Universiẗat, Diss., 2003--Bonn.

The ongoing effort to improve the sensitivity and information content of NMR spectroscopy and MRI has important implications in scientific research and medical diagnostics. In this dissertation, a variety of approaches have been investigated and expanded on in an effort to contribute to this field. First, cryptophanes are cage-shaped molecules that have previously been used to encapsulate molecules of interest for a number of potential applications--including gas sensing and biosensing. In one set of studies, encapsulation of molecular hydrogen gas (H2) has shown different behavior compared to other small organic molecules in C111 (up until now, the smallest cryptophane). The transient, non-covalent binding was studied by variable-temperature NMR at different fields up to 950 MHz. A mathematical model that considers multiple-H2 binding was developed to better understand the physics and binding process, with predictions compared to experimental data (and rationalized in light of quantum chemical calculations on possible H2@C111 complexes). To our knowledge, C111 is the only system to reversibly trap multiple H2 gas molecules non-covalently under mild conditions. In a second series of studies, the interaction of laser-polarized xenon and a water-soluble cryptophane was studied. Despite the low concentration of xenon in aqueous solution, it was possible to achieve polarization transfer from xenon to cryptophane spins via the SPINOE (spin-polarization induced nuclear Overhauser effect). The SPINOE enhancements, along with the 129Xe NMR spectra, provide information about the interaction of the Xe-cryptophane complex (variants of which are now used in so-called xenon biosensors). This was our first in-house successful application of hyperpolarized xenon as a signal source for the spins of other molecules, leading the way to a number of ongoing studies. Although the absolute NMR enhancements obtained via the SPINOE were small, much larger enhancements were studied in a technique that uses para-hydrogen (pH2)--a spin isomer of normal molecular hydrogen)--as the source of spin order. As with the xenon experiments (and the H2 binding experiments), pH2 must be delivered as a gas to a sealed sample prior to performing the NMR experiments. Parahydrogen-induced polarization (PHIP) is an emerging field in enhancing the sensitivity in NMR experiments and may play an important role in MRI studies. Within this field a very recent phenomena of signal amplification by reversible exchange (SABRE) was investigated. The reproducibility of this recent discovery has been examined and new conclusions about the mechanism of this technique are delineated. NMR signal enhancements of nearly ~400-fold are reported. Moreover, a new water soluble NHC-Iridium catalyst was synthesized and investigated in SABRE related studies. We also report the first studies of SABRE-enhancement in biologically tolerable solvents--opening a door to the development of SABRE-hyperpolarized metabolic contrast agents for subsecond molecular imaging in the body. Although much of the above work was motivated by the desire to improve NMR/MRI sensitivity enhancement, other efforts concerned the other side of the equation--improving NMR/MRI information content. The next section concerns our efforts to investigate use of Variable-Angle (VA) NMR to study composite liquid crystal (LC) media comprised of stretched polyacrylamide gels (SAG) and embedded bacteriophage Pf1 particles. This in situ combination exploited the apparent interference between the different solute-aligning properties of the two LC components--yielding composite media with alignment properties that can differ in a tunable manner from those obtained with each medium alone. Characterization of alignment of both large and small molecules provides more insight into the nature of solute alignment that those composite phases introduce--with the goal of developing this approach as a new technique for studying molecular structure and dynamics via the dipolar and quadrupolar couplings that are restored in liquid-crystalline media. Finally the use of SPIONS--superparamagnetic iron oxide nanoparticles--as contrast agents is a relatively new approach to enhance information content in MRI studies; this is particularly true for SPIONs that have been surface-functionalized to achieve an environment-sensitive MR response. Novel surface-functionalized SPIONs were investigated by examining their effect on nuclear spin relaxation in aqueous environments simulating bodily tissues. More specifically, the pH and ionic strength dependent properties of selected dendron-functionalized and polymer-functionalized SPIONs have been examined. Of particular interest to this dissertation is how environment-mediated transient clustering of the SPIONs gives rise to changes in so-called transverse (homogeneous) spin relaxation rates as measured by following the decay of MR signals detected after the application of a series of radio-frequency (RF) pulses. In order to better understand these effects in the context of the SPIONs' behavior, a mathematical model is under development whose predictions are compared with experimental data. Aspects of the model are also compared to transmission electron micrography (TEM) and dynamic light scattering (DLS).

Universiẗat, Diss., 2000--Bonn.

