Separation of isotopically labeled molecules by micro high performance liquid chromatography with a packed micro glass capillary column

Jinno, Kiyokatsu; Hirata, Yukio; Hiyoshi, Yoshihiko

doi:10.1002/jhrc.1240050210

Cited by 18 publications

(5 citation statements)

References 8 publications

(1 reference statement)

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“…Edmonds et al also observed the decreased retention times with small peptides and nucleosides on a C18 stationary phase with a 50/50 ACN/D 2 O mobile phase − All amino acids have at least three exchangeable protons. A decrease in the retention factor between 0.3 and 1% per D atom was observed for different molecules. − This effect will account for a part of the observed decreased retention times of the amino acids.…”

Section: Resultsmentioning

confidence: 95%

“…Several authors found that, when isotopomers are separated on C18 columns using an aqueous mobile phase, the most deuterated compound elutes first. − All amino acids have at least three exchangeable protons. A decrease in the retention factor between 0.3 and 1% per D atom was observed for different molecules. − This effect will account for a part of the observed decreased retention times of the amino acids. The additional difference in our case is that, unlike the C18 stationary phase, the teicoplanin stationary phase is deuterated itself to a significant extent.…”

Section: Resultsmentioning

confidence: 99%

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Retention and Selectivity of Teicoplanin Stationary Phases after Copper Complexation and Isotopic Exchange

et al. 2001

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Teicoplanin is a macrocyclic glycopeptide that is highly effective as a chiral selector for LC enantiomeric separations. Two possible interaction paths were investigated and related to solute retention and selectivity: (1) interactions with the only teicoplanin amine group and (2) role of hydrogen bonding interactions. Mobile phases containing 0.5 and 5 mM copper ions were used to try to block the amine group. In the presence of copper ions, it was found that the teicoplanin stationary phase has a decreased ability to separate most underivatized racemic amino acids. However, it maintained its ability to separate enantiomers that were not alpha-amino acids. It is established that there is little copper-teicoplanin complex formation. The effect of Cu2+ on the enantioseparation of some alpha-amino acids appears to be due to the fact that these solutes are good bidentate ligands and form complexes with copper ions in the mobile phase. Isotopic exchange with deuterium oxide was performed using acetonitrile-heavy water mobile phases. It was found that the retention times of all amino acids were lower with deuterated mobile phases. The retention times of polar or apolar molecules without amine groups were higher with deuterated mobiles phases. In all cases, the enantioselectivity factors were unaffected by the deuterium exchange. It is proposed that the electrostatic interactions are decreased in the deuterated mobile phases and the solute-accessible stationary-phase volume is somewhat swollen by deuterium oxide. The balance of these effects is a decrease in the amino acid retention times and an increase in the apolar solute retention time. The enantioselectivity factors of all of the molecules remain unchanged because all of the interactions are changed equally. We propose a new global quality criterion (the E factor) for comparing and evaluating enantiomeric separations.

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Section: Resultsmentioning

confidence: 95%

Section: Resultsmentioning

confidence: 99%

Retention and Selectivity of Teicoplanin Stationary Phases after Copper Complexation and Isotopic Exchange

et al. 2001

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“…Semipreparative HPLC steroid purification methods combined with optimum GC–C–IRMS conditions can produce excellent chromatographic resolution required for the accurate reporting of δ 13 C values. Consideration is needed, however, of the ability of reversed‐phase HPLC to effectively separate isotopologs60 and how this extends intuitively to the purification of steroids as a potential source of error. Indeed, as important as careful peak definition in the GC–C–IRMS analysis is a prerequisite for accurate results, chromatographic separation by HPLC has been shown to significantly alter the 13 C content of a steroid when the entire peak was not collected 61…”

