Workshop limited to 4 participants (first come, first served)
Contact person:
Silvia Lope-Piedrafita, PhD ()
This course combines a comprehensive series of lectures on the technology of Magnetic resonance spectroscopy and imaging (MRS/MRI) with hands-on laboratory sessions to provide practical demonstrations of key concepts and procedures for preclinical studies.
Whether you are considering MRI as a research tool in your lab or just would like to learn more about MRI, this workshop addresses practical aspects of experimental MRI with laboratory animals and provide valuable hands-on experience on a 7 Tesla Bruker BioSpec spectrometer.
The world’s top 2% of most impact scientists across all fields have been updated in a new list issued in October 2023 by Stanford University (1). Dr. Teodor Parella, head of the SeRMN-UAB, appears in this Stanford list that includes 3230 spanish researchers from all disciplines, around 300 spanish chemists and 81 researchers from the Universitat Autònoma de Barcelona. Dr. Parella is found at the top world-wide 1% scientists list, ranking #80088 in the world, #980 in Spain and #21 at the UAB.
Dr. Parella has also been ranked #6 at UAB, #268 in Spain and #7687 in the world in the field of Chemistry, according toResearch.com, a leading academic platform for researchers, that has just released the 2023 Edition of the Ranking of Best Scientists. The ranking is based on D-index (Discipline H-index) metric, which only includes papers and citation values for an examined discipline. The ranking includes only leading scientists with D-index of at least 40 for academic publications made in the area of Chemistry.
Last 26th July 2023 I successfully defended my PhD Thesis entitled: “PhD Thesis: Implementation of high-resolution MRSI methods in a pre-clinical scanner, and optimization for brain longitudinal studies of therapy response in mice glioma model.”
Abstract
Magnetic resonance spectroscopy and magnetic resonance spectroscopic imaging (MRS/MRSI) are non-invasive diagnostic techniques that use a strong magnetic field and radio waves to examine the chemical composition of living tissue. Working on the same principles as Magnetic Resonance Imaging (MRI), instead of producing images, MRS generates a spectrum of signals that can be used to identify the type and amount of molecules present in a tissue. The utility of MRS and MRSI has already been established in many studies, providing useful information about the chemical makeup of different regions of the brain, and allowing diagnosis of conditions such as Alzheimer’s disease, multiple sclerosis, and brain Glioblastoma (GB) tumors.
Preclinical glioblastoma studies looking forward to improving therapeutic outcomes are necessary since clinical GB has no current cure. These studies can greatly benefit from improved spatial resolution and homogeneity of the acquired MRSI grids. Hence, we can work towards improved acquisition schemes enhancing the quality of acquired data using MRS and MRSI. There exists a methodological consensus among spectroscopy experts where the Localized Adiabatic Spin Echo Refocused (semiLASER) data acquisition strategy has been ranked as the most likely localization technique to improve (pre) clinical MRS. SemiLASER uses adiabatic pulses to selectively excite and refocus the signal from a localized volume of interest in the brain. This results in a higher signal-to-noise ratio (SNR) and better spatial resolution compared to conventional data acquisition sequences.
Partial volume effects can occur in MRSI when the voxel (a 3D volume of interest) being measured contains a mixture of different neighbouring tissue types or compartments, such as grey and white matter or cerebrospinal fluid. This can lead to inaccurate quantification of metabolites, as the signal from one tissue can mix with the signal from another and affect overall pattern recorded. SemiLASER is designed to minimize partial volume effects by using adiabatic pulses to selectively excite and refocus the signal from a small region of interest within the voxel. This allows for more accurate quantification of metabolites within the region of interest, while reducing the contamination of the signal by other tissue types. In addition, semiLASER also employs an outer-volume suppression (OVS) technique to further reduce contamination from outside the region of interest. This involves using additional adiabatic pulses to selectively saturate the signal from outside the volume of interest, so that it does not contribute to the MRSI signal. Overall, the combination of selective excitation and OVS in semiLASER can help improve the accuracy of MRSI measurements and reduce partial volume effects.
Although, the clinical utility of semiLASER has been acknowledged, the pre-clinical use and implementation of semiLASER still remains a less explored area. Our group has a long record of using MRSI in therapy response monitoring of a murine model glioblastoma (the GL261 cell line) using a commercially available MRSI acquisition sequence. In our efforts towards bridging the barriers between pre-clinical and clinical research, we have implemented the clinically verified semiLASER sequence on a pre-clinical 7T Bruker Biospec USR scanner running the ParaVision 5.1 software package, which provides a graphical user interface for sequence programming and data acquisition. The single and multi-voxel semiLASER sequences were implemented with the idea that the developments generated during this PhD project will be replicable by other interested users.