Thesis (Ph. D.)--Miami University, Dept. of Chemistry and Biochemistry, 2007. / Title from second page of PDF document. Includes bibliographical references (p. 87-96).

No description available.

Die Enzyme der Sulfotransferase-Gensuperfamilie (SULT) konjugieren nukleophile Gruppen von kleinen endogenen Verbindungen und Fremdstoffen mit der negativ geladenen Sulfo-Gruppe. Dadurch wird die Polarität dieser Verbindungen erhöht, ihre passive Permeation von Zellmembranen verhindert und somit ihre Ausscheidung erleichtert. Jedoch stellt die Sulfo-Gruppe in bestimmten chemischen Verbindungen eine gute Abgangsgruppen dar. Aus der Spaltung resultierende Carbenium- oder Nitreniumionen können mit DNA oder anderen zellulären Nukleophilen reagieren. In Testsystemen für Mutagenität wurden zahlreiche Verbindungen, darunter Nahrungsinhaltsstoffe und Umweltkontaminanten, durch SULT zu Mutagenen aktiviert. Dabei zeigten sich zum einen eine ausgeprägte Substratspezifität selbst orthologer SULT-Formen unterschiedlicher Spezies und zum anderen Interspezies-Unterschiede in der SULT-Gewebeverteilung. Daher könnten sich die Zielgewebe einer SULT-induzierten Krebsentstehung bei Mensch und Nager unterscheiden. Um die Beteiligung von humanen SULT an der Bioaktivierung von Fremdstoffen im Tiermodell untersuchen zu können, wurden transgene Mauslinien für den Cluster der humanen SULT1A1- und -1A2-Gene sowie für die humane SULT1B1 generiert. Zur Herstellung der transgenen Linien wurden große genomische Konstrukte verwendet, die die SULT-Gene sowie – zum Erreichen einer der Humansituation entsprechenden Gewebeverteilung der Proteinexpression – deren potentielle regulatorische Sequenzen enthielten. Es wurden je drei transgene Linien für hSULT1A1/hSULT1A2 und drei transgene Linien für hSULT1B1 etabliert. Die Expression der humanen Proteine konnte in allen Linien gezeigt werden und fünf der sechs Linien konnten zur Homozygotie bezüglich der Transgene gezüchtet werden. In der molekularbiologischen Charakterisierung der transgenen Linien wurde der chromosomale Integrationsort der Konstrukte bestimmt und die Kopienzahl pro Genom untersucht. Mit Ausnahme einer hSULT1A1/hSULT1A2-transgenen Linie, bei der Kopien des Konstrukts in zwei unterschiedliche Chromosomen integriert vorliegen, wiesen alle Linien nur einen Transgen-Integrationsort auf. Die Untersuchung der Transgen-Kopienzahl ergab, dass die Mauslinien zwischen einer und etwa 20 Kopien des Transgen-Konstrukts pro Genom trugen. In der proteinbiochemischen Charakterisierung wurde gezeigt, dass die transgenen Linien die humanen Proteine mit einer weitgehend der des Menschen entsprechenden Gewebeverteilung exprimieren. Die Intensität der im Immunblot nachgewiesenen Expression korrelierte mit der Kopienzahl der Transgene. Die zelluläre und subzelluläre Verteilung der Transgen-Expression wurden bei einer der hSULT1A1/hSULT1A2-transgenen Linien in Leber, Niere, Lunge, Pankreas, Dünndarm und Kolon und bei einer der hSULT1B1-transgenen Linien im Kolon untersucht. Sie stimmte ebenfalls mit der Verteilung der entsprechenden SULT-Formen im Menschen überein. Da sich die erzeugten transgenen Linien aufgrund ihrer mit dem Menschen vergleichbaren Gewebeverteilung der SULT-Expression als Modellsystem zur Untersuchung der menschlichen SULT-vermittelten metabolischen Aktivierung eigneten, wurde eine der hSULT1A1/hSULT1A2-transgenen Linien für zwei erste toxikologische Untersuchungen eingesetzt. Den Mäusen wurden chemische Verbindungen verabreicht, für die in in-vitro-Versuchen eine hSULT1A1/hSULT1A2-vermittelte Bioaktivierung zu Mutagenen gezeigt worden war. In beiden Untersuchungen wurde die Gewebeverteilung der entstandenen DNA-Addukte als Endpunkt einer gewebespezifischen genotoxischen Wirkung ermittelt. In der ersten Untersuchung wurden 90 mg/kg Körpergewicht 2-Amino-1-methyl-6-phenylimidazo[4,5-b]pyridin – ein in gebratenem Fleisch gebildetes heterozyklisches aromatisches Amin – transgenen sowie Wildtyp-Mäusen oral verabreicht. Acht Stunden nach Applikation wiesen die transgenen Mäuse signifikant höhere Adduktniveaus als die Wildtyp-Mäuse in Leber, Lunge, Niere, Milz und Kolon auf. In der Leber der transgen Mäuse war das Adduktniveau 17fach höher als in der Leber der Wildtyp-Mäuse. Die Leber war bei den transgenen Tieren das Organ mit dem höchsten, bei den Wildtyp-Tieren hingegen mit dem niedrigsten