Section: Issues For Doping Controlmentioning

confidence: 99%

The application of carbon isotope ratio mass spectrometry to doping control

2008

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The administration of synthetic steroid copies is one of the most important issues facing sports. Doping control laboratories accredited by the World Anti-Doping Agency (WADA) require methods of analysis that allow endogenous steroids to be distinguished from their synthetic analogs in urine. The ability to measure isotope distribution at natural abundance with high accuracy and precision has increased the application of Gas Chromatography-Combustion-Isotope Ratio Mass Spectrometry (GC-C-IRMS) to doping control in recent years. GC-C-IRMS is capable of measuring the carbon isotope ratio (d 13 KEYWORDS: endogenous steroids; doping control; carbon isotope ratio (υ 13 C); gas chromatography-combustion-isotope ratio mass spectrometry (GC-C-IRMS) THE ENDOGENOUS STEROID CHALLENGEDrug use and abuse is an unfortunate feature of sports. The fight against doping in sports is a challenge faced by governments, sports authorities, laboratories, coaches and athletes who wish to contribute to fair, healthy and legal competition. The abuse of synthetic steroid copies is one of the most important issues for doping control. Before 1982, only the use of xenobiotic steroids had been banned, with a positive result requiring unequivocal proof provided by techniques such as GC-MS that the banned substance and/or its metabolites were present in a urine sample. Following the prohibition of testosterone (T; androst-4-ene-17ˇ-ol-3-one) administration by the International Olympic Committee and subsequently by the World Anti-Doping Agency (WADA), 1 new areas of research have been established within doping control laboratories.Methods for the detection of administered synthetic steroids by molecular spectrometry techniques such as GC-MS are limited by their ability to identify and quantify. The GC-MS steroid profiles are, however, an important Ł Correspondence to: Adam T. Cawley, National Measurement Institute, 1 Suakin Street, Pymble NSW 2073, Australia. E-mail: Adam.Cawley@measurement.gov.au means of screening all urine samples entering the laboratory to eliminate those of nonsuspicious nature. The objective for doping control laboratories is, therefore, to maximize the detection of synthetic steroid doping violations while minimizing the additional resources required from stakeholders. This can be achieved with advanced GC-MS steroid profiling strategies based on the analysis of several urinary steroids that can originate from the endocrine system. 2 -7 Control of any endogenous substance requires the establishment of 'normal' reference intervals and subsequent statistical determination of what constitutes an 'abnormal' state. The challenge for doping control is presented by steroid administration altering the human endocrine system in ways that are not yet fully understood, since they are largely dependent on the individual athlete.Synthetic copies of endogenous steroids, namely, dehydroepiandrosterone [(DHEA); androst-5-ene-3ˇ-ol-17-one], androstenedione (ADIONE; androst-4-ene-3,17-dione), 4-androstenediol (4-ADIOL; androst-4-ene-3ˇ,17...

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“…have prepared packed microcapillary columns which are semipermeable, and operated extremely long columns up to 100 m under moderate pressure [3-51. Recently, fused silica [6,7] and Pyrex glass capillaries [8,9] have been utilized as column materials and effectively packed with microparticles. While instrumentation suitable for each type of column, including sampling, detection, and connection system, is necessary to realize its efficiency, column materials as well as packing procedures seem to be an important factor in preparing such long, efficient columns.…”

Section: Introductionmentioning

confidence: 99%

High resolution liquid chromatography with a packed micro glass capillary column

Hirata

Jinno

1983

J. High Resol. Chromatogr.

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SummarySingle, long columns (1-5 m) can be prepared efficiently using reversed phase packings (3-10 pm particle diameter). 1-m columns packed with 3 and 10 pm packing provide 1 10 000 and 50 000 theoretical plates, respectively. Very efficient columns can resolve highly complex mixtures and difficult-to-separate compounds. Temperature gradient elution is a powerful technique for LC with microbore columns.

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Separation of isotopically labeled molecules by micro high performance liquid chromatography with a packed micro glass capillary column

Cited by 18 publications

References 8 publications

Retention and Selectivity of Teicoplanin Stationary Phases after Copper Complexation and Isotopic Exchange

Retention and Selectivity of Teicoplanin Stationary Phases after Copper Complexation and Isotopic Exchange

The application of carbon isotope ratio mass spectrometry to doping control

High resolution liquid chromatography with a packed micro glass capillary column

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