This thesis describes three automated precolumn derivatization procedures for the analysis of carboxylic group-containing compounds. After derivatization with a suitable label, the derivatives are separated on reversed-phashed HPLC and detected by fluorescence. ... Zie: Summar

Wolf, Johannes Hendrik

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Automated precolumn derivatization procedures in HPLC for biomedical and clinical applications

University of Groningen Research Database

  
 University of Groningen
Automated precolumn derivatization procedures in HPLC for biomedical and clinical
applications
Wolf, Johannes Hendrik
IMPORTANT NOTE: You are advised to consult the publisher's version (publisher's PDF) if you wish to cite from
it. Please check the document version below.
Document Version
Publisher's PDF, also known as Version of record
Publication date:
1992
Link to publication in University of Groningen/UMCG research database
Citation for published version (APA):
Wolf, J. H. (1992). Automated precolumn derivatization procedures in HPLC for biomedical and clinical
applications Groningen: s.n.
Copyright
Other than for strictly personal use, it is not permitted to download or to forward/distribute the text or part of it without the consent of the
author(s) and/or copyright holder(s), unless the work is under an open content license (like Creative Commons).
Take-down policy
If you believe that this document breaches copyright please contact us providing details, and we will remove access to the work immediately
and investigate your claim.
Downloaded from the University of Groningen/UMCG research database (Pure): http://www.rug.nl/research/portal. For technical reasons the
number of authors shown on this cover page is limited to 10 maximum.
Download date: 10-02-2018
This thesis describes three automated precolumn derivatization procedures for the
analysis of carborylic group-containing compounds. After derivatization with a suitable
label, the derivatives are separated on reversed-phase HPLC and detected by
fluorescence.
In the analysis of organic compounds chromatographic techniques play an predominant
role; for the analpis of polar compounds High Performance Liquid Chromatography,
HPLC, is the method of choice. A major disadvantage of the technique is the lack
of sensitive and selective detectors. Derivatization, chemical and/or physical alteration
of the analyte, is often used to enhance sensitivity or selectivity. In Chapter 1 the
pros and cons of derivatization are given. Next the automation of such reactions
is discussed.
Chapter 2 deals with use of 4-bromomethyl-7-methorycoumarin as labelling agent.
In Section 2.1 a review of the applications of this compound and its analogues as
derivatization agents is given. The next two sections discuss the optimization of the
reaction and its automation. In Section 2.2 the role of the various constituents of
the reaction mixture is reported along with the optimization of the HPLC separation;
analytical data such as repeatability, detection limits and linearity are presented.
The last section deals with the application of the method to the determination of
the anti-epilectical drug valproic acid in human serum and in rat blood and brain
dialysates. The present procedure for human serum is compared with an immuno-
assay method routinely used in clinical laboratories. The animal studies were carried
out to investigate the use of valproic acid as a model compound in pharmacological
studies on blood-brain passage. Valproate could be determined in both blood and
dialysates and the automated method has recently been used for the determination
of the cellular and extra-cellular concentration of valproic acid in freely moving rats
[50]. This study is not included in this thesis.
An important characteristic of 4-bromomethyl-7-methoxycoumarin is that it is only
reactive in aprotic, i.e. non-polar, solvents. For that reason two highly interesting
polar neuroactive compounds, N-acetylaspartate and N-acetylaspartylglutamate,
can not be determined by means of procedures involving this label. For these analytes
we developed a procedure using 2-aminoanthracene as label and carbodiimide as
coupling agent; this work is presented in Chapter 3. The reaction is fast and requires
only minimal sample pretreatment. N-Acetylaspartate and N-acetylaspartylglutamate
levels in several brain regions and spinal cord were similar to those so far reported.
Attempts to investigate whether the present procedure is more sensitive than the
well known reaction with o-phthaldialdehyde for the determination of amino acids
was not successful; the amino acids did not react with 2-aminoanthracene. An
I
91
increase in sensitivitywas, however, found using naphthalene-2,3-dicarboxaldehyde
as label. In Chapter 4 relevant analytical data are given on the automated procedure
for the <ietermination of several amino group-containing compounds uch as amino
acids, small peptides, brain amines and some drugs.
The present hesis shows that commercial equipment and reagents are available which
allow the set-up of fully automated on-line precolumn derivatization in combination
with HPLC and fluorescence detection. We believe that the approach of automated
on-line precolumn derivatization HPLC analysis carried out with, mainly, commer-
cially available quipment will become an attractive alternative to off-line precolumn
as well as on-line postcolumn procedures in many fields of research. Although the
examples in the present work all deal with (bio)medical applications, the procedures
can be easily be modified to be effective in areas such as environmental, agricultural
or pharmaceutical analysis.
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ARTS repository - University of Groningen