DNA-Adduktniveau. In der zweiten Untersuchung (Pilotstudie mit geringer Tierzahl) wurde transgenen und Wildtyp-Mäusen 19 mg/kg Körpergewicht des polyzyklischen aromatischen Kohlenwasserstoffs 1-Hydroxymethylpyren – ein Metabolit der Nahrungs- und Umweltkontaminante 1-Methylpyren – intraperitoneal verabreicht. Nach 30 Minuten wurden, verglichen mit den Wildtyp-Mäusen, bis zu 25fach erhöhte Adduktniveaus bei den transgenen Mäusen in Leber, Niere, Lunge und Jejunum nachgewiesen. Somit konnte anhand einer in dieser Arbeit generierten transgenen Mauslinie erstmals gezeigt werden, dass die Expression der humanen SULT1A1/hSULT1A2 tatsächlich sowohl auf die Stärke als auch die Zielgewebe der DNA-Adduktbildung in vivo eine Auswirkung hat. / The enzymes of the sulfotransferase gene superfamily (SULT) conjugate nucleophilic groups of small endogenous compounds and xenobiotics with the negatively charged sulfo group. Thus, the polarity of the compounds is increased, their passive permeation of cell membranes is hindered and their excretion facilitated. The sulfate groups, however, form a good leaving group in certain chemical linkages due to their electron-withdrawing characteristics. Carbenium or nitrenium ions resulting from a spontaneous cleavage may react with DNA and other cellular nucleophiles. In test systems for mutagenicity, a large amount of compounds including ingredients of nutrition and environmental contaminants were activated to mutagens by SULT. A pronounced substrate specificity even of orthologous SULT forms of different species was evidenced. Also, the tissue distribution of SULT exhibited pronounced interspecies differences. The target tissues of a SULT induced carcinogenesis might thus be different in humans and rodents. To investigate the involvement of human SULT in the bioactivation of xenobiotics in an animal model, transgenic mouse lines for the human SULT1A1- and -1A2 gene cluster as well as for human SULT1B1 were generated. For the construction of the transgenic lines, large genomic constructs were used, containing the SULT genes plus their potential regulatory sequences to cause a tissue distribution of protein expression corresponding to the situation in humans. Three transgenic lines for hSULT1A1/hSULT1A2 and three transgenic lines for hSULT1B1 were established. The expression of the human proteins could be shown for all lines and except for one line, all could be bred to transgene homozygosity. By molecular biological characterization of the transgenic lines, the chromosomal integration locus of the constructs was identified and the copy number per genome was investigated. With the exception of one hSULT1A1/hSULT1A2 transgenic line, where the construct had integrated into two different chromosomes, all lines exhibited just one transgene integration locus. By investigating the transgene copy number it was deduced that the mouse lines carry between one and 20 copies of the transgene construct per genome. The protein biochemical characterization showed that the transgenic mouse lines express the human proteins with a tissue distribution largely similar to the distribution in humans. The intensity of the proteins detected by immunoblotting correlated with the copy number of the transgenes. The cellular and subcellular distribution of the transgene expression was investigated for one of the hSULT1A1/1A2 transgenic lines in liver, kidney, lung, pancreas, small intestine and colon and for one of the hSULT1B1 transgenic lines in colon. It also accorded with the distribution of the respective SULT in humans. Owing to the similarity of transgene expression to the corresponding human tissue distribution, the transgenic lines were considered suitable as model systems for the investigation of the human SULT-mediated metabolic activation. One of the hSULT1A1/hSULT1A2 transgenic lines was used in two first toxicological investigations with chemical compounds for which in vitro experiments had demonstrated a hSULT1A1/hSULT1A2 mediated bioactivation. In both investigations, the tissue distribution of the resulting DNA adducts was determined as an end point for a tissue-specific genotoxic effect. For the first investigation, 90 mg/kg bodyweight of 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine – a heterocyclic amine formed in cooked meat – were orally administered to transgenic and wild type mice. Eight hours