIR) spectroscopy, and NMR. The CD and FTIR spectra clearly indicate that isolated F2 has about 30-45% of its residues involved in secondary structures stabilized by conventional hydrogen bonds. The characteristics of the NMR spectrum (line broadening, absence of structure-induced chemical shifts, absence of nuclear Overhauser effects in the amide region, few dipolar interactions between the side-chain protons) suggest that isolated F2 is oscillating between several conformations in rapid equilibrium. The rate of amide proton exchange has been studied by one-dimensional NMR, which indicates a significant extent of proton protection, with, however, protection factors that can be estimated to be at most 60 and more probably closer to 10. Thus, F2 appears to exist as a molten globule that exhibits very low amide proton protection and yet contains a large fraction of its residues involved in authentic secondary structures stabilized by hydrogen bonds. Such a state is likely to correspond to the earliest structured folding intermediates thus far characterized.

Amides↗

PubMed1995

Beware of connective tissue proteins: assignment and implications of collagen absorptions in infrared spectra of human tissues.

Infrared spectra of human central nervous system tissue and human breast carcinoma are presented. The spectra are discussed in terms of the composition of the tissues. It is shown that differences between spectra of white and grey matter can be rationalised on the basis of differences in lipid content. Spectra of the choroid plexus and arachnoid villus of the meninges show a series of absorptions not observed in other CNS tissue. These absorptions are discussed in terms of the connective tissue content of the samples. We demonstrate that the presence of collagen results in the appearance of a series of characteristic absorptions which may be mis-assigned as DNA phosphate absorptions. The implications of the presence of collagen in tissues for the diagnosis of disease states by IR spectroscopic methods, with particular reference to cancer, is discussed.

Brain Chemistry↗

PubMed1995

Arthritis diagnosis based upon the near-infrared spectrum of synovial fluid.

Synovial fluid aspirates have been characterized by measuring their visible/near-infrared spectra (400-2500 nm). The hypothesis tested in this study is that the spectra contain sufficient information to serve as an aid in the diagnosis and/or staging of arthritic disorders. The concentrations of all major constituents are carried implicitly in the spectra, and in this sense this approach is similar in spirit to conventional synovial fluid analysis. The distinguishing feature of this method is that we have not converted the raw data (spectra) explicitly to analytical information. Rather, we have used automated pattern recognition methods to identify significant characteristics of the spectra themselves. A total of 109 spectra were measured and split into three classes according to the disease (osteoarthritis, rheumatoid arthritis, or spondyloarthropathy) affecting the patient from whom the synovial fluid sample was taken. An automated classification method was then trained by correlating features derived from these spectra to the clinical diagnoses. The robustness of the classification was validated using the leave-one-out cross-validation method, i.e., by training on all but one of the spectra and using the resulting model to predict the classification for the spectrum that is left out. The result derived by following this procedure for each of the spectra was that 105 of the 109 predicted classifications correctly matched the clinical diagnosis. These results suggest that the near-infrared spectrum of synovial fluid is sufficient to allow diagnosis of the disease affecting the joint from which the aspirate is drawn.

Arthritis, Rheumatoid↗

PubMed1995

Neural network classification of infrared spectra of control and Alzheimer's diseased tissue.

Artificial neural network classification methods were applied to infrared spectra of histopathologically confirmed Alzheimer's diseased and control brain tissue. Principal component analysis was used as a preprocessing technique for some of these artificial neural networks while others were trained using the original spectra. The leave-one-out method was used for cross-validation and linear discriminant analysis was used as a performance benchmark. In the cases where principal components were used, the artificial neural networks consistently outperformed their linear discriminant counterparts; 100% versus 98% correct classifications, respectively, for the two class problem, and 90% versus 81% for a more complex five class problem. Using the original spectra, only one of the three selected artificial neural network architectures (a variation of the back-propagation algorithm using fuzzy encoding) produced results comparable to the best corresponding principal component cases: 98% and 85% correct classifications for the two and five class problems, respectively.

Alzheimer Disease↗