Biomedical subjects

D W Armstrong

Publications and source records attributed to D W Armstrong.

At least 73 records · Page 4Linked to original sources

PubMed1995

Determination and use of Rohrschneider-McReynolds constants for chiral stationary phases used in capillary gas chromatography.

The overall polarity of 22 chiral stationary phases (CSPs) used in capillary open tubular gas chromatography columns was estimated using the first five Rohrschneider-McReynolds constants. Most of the columns (i.e., 18) were wall-coated, and four were of the wall-immobilized or of the so-called "bonded" type. A wall-coated capillary squalane column was specially prepared as a polarity reference column. All but two of the CSPs were based on derivatized cyclodextrins (CDs) of different sizes. The overall properties of the CSPs are discussed in terms of the five Rohrschneider-McReynolds constants and their average values. It was found that the derivatized cyclodextrin CSP polarity increased with the CD ring size. The bonded CSPs were significantly less polar than their coated homologues due to the apolar polymer used to immobilize the CD rings. The retention behavior of 14 compounds was studied at 100 degrees C on the capillary columns. Retention parameters are clearly related to the McReynolds constants. Conversely, the enantiomeric resolution capability of a given stationary phase is not related to the constants. The enantioselective resolution mechanism critically depends on the solute structure and on the nature of the CSP.

Chromatography, Gas↗

PubMed1995

Highly enantioselective capillary electrophoretic separations with dilute solutions of the macrocyclic antibiotic ristocetin A.

Ristocetin A is one of a series of structurally related amphoteric, glycopeptide, macrocyclic antibiotics. These compounds have several features that make them attractive as chiral selectors. These include spatially oriented functional groups that are known to provide the types of interactions that are conducive to enantio-recognition, a somewhat rigid "pocket" that can provide a site for hydrophobic interactions and polar, flexible arms (i.e., pendent sugar moieties) that can rotate to hydrogen bond and otherwise interact with a variety of chiral analytes. In addition, these compounds are sufficiently soluble in water, aqueous buffers and aqueous-organic solvents that are commonly used in capillary electrophoresis (CE). The use and optimization of ristocetin A as a chiral selector in CE is discussed. Over 120 racemates are resolved including a variety of N-blocked amino acids, non-steroidal anti-inflammatory compounds and a large number of biologically important compounds containing carboxylic acid groups (e.g., mandelic acid derivatives, lactic acid derivatives, folinic acid, tropic acid).

Carbohydrate Sequence↗

PubMed1994

Use of a macrocyclic antibiotic, rifamycin B, and indirect detection for the resolution of racemic amino alcohols by CE.

Ansamycins are a very specific class of macrocyclic antibiotics of which the rifamycins are among the better known members. Rifamycins bind to and inhibit DNA polymerase. Rifamycin B (the most easily obtained ansamycin) is negatively charged and is shown to associate with and enantioselectively resolve several chiral amino alcohols including terbutaline, isoproterenol, bamethan, metaproterenol, synephrine, metanephrine, salbutamol, epinephrine, norphenylephrine, ephedrine, psi-ephedrine, octopamine, norepinephrine, normetanephrine, metoprolol, alprenolol, atenolol, and oxprenolol. A description of the structure and properties of rifamycins, in general, and rifamycin B, in particular, is given. The complexation and chiral recognition of the aforementioned racemic compounds by rifamycin B is afforded by multiple interactions of which charge-charge, hydrogen-bonding, and hydrophobic inclusion interactions most likely dominate in hydroorganic solvents. The effect of various experimental factors on enantiomeric resolution is discussed in terms of optimizing the CE separations. Since most chiral antibiotic macrocycles are ionizable, somewhat flexible, and contain hydrophobic and hydrophilic moieties, they tend to be significantly affected by variations in the solution environment.

Amino Alcohols↗

PubMed1994

Sensitive enantiomeric separation of aliphatic and aromatic amines using aromatic anhydrides as nonchiral derivatizing agents.