The implemented SV-semiLASER and MRSI-semiLASER sequences for preclinical acquisitions were optimised to perform high resolution MRSI of living mouse brain. For this, sequences were duly verified and tested first in phantoms and later in-vivo, in wild-type (wt) and tumor bearing (GL261) mice. To do so, the Bruker pulse sequence implementation was first studied in detail to become familiar with the Bruker programming environment and a test sequence PRESS_Slice to localize the slice dimension was developed by modifying the Bruker stock PRESS sequence for single voxel localization. After careful evaluation of test sequence results, the semiLASER single and multi-voxel sequences were also implemented using a similar strategy.
The implemented SV-semiLASER sequence provided a ca. 1.4-fold improvement in SNR in phantoms and ca. 1.3-fold improvement in SNR for in-vivo subjects, in comparison to the stock Bruker PRESS (single volume acquisition) sequence. The MRSI-semiLASER sequence resulted in a ca. 1.3-fold improvement of SNR in phantoms and in-vivo subjects compared to the stock Bruker CSI-PRESS sequence. Combined with phase encoding strategies and volume reduction methods, higher spatial resolution and SNR was achieved for the implemented MRSI-semiLASER. The quantification analysis of the results was done using MATLAB based post-processing tools specially designed to process Bruker datasets and solutions for a faster post processing pipeline were proposed. The single voxel MRSI-semiLASER sequences were further simulated using NMRSCOPE-B virtual simulator, a jMRUI plug-in which positively correlated with the experimental results. Preliminary nosological images obtained using MRSI-semiLASER datasets and the SpectraClassifier tool previously developed in our group, and trained with GL261 tumors using already available CSI-PRESS data, suggested those classifiers could be robust enough to recognize the tumor region acquired with the semi-LASER sequence. Still, classifiers may require retraining for the evaluation of response to therapy, which is an ongoing project within the group.
The thesis dissertation can be downloaded in PDF format using the link below or from the official TXD and Teseo repositories (currently in progress):
I would like to thank the financial support by the European Comission Marie Curie Initial Training Networks (ITN, call H2020-MSCA-ITN-2018, grant 813120 to project INSPiRE-MED); by the Ministry of Science and Insdustry (MCIN/AEI/10.13039/501100011033) (APC); and by Centro de Investigación Biomédica en Red—Bioingeniería, Biomateriales y Nanomedicina (CIBER-BBN http://www.ciber-bbn.es/en, CB06/01/0010), an initiative of the Instituto de Salud Carlos III (Spain) co-funded by the European Regional Development Fund (ERDF). I was recipient of a Marie Skłodowska-Curie early-stage researcher fellowship of the INSPiRE-MED project (Grant agreement ID: 813120)
Our former PhD student Kumar Motiram has been awarded an extraordinary prize by the Department of Chemistry for his PhD thesis entitled “Advances in NMR spectroscopic methodology and applications: time-efficient methods, ultra long-range heteronuclear correlation experiments and enantiospecific analysis of complex mixtures” that he defended on October 2021.
The Doctoral Commission, meeting on 20 June 2023, has awarded the extraordinary prize for theses of the doctoral programme in Chemistry defended during the academic year 2021-22… (News from the Department of Chemistry, UAB).
His thesis goals were i) the development of Nuclear Magnetic Resonance (NMR) experiments focused on efficiency in terms of time the ii) establishing new pulse sequences that facilitate the study of long-distance coupling constants fundamental for structural elucidation, iii) the development of a reliable method that allows the differentiated analysis of enantiomers (enantiospecific) directly from its original mixture (in situ) and from multiple molecules simultaneously (multicomponent).
the development of Nuclear Magnetic Resonance (NMR) experiments focused on efficiency in terms of time;
establishing new pulse sequences that facilitate the study of long-distance coupling constants fundamental for structural elucidation;
the development of a reliable method that allows the differentiated analysis of enantiomers (enantiospecific) directly from its original mixture (in situ) and from multiple molecules simultaneously (multicomponent).
The NMR experiments developed using the MFA (Multiple Fid Acquisition) approach based on the “afterglow magnetization”, allows a considerable reduction of the experimental time by acquiring several experiments at the same time, which are stored in different FIDs that can be visualized separately. In this thesis several works are presented that allow to make a structural elucidation of organic molecules in a fast, simple and unambiguous way, among them MFA-COSY / RELAY3, MFA-COSY / TOCSY, MFA-HMBC / HMBC-COSY, MFA-MBOB -COSY, MFA-TOCSY / TOCSY, MFA-HSQC / HSQC and the MFA-HSQC / Pure-shiftHSQC. Furthermore, with an adequate combination of MFA and “Spectral Aliasing” (SA), a new experiment is presented, which in addition to the experimental time improves spectral resolution and facilitates structural identification. The SA, despite being a powerful experiment to avoid signal overlapping has an important disadvantage related to the identification of each signal, to avoid this problem, for a few extra seconds, we acquired the two heteronuclear experiments in 2D, the HSQC with “Spectral Aliasing” and the standard HSQC to facilitate signal assignment.