   University of GroningenAutomated precolumn derivatization procedures in HPLC for biomedical and clinicalapplicationsWolf, Johannes HendrikIMPORTANT NOTE: You are advised to consult the publisher's version (publisher's PDF) if you wish to cite fromit. Please check the document version below.Document VersionPublisher's PDF, also known as Version of recordPublication date:1992Link to publication in University of Groningen/UMCG research databaseCitation for published version (APA):Wolf, J. H. (1992). Automated precolumn derivatization procedures in HPLC for biomedical and clinicalapplications. s.n.CopyrightOther than for strictly personal use, it is not permitted to download or to forward/distribute the text or part of it without the consent of theauthor(s) and/or copyright holder(s), unless the work is under an open content license (like Creative Commons).The publication may also be distributed here under the terms of Article 25fa of the Dutch Copyright Act, indicated by the “Taverne” license.More information can be found on the University of Groningen website: https://www.rug.nl/library/open-access/self-archiving-pure/taverne-amendment.Take-down policyIf you believe that this document breaches copyright please contact us providing details, and we will remove access to the work immediatelyand investigate your claim.Downloaded from the University of Groningen/UMCG research database (Pure): http://www.rug.nl/research/portal. For technical reasons thenumber of authors shown on this cover page is limited to 10 maximum.Download date: 06-06-2022This thesis describes three automated precolumn derivatization procedures for theanalysis of carborylic group-containing compounds. After derivatization with a suitablelabel, the derivatives are separated on reversed-phase HPLC and detected byfluorescence.In the analysis of organic compounds chromatographic techniques play an predominantrole; for the analpis of polar compounds High Performance Liquid Chromatography,HPLC, is the method of choice. A major disadvantage of the technique is the lackof sensitive and selective detectors. Derivatization, chemical and/or physical alterationof the analyte, is often used to enhance sensitivity or selectivity. In Chapter 1 thepros and cons of derivatization are given. Next the automation of such reactionsis discussed.Chapter 2 deals with use of 4-bromomethyl-7-methorycoumarin as labelling agent.In Section 2.1 a review of the applications of this compound and its analogues asderivatization agents is given. The next two sections discuss the optimization of thereaction and its automation. In Section 2.2 the role of the various constituents ofthe reaction mixture is reported along with the optimization of the HPLC separation;analytical data such as repeatability, detection limits and linearity are presented.The last section deals with the application of the method to the determination ofthe anti-epilectical drug valproic acid in human serum and in rat blood and braindialysates. The present procedure for human serum is compared with an immuno-assay method routinely used in clinical laboratories. The animal studies were carriedout to investigate the use of valproic acid as a model compound in pharmacologicalstudies on blood-brain passage. Valproate could be determined in both blood anddialysates and the automated method has recently been used for the determinationof the cellular and extra-cellular concentration of valproic acid in freely moving rats[50]. This study is not included in this thesis.An important characteristic of 4-bromomethyl-7-methoxycoumarin is that it is onlyreactive in aprotic, i.e. non-polar, solvents. For that reason two highly interestingpolar neuroactive compounds, N-acetylaspartate and N-acetylaspartylglutamate,can not be determined by means of procedures involving this label. For these analyteswe developed a procedure using 2-aminoanthracene as label and carbodiimide ascoupling agent; this work is presented in Chapter 3. The reaction is fast and requiresonly minimal sample pretreatment. N-Acetylaspartate and N-acetylaspartylglutamatelevels in several brain regions and spinal cord were similar to those so far reported.Attempts to investigate whether the present procedure is more sensitive than thewell known reaction with o-phthaldialdehyde for the determination of amino acidswas not successful; the amino acids did not react with 2-aminoanthracene. AnI91increase in sensitivitywas, however, found using naphthalene-2,3-dicarboxaldehydeas label. In Chapter 4 relevant analytical data are given on the automated procedurefor the <ietermination of several amino group-containing compounds uch as aminoacids, small peptides, brain amines and some drugs.The present hesis shows that commercial equipment and reagents are available whichallow the set-up of fully automated on-line precolumn derivatization in combinationwith HPLC and fluorescence detection. We believe that the approach of automatedon-line precolumn derivatization HPLC analysis carried out with, mainly, commer-cially available quipment will become an attractive alternative to off-line precolumnas well as on-line postcolumn procedures in many fields of research. Although theexamples in the present work all deal with (bio)medical applications, the procedurescan be easily be modified to be effective in areas such as environmental, agriculturalor pharmaceutical analysis.Dit prordie een:fluorescIn de ateen belPerfornnadeelDerivatinteraclgevoelileen ov€ingegaaseringsHoofdsvoor arefmee rgrafenHierincompolals lineparagÍzvan valwaarbijOok wrratten,deze stin dit 1Eenbealleeninteresniet mhet reagebruilagens !monst(neurolgehaltt92