New pre-column derivatizing reagents: phthalic anhydride, 3-nitrophthalic anhydride, diphenic anhydride, 1,8-naphthalic anhydride and diphenylmaleic anhydride have been developed for resolving chiral compounds having amine groups. Although all of these agents produce derivatives with high molar absorptivities, the later two also fluoresce. Upon derivatization, aromatic analytes containing free carboxylic groups are produced. Both of these moieties enhance chiral recognition on cyclodextrin-based columns. The derivatization reaction is carried out at room temperature by shaking a buffered aqueous solution of a sample with an acetonitrile solution of the reagent. The reaction is fast and proceeds without any detectable racemization. The labeled compounds have favorable chromatographic properties which are demonstrated by resolution of a number of chiral compounds on cyclodextrin-bonded phases operated with non-aqueous polar organic eluents. The selectivity and good efficiency of this system contributes to its high sensitivity and in its applicability for detecting low levels of enantiomeric impurities. The detection limit is in the picomole range and less than 0.1% enantiomeric impurities can be determined in some cases.

Amines↗

PubMed1994

Evaluation of enantiomeric purity of selected amino acids in honey.

Highly sensitive and accurate HPLC methods were used for the determination of total amounts of proline, leucine and phenylalanine and their enantiomeric ratios in a variety of different honey samples. Significant amounts of D-leucine and D-phenylalanine and relatively low concentrations of D-proline have been found in honeys of different botanical and geographical origins. It is suggested that the enantiomeric ratios of amino acids could be used to test for storage effects, age, and the quality of the processing of the honey.

Amino Acids↗

PubMed1994

Evaluation of free D-glutamate in processed foods.

Monosodium glutamate (MSG) is added to many processed foods at significant levels for flavor enhancement. It is also naturally occurring at high levels in some foods. The enantiomeric composition of free glutamate in foods was examined and all foods analyzed were found to contain D-glutamate. The relative percent of D-glutamate in the food products studied depended on the origin of the glutamate. Foods to which MSG was added by the manufacturer had a high total level of MSG but a lower relative percentage of the D-enantiomer (usually less than 0.8%). In comparison, fermented foods tend to have high relative levels of D-glutamate but a lower total amount of the amino acid. The relative percent of D-glutamate in nonfermented foods containing no added MSG was also found to be low compared to fermented products. In some cases the percent D-glutamate could be related to the relative amounts of other food ingredients such as cheese.

Fermentation↗

PubMed1994

Evaluation of the macrocyclic antibiotic vancomycin as a chiral selector for capillary electrophoresis.

Vancomycin is one of a family of related macrocyclic glycopeptide antibiotics that were discovered by scientists at the Eli Lilly Company in the 1950s. It has been used to treat severe staphylococcal infections, particularly when bacterial resistance to other antibiotics has developed. Vancomycin is a naturally occurring chiral compound and has a number of stereogenic centers. Furthermore, it contains a variety of functionalities that are known to be useful for enantioselective interactions (e.g., hydrogen bonding groups, hydrophobic pockets, aromatic groups, amide linkages, etc.). The physiochemical properties of vancomycin, including its stability in solution, are discussed as they pertain to capillary electrophoresis. Over 100 racemates were resolved including many nonsteroidal antiinflammatory drugs, antineoplastic compounds and N-derivatized amino acids. Many of these compounds had very high resolution factors. Optimization and the effect of different experimental parameters on the enantioselective separations are discussed.

Amino Acid Sequence↗

PubMed1993

Enantiomeric separation of fluorescent, 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate, tagged amino acids.

A new derivatizing reagent, 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate (AQC), has been used successfully for chromatographic enantioseparation of 31 amino acids on cyclodextrin bonded stationary phases. AQC reacts with both primary and secondary amino acids to produce stable and highly fluorescent derivatives suitable for efficient and sensitive chromatographic determinations. The derivatization reaction proceeds without detectable racemization. The detection limit is in the femtomole range and approximately 0.0075% of the D-enantiomer in an excess of the L-enantiomer is detectable. High resolution values are needed when determining trace enantiomeric impurities.

Amino Acids↗

PubMed1993

High-performance liquid chromatographic enantioseparation of glycyl di- and tripeptides on native cyclodextrin bonded phases. Mechanistic considerations.