after application, the transgenic mice exhibited significantly higher adduct levels than the wild type controls in liver, lung, kidney, spleen and colon. The adduct level in the liver of the transgenic mice exceeded that in the wild type liver by a factor of 17. Furthermore, the liver was the organ with the highest adduct level in the transgenic mice and with the lowest adduct level in the wild type mice. For the second investigation (a pilot study with few animals), 19 mg/kg bodyweight of the polycyclic aromatic hydrocarbon 1-hydroxymethylpyrene – a metabolite of the nutritional and environmental contaminant 1-methylpyrene – were administered intraperitoneally to transgenic and wild type mice. After 30 minutes, up to 25 fold higher adduct levels compared to the wild type were detected in liver, kidney, lung and jejunum of the transgenic mice. Thus, by means of one of the transgenic mouse line generated in this thesis, it could be shown for the first time that the expression of human SULT1A1/SULT1A2 has in fact an impact on the strength as well as on the target tissue of DNA-adduct generation in vivo.

Eine besondere Rolle im Fremdstoffmetabolismus hat die SULT1A1 beim Menschen aufgrund der hohen Expression und breiten Gewebeverteilung. Während die humane SULT1A1 in sehr vielen Geweben exprimiert wird, wurde die murine SULT1A1 vor allem in der Leber, Lunge und Colon gefunden. Neben der Gewebeverteilung spielt auch der Polymorphismus im humanen SULT1A1-Gen eine bedeutende Rolle. Der häufigste Polymorphismus in diesem Gen führt zu einer Aminosäuresubstitution von Arginin zu Histidin an Position 213. Die Genvariante mit Histidin (auch als SULT1A1*2 bezeichnet) codiert für ein Protein mit einer geringen Enzymaktivität und einer reduzierten Enzymmenge in Thrombocyten. Über den Einfluss dieser allelischen Varianten in anderen Geweben ist bislang wenig bekannt. In vorausgegangenen epidemiologischen Studien wurden mögliche Korrelationen zwischen den Genvarianten und der Krebsentstehung in verschiedenen Geweben untersucht. Diese Daten liefern jedoch widersprüchliche Ergebnisse zum Krebsrisiko. Aufgrund der strittigen epidemiologischen Daten sollten Tiermodelle generiert werden, um die häufigsten SULT1A1-Allele hinsichtlich der Empfindlichkeit gegenüber Nahrungs- und Umweltkanzerogenen zu untersuchen. Zur Erzeugung transgener (tg) Mauslinien wurde mittels Mikroinjektion der codierenden Genbereich und große flankierende Humansequenzen stromaufwärts und stromabwärts in das Mausgenom integriert. Es wurden mehrere Mauslinien hergestellt. Zwei davon, die Mauslinie 31 mit dem SULT1A1*1-Allel und die Mauslinie 28 mit dem SULT1A1*2-Allel, wurden eingehend analysiert. In beiden Linien wurde eine identische Kopienzahl des Transgens ermittelt. Proteinbiochemische Charakterisierungen zeigten eine weitgehend dem Menschen entsprechende Gewebeverteilung und zelluläre und subzelluläre Lokalisation der humanen SULT1A1 in der Linie (Li) 28. In Li 31 wurden Unterschiede zu Li 28 sowohl in der Gewebeverteilung als auch in der zellulären Lokalisation des exprimierten humanen Proteins ermittelt. Dabei war die Expression auf Proteinebene in der SULT1A1*2-tg Linie generell stärker als in der SULT1A1*1-Linie. Dieses Ergebnis war überraschend, denn in humanen Thrombocyten führt das SULT1A1*1-Allel zu einem höheren Gehalt an SULT1A1-Protein als das SULT1A1*2-Allel. Zur Analyse der unterschiedlichen Proteinexpressionen in den tg Mauslinien wurde die cDNA und der 5´-flankierende Bereich des SULT1A1-Gens sequenziert. In beiden tg Linien entsprach die Sequenz der cDNA der Referenzsequenz aus der Gendatenbank (Pubmed). In der 5´-flankierenden Region wurden bekannte Polymorphismen analysiert und unterschiedliche Haplotypen in den tg Linien an den Positionen -624 und -396 ermittelt. Dabei wurde in der Li 31 der Haplotyp detektiert, der in der Literatur mit einer höheren SULT1A1-Enzymaktivität beschrieben wird. Der mögliche Zusammenhang zwischen Transkriptionsrate und Proteinexpression wurde in RNA-Expressionsanalysen im codierenden und 5´-nicht codierenden Bereich (mit den alternativen Exons 1B und 1A) untersucht. Im codierenden Bereich und im Exon 1B konnte in den untersuchten Organen eine höhere RNA-Expression in der Li 28 im Vergleich zur Li 31 ermittelt werden. Außer in der Lunge wurde für Exon 1B eine identische RNA-Expression detektiert. RNA, die Exon 