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Evaluation of ethylammonium nitrate as background electrolyte in capillary electrophoresis
KTH, School of Chemical Science and Engineering (CHE), Chemistry, Applied Physical Chemistry. (Analytisk kemi)
KTH, School of Chemical Science and Engineering (CHE), Chemistry, Applied Physical Chemistry.
KTH, School of Chemical Science and Engineering (CHE), Chemistry, Applied Physical Chemistry. KTH, Superseded Departments (pre-2005), Chemistry.ORCID iD: 0000-0002-3444-9987
(English)Manuscript (preprint) (Other academic)
National Category
Analytical Chemistry
Research subject
Chemistry
Identifiers
URN: urn:nbn:se:kth:diva-215628OAI: oai:DiVA.org:kth-215628DiVA, id: diva2:1148463
Note

QC 20171013. QC 20191018

Available from: 2017-10-11 Created: 2017-10-11 Last updated: 2019-10-18Bibliographically approved
In thesis
1. Bioanalytical separation using capillary electrophoresis: Applications with microbubbles and proteins
Open this publication in new window or tab >>Bioanalytical separation using capillary electrophoresis: Applications with microbubbles and proteins
2017 (English)Licentiate thesis, comprehensive summary (Other academic)
Abstract [en]

In this thesis the possibilities of using capillary electrophoresis as a separation technique for analysis of proteins and microbubbles is presented.

A complete analytical process consists of five necessary steps of which one is the actual analysis step. For this step a suitable analytical technique is needed. Capillary electrophoresis (CE) is one of the common analytical separation techniques used for analysis of a diversity of analytes, and can be both used in routine analysis and for research purposes. The reason for using CE, compared to other liquid-based separation techniques, is mainly short analysis time, high resolution, and negligible sample volumes and solvent waste. Depending on the characteristics of the analytes, and the sample matrix, different modes of CE can be used, where capillary zone electrophoresis (CZE) is the most employed one. The basic principle of CZE is separation of the analytes due to differences in total mobility, which is dependent on the charge and size of the analytes, and the electroosmotic flow (EOF). The EOF can be controlled by several parameters e.g. choice of background electrolyte (BGE), and the optimization of the parameters has been discussed throughout the thesis.

To improve the properties of the BGE, an ethylammonium nitrate (EAN) water solution was used as BGE for CE analysis in Paper I. The precision of the EOF with this method was determined by adjusting the pH of the BGE, the concentration of EAN in the BGE, and the electric field. Model proteins were thereafter analysed using the optimal parameters yielding a precision sufficient for routine control.

One example of the applications of CE is separation of novel contrast agents, which consist of polyvinyl alcohol microbubbles (PVA-MBs). In Paper II, a method for analysis of PVA-MBs in biological samples using CE with UV-detection was developed. It was also established that intact PVA-MBs could be distinguished from ultrasound degraded PVA-MBs in the same set-up.

Place, publisher, year, edition, pages
Stockholm: KTH Royal Institute of Technology, 2017
Series
TRITA-CHE-Report, ISSN 1654-1081 ; 2017:32
Keywords
Background electrolyte, capillary electrophoresis, contrast agents, electroosmotic flow, ethylammonium nitrate, ionic liquids, microbubbles, particles, polyvinyl alcohol microbubbles, and separation
National Category
Analytical Chemistry
Research subject
Chemistry
Identifiers
urn:nbn:se:kth:diva-215194 (URN)978-91-7729-560-0 (ISBN)
Presentation
2017-11-23, K52, Teknikringen 28, plan 5, Stockholm, 10:00 (English)
Supervisors
Note

QC 20171012

Available from: 2017-10-12 Created: 2017-10-11 Last updated: 2017-10-12Bibliographically approved
2. Bioanalysis using capillary electrophoresis and mass spectrometry: Applied on proteins, protein nanofibrils and polyvinyl alcohol microbubbles
Open this publication in new window or tab >>Bioanalysis using capillary electrophoresis and mass spectrometry: Applied on proteins, protein nanofibrils and polyvinyl alcohol microbubbles
2019 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

The sequencing of the genome of various species, including the human species, have led to increased understanding about how a protein structure is generated, and how specific structures are related to the proteins’ functionality. In paper I and II of this thesis, the folding of proteins in vitro to form hierarchical nanostructures, which in vivo often have a pathological effect, have been studied. Protein isolates from soybean and potato, that are byproducts from oil and starch production, respectively, were used as a starting material for protein nanofibril (PNF) formation, and mass spectrometry was used to identify the building blocks that are included in the formed PNF. The five peptides identified in soybean PNF and the six peptides identified in potato PNF originated from the major seed storage proteins for the respective crop.