Di- and tripeptides containing at least one glycine moiety were separated into enantiomers on alpha-, beta- and gamma-cyclodextrin bonded phases after their precolumn derivatization with 9-fluorenylmethyl chloroformate (FMOC-Cl). It is shown that the choice of a suitable cyclodextrin bonded phase used with a nonaqueous polar mobile phase offers a wide range of possibilities to optimize enantioselectivity. Use of the FMOC derivative greatly enhances sensitivity, stability and enantioselectivity. It is postulated that under reversed-phase conditions the inclusion complex formation between the hydrophobic aromatic part of the FMOC-functionalized peptides and non-polar cyclodextrin cavity interior is the major factor contributing to retention. However, under these conditions there seems to be insufficient interaction between the hydrophilic peptide chain and cyclodextrin hydroxyls. Hence no chiral recognition and enantiomeric separation is observed. In systems operating with polar organic mobile phases the inclusion complex is suppressed as the cyclodextrin cavity is largely occupied by the mobile phase. The enantioselectivity observed is caused by hydrogen bonding between peptide chain and the hydroxyl groups at the mouth of the cyclodextrin. The stereospecific interactions depend very strongly on the cyclodextrin size, the length of the peptide chain and the mobile phase composition.

Amino Acid Sequence↗

PubMed1993

D-amino acid levels in human physiological fluids.

Plasma, urine, cerebrospinal fluid (CSF), and amniotic fluid were examined to determine whether free D-amino acids were present and if so at what levels. It was found that D-amino acids exist in all physiological fluids tested, but that their level varied considerably. The lowest levels of D-amino acids were usually found in amniotic fluid or CSF (almost always < 1% of the corresponding L-amino acid). The highest levels were found in urine (usually tenth percent to low percent levels). Pipecolic acid seemed to be different from the other amino acids tested in that it was excreted primarily as the D-enantiomer (often > 90%). Correspondingly high levels of D-pipecolic acid were not found in plasma. Some of the trends found in this work seemed to be analogous to those found in a recent rodent study.

Adult↗

PubMed1993

Relevance of enantiomeric separations in environmental science.

A significant number of all organic chemicals that are released into the environment are racemic mixtures. Most environmental regulations and scientific environmental studies treat racemic mixtures as though they were single, pure compounds. This can lead to incorrect toxicological, distribution, degradation and other data. A series of new enantioselective chromatographic techniques have been developed that allow the facile separation and quantitation of chiral compounds of environmental importance. Nineteen racemic compounds that have been or currently are being released to the environment are resolved. These include: rodenticides--Warfarin, Coumachlor and Coumafuryl; insecticides--Crufomate, Bulan, Fonofos, Mitotane; insect repellent--Ethohexadiol; herbicides and fungicides--Ancymidol, Silvex, Napropamide, phenyl mercuric lactate, 2-[3-chlorophenoxy]propionamide, and 2-chloropropionic acid; and halocarbons-1,2-dichloropropane, 2-bromo-1-chloropropane, 1,2-dibromo-3-chloropropane, 2,3-dichlorobutane and alpha-1,2,3,4,5,6-hexachlorocyclohexane. Several examples are given to illustrate the importance of enantioselective measurements of these and other compounds. Choosing the proper chromatographic technique and chiral stationary phase based on analyte structure is also discussed.

Journal Article↗

PubMed1993

Stereochemistry of pipecolic acid found in the urine and plasma of subjects with peroxisomal deficiencies.

Recently it was found that normal adults excrete pipecolic acid primarily as the D-enantiomer even though it is present in the blood stream mainly as the L-enantiomer (i.e. > 98% L). This study of pipecolic acid stereochemistry was extended to subjects with peroxisomal deficiencies since they are known to have high levels of pipecolic acid in their physiological fluids. Also, pipecolic acid stereochemistry was examined in young normal subjects since this group was not considered previously. It was found that the stereochemical composition of pipecolic acid in plasma was very similar for all subjects tested (i.e. > 98% of the L-enantiomer). However, the stereochemical composition of excreted pipecolic varied considerably. Urine samples from subjects with the most severe peroxisomal deficiency, i.e. cerebralhepatorenyl (Zellweger) syndrome (CHRS) contained little D-pipecolic acid. In fact the enantiomeric ratios for pipecolic acid in the urine and plasma of these subjects were very similar. This was not the case for normal subjects. Levels of D-pipecolic acid in the urine of subjects with 'less severe' peroxisomal deficiencies tended to be somewhat higher but they did not approach the levels found in normal adults. Several possible reasons for these results are discussed.