1A enthielt, wurde in allen untersuchten Organen der Li 28, aber nur in der Lunge bei der Li 31 gefunden. In beiden tg Linien konnten mit den Exon 1A-Primern jedoch auch größere PCR-Produkte ermittelt werden. Dieser Unterschied im Exon 1A und mögliche Spleißvarianten könnten damit für die unterschiedliche Proteinexpression des humanen SULT1A1-Proteins in den beiden tg Mauslinien sein. Die in dieser Arbeit generierten und charakterisierten tg Mausmodelle wurden in einer toxikologischen Studie eingesetzt. Es wurde das heterozyklische aromatische Amin 2-Amino-1-methyl-6-phenylimidazo-[4,5-b]pyridin (PhIP) verwendet. PhIP wird beim Erhitzen und Braten von Fleisch und Fisch gebildet und könnte mit der erhöhten Krebsentstehung im Colon in der westlichen Welt im Zusammenhang stehen. Mittels 32P-Postlabelling sollte der Einfluss der zusätzlichen Expression der humanen SULT-Proteine auf die PhIP-DNA-Adduktbildung analysiert werden. Dabei wurden mehr DNA-Addukte in den tg Tieren als in den Wildtyp-Mäusen ermittelt. Die Konzentration der gebildeten DNA-Addukte korrelierte mit der Expressionsstärke des humanen SULT1A1-Proteins in den tg Mäusen. An den in dieser Arbeit generierten tg Mauslinien mit den häufigsten allelischen Varianten des SULT1A1-Gens konnten Unterschiede auf RNA- und Protein-Ebene ermittelt werden. Zudem konnte gezeigt werden, dass die Expression der humanen SULT1A1 eine Auswirkung sowohl auf die Stärke als auch das Zielgewebe der DNA-Adduktbildung in vivo hat. / In humans, SULT1A1 and its polymorphic variants play an important role in xenobiotic metabolism and display a broad tissue distribution and high expression level. This enzyme is expressed in almost every human organ whereas in mice SULT1A1 can only be detected in liver, lung and colon. The most common polymorphism of this gene leads to an amino acid substitution from arginine to histidine at the position 213. In platelets, the allele encoding histidine (also designated as SULT1A1*2) is associated with both low activity and low thermal stability of the SULT protein. However, so far only little is known about the significance of these allelic variants in the other tissues with hSULT1A1 expression. Previous epidemiological studies have made attempts to correlate SULT1A1 allelic variants and cancer development, their data, however, have been contradictory for an appropriate cancer risk assessment. In this thesis, we addressed the effect of the hSULT1A1 genetic variability on the susceptibility to nutritional and environmental carcinogens using transgenic (tg) mouse models. We generated tg mice carrying the most common allelic variants of the human SULT1A1 gene. The coding region and large flanking human sequences upstream and downstream of the hSULT1A1 gene were integrated randomly into the mouse genome by microinjection. Several tg mouse lines were generated. Two of them, line (li) 31 with the SULT1A1*1 allele and li 28 with the SULT1A1*2 allele, were analysed in detail. At first, an identical transgene copy number was detected in both lines. Furthermore, biochemical characterization of li 28 showed that the tissue distribution, the cellular and subcellular localisation of the protein were very similar to those in humans. In contrast, li 31 exhibited differences in tissue distribution and cellular localisation of the human protein compared to li 28. The protein expression level in the tg line with SULT1A1*2 (li 28) was generally higher than in SULT1A1*1 (li 31) mice. These results were surprising since the SULT1A1*1 allele in human platelets usually leads to a higher amount of SULT1A1 protein compared to the SULT1A1*2 allele. To investigate these differences, we sequenced the cDNA and 5´-flanking region of the SULT1A1 gene. In both tg mouse lines, the cDNA sequence was identical to the reference sequence from the gene databank (Pubmed). We subsequently analysed the common polymorphisms of the 5´-flanking region, and determined different haplotypes at position -624 and -396 in the tg mouse lines. According to the literature, the haplotype associated with a higher SULT1A1 enzyme activity, we detected in li 31. We analyzed the possible correlation between gene transcription and protein expression by measuring RNA expression levels of the coding and the non-coding region (with alternative exons 1B and 1A). We detected a higher RNA expression level of the