The use of ionic liquids has increased for improvement of the performance of different separation techniques due to their adjustable properties, and good solvating ability. In paper III, an ionic liquid and water mixture was used as background electrolyte in capillary electrophoresis for protein separation. The system showed high reproducibility at basic conditions, and could potentially be used for routine control analysis.

Many diseases and injuries require clinical diagnosis techniques e.g. ultrasound imaging, to be detected, and for the physician to be able to decide the correct therapy. To increase the resolution of such imaging techniques, contrast agents can be used. In paper IV-VI, a newly developed contrast agent consisting of air-filled microbubbles stabilized with a shell of polyvinyl alcohol (PVA-MBs) was studied. Development of a capillary electrophoretic method for analysis of the PVA-MBs with the intentions to be used for clinical diagnosis is performed, where different detectors such as a UV detector, a UV area imaging detector and an in-house constructed microscope are used to increase the sensitivity of detection for the PVA-MBs. The developed method could be used for quantification of the contrast agent, since individual PVA-MBs were visible using the imaging detectors. Findings regarding the mobility of the PVA-MBs in human blood plasma and in water implies that a protein corona was formed around the MBs.

Abstract [sv]

Sekvensering av genomet för många arter, inklusive den mänskliga, har lett till en ökad förståelse för hur proteiners strukturer är genererade, och hur specifika proteinstrukturer är relaterade till proteinernas funktioner. I artikel I och II i denna avhandling studeras proteinveckning till hierarkiska nanostrukturer in vitro, vilka vanligtvis har en patologisk effekt in vivo. Proteinisolat från sojabönor och potatis, d.v.s. biprodukter från respektive olje- och stärkelseproduktion, användes som startmaterial för tillverkningen av protein-nanofibriller (PNF). Masspektrometri användes för att identifiera byggstenarna i dessa nanofibriller. De fem identifierade peptiderna från sojabönans PNF och de sex peptiderna från potatis PNF har sitt ursprung i de större lagringsproteinerna från respektive gröda.

Användningen av jonvätskor har ökat i avseende att förbättra prestandan för olika separationstekniker på grund av jonvätskors justerbara egenskaper och effektivitet vid användning som lösningsmedel. I artikel III används en utspädd jonvätska som bakgrundselektrolyt i kapillärelektrofores för separation av proteiner. Systemet uppvisade så god reproducerbarhet vid alkaliska förhållanden att det potentiellt skulle kunna användas vid rutinanalyser.

Många sjukdomar och skador kräver kliniska diagnostiska tekniker såsom ultraljudsavbildning (sonografi), för att rätt terapi ska kunna tillämpas. För att förbättra upplösningen för ultraljudsavbildning kan kontrastmedel användas. I artikel IV-VI studerades nyligen utvecklade mikrobubblor av luft med ett hårt och stabiliserande skal av polyvinylalkohol (PVA-MBs) som kontrastmedel. En kapillärelektroforetisk metod har tagits fram för analys av dessa PVA-MBs som i avsikt att använda dessa i klinisk diagnostik. Tre olika detektorer har utvärderats för att uppnå känslig detektion av detta kontrastmedel: en UV-detektor, en ytavbildande UV-detektor och en egen-konstruerad mikroskopdetektor. Den utvecklade kapillärelektroforetiska metoden kan användas för kvantifiering av PVA-MBs då känsligheten för de avbildande detektorerna var så hög att individuella MB var synliga. Den elektroforetiska mobiliteten för PVA-MB varierade när PVA-MBs suspenderats i vatten med suspension i humant blodplasma, vilket indikerar att en proteinkorona kring PVA-MB bildades.

Place, publisher, year, edition, pages
Stockholm: KTH Royal Institute of Technology, 2019. p. 43
Series
TRITA-CBH-FOU ; 2019:49
Keywords
: amyloid, capillary electrophoresis, contrast agents, detection, human blood plasma, ionic liquids, mass spectrometry, microbubbles, polyvinyl alcohol microbubbles, protein nanofibrils, proteins
National Category
Analytical Chemistry
Research subject
Chemistry
Identifiers
urn:nbn:se:kth:diva-257962 (URN)978-91-7873-311-8 (ISBN)
Public defence
2019-10-24, sal F3, Lindstedtsvägen 26, Stockholm, 10:00 (English)
Opponent
Supervisors
Note

QC 2019-09-20

Available from: 2019-09-20 Created: 2019-09-18 Last updated: 2019-09-20Bibliographically approved

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