Adult↗

PubMed1992

Pressure-induced conformational changes in an antigen and an antibody and the implications on their use for hyperbaric immunoadsorption.

Pressure-induced conformational changes in two proteins, bovine serum albumin (BSA) and immunoglobulin G (IgG), were studied to assess the application of hyperbaric manipulation to the dissociation of antigen-antibody complexes. Antigen-antibody dissociation is important in the product-recovery phase of immunoadsorption, an affinity purification process. Three techniques were used in parallel for this study, including fluorescence, Fourier transform infrared (FTIR) spectroscopy, and the enzyme-linked immunosorbent assay (ELISA). Employing a fluorescent probe, fluorescent intensity measurements were used to detect protein conformational changes. FTIR spectroscopy was used to determine changes in protein secondary structure induced by high pressure, while the ELISA test was used to examine antibody recognition after the proteins had been pressure-treated. The results from this work demonstrate that IgG is resistant to conformational changes induced by pressures below 2 kbar. In contrast, BSA undergoes reversible conformational changes in this pressure range. However, these conformational changes are not reflected in tests measuring antibody recognition. These findings indicate that IgGs have the potential to be used as recycled ligands in immunoadsorption separation processes. Different antigens that are being considered for purification by immunoadsorption and separated by means of high pressure could be screened by the methods disclosed to determine their stability under high pressure conditions.

Adsorption↗

PubMed1992

FTIR spectroscopic kinetic analysis of alkaline phosphatase under hyperbaric manipulation.

For the first time, high-pressure infrared spectroscopy has been used in an enzyme kinetics study. This technique allows not only the investigation of kinetics under very high pressure, but it also allows simultaneous determinion of changes in the secondary structure of enzymes at the corresponding pressures. In the present study, a classical enzyme reaction, the conversion of p-nitrophenol phosphate into p-nitrophenol by alkaline phosphatase was selected to demonstrate the potential of infrared spectroscopy as an alternative physical method in the high-pressure study of enzyme kinetics. The rate constants of this enzyme reaction have been determined as a function of pressure in the pressure range 0.001-14 kbar. The first-order rate constants thus obtained increases with increasing pressure up to 8.3 kbar. At this pressure, the reaction rate decreases abruptly due to the denaturation of the enzyme arising from the conformational changes of some alpha-helical segments in the enzyme molecules into beta-sheet structure. The present results suggest that the pressure-enhanced overall hydrogen-bond strength in the amide groups of the enzyme is one of the factors which stimulate the enzyme activity. Moreover, the dissociation of the dimeric enzyme into its subunits does not inhibit the enzyme activity but only attributes to a slight change in activation volume.

Alkaline Phosphatase↗

PubMed1992

Efficient enantioselective separation and determination of trace impurities in secondary amino acids (i.e., imino acids).

An R-(-)-1-(1-naphthyl)ethyl carbamoylated-beta-cyclodextrin bonded phase in conjunction with a nonaqueous polar mobile phase was used for the highly selective enantioseparation of a number of secondary amino acids after their pre-column derivatization with 9-fluorenylmethyl chloroformate (FMOC). Under the conditions employed, the FMOC reagent served to "lock" the imino acid into their existing conformation thereby preventing the possibility of racemization. Furthermore, it served to increase the sensitivity to the point that trace level enantiomeric impurities were easily detected. Compared with separations that use traditional reversed-phase solvents, this method showed several advantages: higher selectivity towards the imino acid enantiomers investigated, shorter analysis times, faster equilibration of the column, more stable baseline and more sensitive fluorescence detection. The detection limits for FMOC derivatives of proline, trans-4-hydroxyproline, cis-4-hydroxyproline, pyroglutamic acid, 3,4-dehydroproline, thiaproline, penicillamine acetone adduct and pipecolic acid are in the low femtomole range. The method was used for evaluation of enantioselectivity of a number of "optically pure" commercial imino acid standards. Enantiomeric impurities as low as 0.0001% (1 ppm) can be determined in some cases. High precision determination of trace levels of D-imino acids in the presence of large amounts of corresponding (opposite) L enantiomer at 1, 0.1, 0.01% and below are demonstrated.

Amino Acids↗

PubMed1992

Chiral separation of nipecotic acid amides.