English

University of Groningen

   University of GroningenAutomated precolumn derivatization procedures in HPLC for biomedical and clinicalapplicationsWolf, Johannes HendrikIMPORTANT NOTE: You are advised to consult the publisher's version (publisher's PDF) if you wish to cite fromit. Please check the document version below.Document VersionPublisher's PDF, also known as Version of recordPublication date:1992Link to publication in University of Groningen/UMCG research databaseCitation for published version (APA):Wolf, J. H. (1992). Automated precolumn derivatization procedures in HPLC for biomedical and clinicalapplications. Groningen: s.n.CopyrightOther than for strictly personal use, it is not permitted to download or to forward/distribute the text or part of it without the consent of theauthor(s) and/or copyright holder(s), unless the work is under an open content license (like Creative Commons).Take-down policyIf you believe that this document breaches copyright please contact us providing details, and we will remove access to the work immediatelyand investigate your claim.Downloaded from the University of Groningen/UMCG research database (Pure): http://www.rug.nl/research/portal. For technical reasons thenumber of authors shown on this cover page is limited to 10 maximum.Download date: 12-11-2019This thesis describes three automated precolumn derivatization procedures for theanalysis of carborylic group-containing compounds. After derivatization with a suitablelabel, the derivatives are separated on reversed-phase HPLC and detected byfluorescence.In the analysis of organic compounds chromatographic techniques play an predominantrole; for the analpis of polar compounds High Performance Liquid Chromatography,HPLC, is the method of choice. A major disadvantage of the technique is the lackof sensitive and selective detectors. Derivatization, chemical and/or physical alterationof the analyte, is often used to enhance sensitivity or selectivity. In Chapter 1 thepros and cons of derivatization are given. Next the automation of such reactionsis discussed.Chapter 2 deals with use of 4-bromomethyl-7-methorycoumarin as labelling agent.In Section 2.1 a review of the applications of this compound and its analogues asderivatization agents is given. The next two sections discuss the optimization of thereaction and its automation. In Section 2.2 the role of the various constituents ofthe reaction mixture is reported along with the optimization of the HPLC separation;analytical data such as repeatability, detection limits and linearity are presented.The last section deals with the application of the method to the determination ofthe anti-epilectical drug valproic acid in human serum and in rat blood and braindialysates. The present procedure for human serum is compared with an immuno-assay method routinely used in clinical laboratories. The animal studies were carriedout to investigate the use of valproic acid as a model compound in pharmacologicalstudies on blood-brain passage. Valproate could be determined in both blood anddialysates and the automated method has recently been used for the determinationof the cellular and extra-cellular concentration of valproic acid in freely moving rats[50]. This study is not included in this thesis.An important characteristic of 4-bromomethyl-7-methoxycoumarin is that it is onlyreactive in aprotic, i.e. non-polar, solvents. For that reason two highly interestingpolar neuroactive compounds, N-acetylaspartate and N-acetylaspartylglutamate,can not be determined by means of procedures involving this label. For these analyteswe developed a procedure using 2-aminoanthracene as label and carbodiimide ascoupling agent; this work is presented in Chapter 3. The reaction is fast and requiresonly minimal sample pretreatment. N-Acetylaspartate and N-acetylaspartylglutamatelevels in several brain regions and spinal cord were similar to those so far reported.Attempts to investigate whether the present procedure is more sensitive than thewell known reaction with o-phthaldialdehyde for the determination of amino acidswas not successful; the amino acids did not react with 2-aminoanthracene. AnI91increase in sensitivitywas, however, found using naphthalene-2,3-dicarboxaldehydeas label. In Chapter 4 relevant analytical data are given on the automated procedurefor the <ietermination of several amino group-containing compounds uch as aminoacids, small peptides, brain amines and some drugs.The present hesis shows that commercial equipment and reagents are available whichallow the set-up of fully automated on-line precolumn derivatization in combinationwith HPLC and fluorescence detection. We believe that the approach of automatedon-line precolumn derivatization HPLC analysis carried out with, mainly, commer-cially available quipment will become an attractive alternative to off-line precolumnas well as on-line postcolumn procedures in many fields of research. Although theexamples in the present work all deal with (bio)medical applications, the procedurescan be easily be modified to be effective in areas such as environmental, agriculturalor pharmaceutical analysis.Dit prordie een:fluorescIn de ateen belPerfornnadeelDerivatinteraclgevoelileen ov€ingegaaseringsHoofdsvoor arefmee rgrafenHierincompolals lineparagÍzvan valwaarbijOok wrratten,deze stin dit 1Eenbealleeninteresniet mhet reagebruilagens !monst(neurolgehaltt92

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Automated precolumn derivatization procedures in HPLC for biomedical and clinical applications

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