coding region and exon 1B in li 28 compared to li 31, whereas RNA for exon 1A was only found in li 28 in all investigated tissues, but only in lung in li 31. Furthermore we detected with exon 1A-primers larger RNA in both lines. These differences in exon 1A expression accompanied by potential splicing variants could be responsible for the different expression and activity of the human SULT1A1 protein in both tg mouse lines. In order to validate our generated and characterized tg mouse models as toxicological in vivo models, we used them for the evaluation of the heterocyclic aromatic amine 2-amino-1-methyl-6-phenylimidazo-[4,5-b]pyridine (PhIP). PhIP is typically generated during heating and roasting of meat and fish and is suggested to be associated with an increased colon cancer incidence in the western world. We measured the impact of the additionally expressed human SULT proteins on the PhIP-DNA adduct level by 32P-postlabelling. We detected significantly higher DNA adduct levels in tg compared to wildtype mice, which correlated positively with the expression pattern of the human SULT1A1 protein in the tg mice. In conclusion, in this thesis, we have successfully generated and validated the transgenic mouse lines carrying the most common allelic variants of the human SULT1A1 gene. Interestingly, these lines exhibited differences in both the SULT1A1 RNA and protein levels. Using these transgenic mouse models as in vivo toxicological tools we have shown that the expression of human SULT1A1 in mice has a decisive impact on the strength and the target tissue of DNA adducts.

Universiẗat, Diss., 2005--Bonn.

No description available.

Abstract The subject of this thesis is flow imaging by methods based on the nuclear magnetic resonance (NMR) phenomenon. The thesis consists of three related topics: In the first one the feasibility of measuring velocity maps and distributions inside a microfluidic device by pulsed field gradient (PFG) NMR has been demonstrated. The second topic was to investigate microfluidic gas flow using a combination of a special detection technique and a powerful signal enhancement method. The third topic is related to the unambiguous determination of velocities under challenging experimental conditions and introduces a new, improved velocity imaging sequence. In the first part, well established imaging methods have been used to study water flow inside a micromixer. A surface coil matching the region of interest of the mixer was home built and used in the measurements in order to gain a better signal-to-noise ratio. Velocities inside the mixer have been measured by phase-encoding velocity, with unprecedented spatial resolution. Two dimensional NMR imaging and velocity maps revealed clogging and different manufacturing qualities of the mixers. In addition to the velocity maps, which display an average velocity for spins within one pixel, complete velocity distributions (so called average propagators) were measured. It was found that in the absence of spatial resolution in the third dimension, the propagator data can provide valuable insight to the flow system by revealing overlapping flow passages. The next topic was gas flow inside a microfluidic device. It was investigated by time-of-flight flow imaging. The measurement of the weak gas signal was enabled by the use of two signal enhancement techniques: remote detection NMR and parahydrogen induced polarization (PHIP). The results demonstrate that a very significant signal enhancement can be achieved by this technique. In the future it may enable the investigation of interesting chemical reactions inside microreactors. The third and last topic of the thesis deals with measuring flow by the so called multiecho sequences. When multiecho sequences are used in combination with phase encoding velocity, an error may be introduced: the multiecho sequence may produce a cumulative error to the phase of the magnetization, if it is sensitive to RF pulse imperfections. The problem has been elaborately explained and various solutions discussed, among the newly proposed one. Experimental results demonstrate the performance of the new velocity imaging sequence and show that the new sequence enables the unambiguous determination of velocities even in challenging experimental conditions resulting from inhomogeneous radio frequency fields of the measurement coils.