1-Decyl-3-(N,N-diethylcarbamoyl)piperidine (1) and alpha,alpha'-bis[3-(N-benzyl-N-methylcarbamoyl)piperidinol]-p-xyle ne (2) represent mono-N-substituted and bis-N-substituted carbamoylpiperidines, or nipecotic acid amides, respectively. Initially, several attempts were made to resolve these compounds using beta-cyclodextrin, cellulose carbamate and Pirkle-type columns. However, the interactions of the stereoisomers of the two compounds with these stationary phases did not differ enough to permit satisfactory separations. Baseline resolution was achieved using an alpha 1-acid glycoprotein (AGP) chiral column. The mobile phase was phosphate buffer (pH 7.0). Tetrabutylammonium (TBA) was used as the cationic modifier and ethanol as the uncharged modifier. Circular dichroism was used to identify the enantiomers. Compound 1 was resolved into positive and negative enantiomers and 2 into positive and negative enantiomers and a meso diastereomer. The influence of pH, buffer ionic strength, cationic and uncharged modifier concentrations on retention, chiral selectivity and resolution were evaluated. Based on the results, it is suggested that both ionic and hydrophobic interactions may be responsible for retention and resolution.

Buffers↗

PubMed1992

Empirical procedure that uses molecular structure to predict enantioselectivity of chiral stationary phases.

A total of 121 racemic compounds were separated in the normal-phase mode on a (S)-(1-naphthylethyl)carbamoylated beta-cyclodextrin (S-NEC-beta-CD) bonded phase and 74 on the R equivalent (R-NEC) chiral stationary phase (CSP). All compounds are of the type that have four substituents on a stereogenic center, rather than an "axis of chirality". It is shown that the binary solvent pair used as the mobile phase has a significant influence on chiral recognition. However, the proportions of the components of a specific pair have little effect. From the results, the individual contributions to chiral recognition by these CSPs were estimated for 81 different substituents of the stereogenic center. Varying the arrangement of these 81 substituents could produce over 1.6 million compounds. Hydrogen was chosen as the reference substituent and was assigned a 0 cal/mol free energy. The chiral recognition increased when sp2-hybridized carbons were connected to the stereogenic center. Conversely, sp3-hybridized carbons decreased the enantioselectivity. Amido groups increased the chiral recognition, especially when associated with pi-acid (3,5-dinitrobenzoyl) or pi-basic (naphthyl) groups. This approach does not allow one to know which enantiomer elutes first. However, the "substituent energy" list for chiral compounds can be used to obtain an estimated value for the enantioselectivity of a compound by adding the energy contributions of the four substituents connected to the stereogenic center. In this way one can predict a priori whether or not a compound will separate on a CSP and estimate its separation factor (alpha). Theoretically, this approach can be used for most CSPs, provided a sufficient data base is generated on them.

Carbamates↗

PubMed1992

Human cold air habituation is independent of thyroxine and thyrotropin.

Thyroxine (T4) is required in species possessing brown adipose tissue (BAT) for the maintenance of cold tolerance and adaptation. In humans, who possess negligible quantities of BAT, the importance of T4 has not been demonstrated. We studied the effects of decreased serum T4 and thyrotropin (TSH) on human cold habituation after repeated cold air exposures. Eight men (T3+) received a single daily dose of triiodothyronine (T3; 30 micrograms/day), and another eight men (T3-) received a placebo. All 16 normal thyroid men underwent a standardized cold air test (SCAT) under basal conditions in January and again in March after eighty 30-min 4.4 degrees C air exposures (10/wk). Measurements of basal metabolic rate (BMR), O2 consumption (VO2), mean arterial pressure (MAP), plasma norepinephrine (NE), serum TSH, free and total T4, and free and total T3 were repeated before and after 8 wk of exposure. TSH, free T4, and total T4 were 50% lower for T3+ than for T3- subjects. Total and free T3 were not different between groups. BMR was unchanged after habituation, whereas the cold-stimulated VO2, MAP, and NE were significantly reduced for all subjects in March. The relationship between VO2 and NE (r2 = 0.44, P less than 0.001) during the initial SCAT was unchanged with habituation. We suggest that human cold habituation is independent of major changes in circulating T4 and TSH.

Acclimatization↗