Furthermore, in terms of improving the spectral resolution, this thesis presents two experiments following the Pure-shift methodology to eliminate the proton-proton coupling constant, using BIRD (BIlinear Rotation Decoupling) to perform heteronuclear decoupling, minimizing signal overlapping. The novelty of this work is based on the detection of multiple nuclei in the same 2D spectrum, nitrogen and carbon in the indirect dimension (F1) and proton in F2, is what is known as “Time-Shared NMR experiments”. In addition, using the same approach, a second experiment is presented that allows the calculation (via direct observation) of the heteronuclear proton-carbon and proton-nitrogen coupling constants simultaneously. The measurement of long-distance heteronuclear coupling constants remains a challenge in NMR spectroscopy, due to their tiny values and to the difficulty of their measurement. A modification in the LR-HSQMBC (changing some 180º pulses of the experiment by selective pulses irradiated in an area of the spectrum) allows the measurement of very small coupling constants (of up to 6 separation bonds). In this work, the advantages of the new LR-selHSQMBC NMR experiment are exposed and the advantages and disadvantages of both experiments are compared.
Finally, this thesis presents an innovative work related to the enantiospecific and simultaneous detection of multiple pairs of enantiomers in a mixture without prior separation or derivatization of the sample components and with minimal sample manipulation. This method is based on NMR spectroscopy and on the use of a chiral solvating agent (CSA) as chiral auxiliary. This work shows, as a proof of concept, the simultaneous enantiospecific detection of multiple enantiomeric pairs directly within the original mixture. This is demonstrated with an aqueous mixture of the essential amino acids in their D and L forms.
I would like to thank the financial support for this research by Spanish MINECO projects “Diseño y Aplicación de Nuevas Metodologías en Resonancia Magnética Nuclear” (CTQ2015-64436-P) and “Metodologías Modernas en Resonancia Magnética Nuclear de Moleculas Pequeñas” (PGC2018-095808-B-I00) and for the grant BES-2016-078903 awarded by Agencia Estatal de Investigación.
Some of our recent research work was presented at the European NMR meeting Euromar 2021 that was going to take place at Portoroz (Slovenia), but which was finally virtual from the 5th to the 8th of July 2021.
To date, the enantiospecific analysis of mixtures necessarily requires prior separation of the individual components. The simultaneous enantiospecific detection of multiple chiral molecules in a mixture represents a major challenge, which would lead to a significantly better understanding of the underlying biological processes; e.g. via enantiospecifically analyzing metabolites in their native environment. Here, we report on the first in situ enantiospecific detection of a thirty-nine-component mixture. As a proof of concept, eighteen essential amino acids (AAs) at physiological concentrations were simultaneously enantiospecifically detected using NMR spectroscopy and a chiral solvating agent. This work represents a first step towards the simultaneous multicomponent enantiospecific analysis of complex mixtures, a capability that will have substantial impact on metabolism studies, metabolic phenotyping, chemical reaction monitoring, and many other fields where complex mixtures containing chiral molecules require efficient characterization.
L. T. Kuhn, K. Motiram-Corral, T. J. Athersuch, T. Parella, M. Pérez-Trujillo, Angew. Chem. Int. Ed.59 (2020) 23615.
Some of the SeRMN staff has presented our recent research work at the biannual Spanish and IberAmerican NMR meeting, 10th GERMN biennial /9th IberAmerican/7th Iberian NMR Meeting. This year it was a virtual meeting taking place from 26 to 29 April 2021.
Pau Nolis presented an oral communication entitled “Reducing experimental time using Multiple Fid Acquisition“. P. Nolis, K. Motiram-Corral, M. Pérez-Trujillo, T. Parella.
Speeding-up NMR molecular analysis is an important research field which has been continuously advancing since NMR early days. The relevant benefits are clear and evident: i) reduce analysis time per sample => reduce analysis cost; ii) gain spectrometer time to analyze new samples => improve spectrometer efficiency. Multiple FID Acquisition (MFA) strategy consists in the design of NMR pulse sequence experiments accommodating N acquisition windows, each registering different relevant structural information. This strategy is faster than perform a traditional sequential acquisition of N separated experiments. Several design strategies and practical experiments will be shown and discussed.
Míriam Pérez-Trujillo presented an oral communication entitled “Simultaneous Enantiospecific Detection of Multiple Metabolites in Mixtures using NMR Spectroscopy“. L. T. Kuhn, K. Motiram-Corral, T. J. Athersuch, T. Parella, M. Pérez-Trujillo.
Chirality plays a fundamental role in nature, but its detection and quantification still face many limitations. To date, the enantiospecific analysis of mixtures necessarily requires prior separation of the individual components. The simultaneous enantiospecific detection of multiple chiral molecules in a mixture represents a major challenge, which would lead to a significantly better understanding of the underlying biological processes; e.g. via enantiospecifically analyzing metabolites in their native environment. Here, we report on the first in situ enantiospecific detection of a thirty-ninecomponent mixture. As a proof of concept, eighteen essential amino acids (AAs) at physiological concentrations were simultaneously enantiospecifically detected using NMR spectroscopy and a chiral solvating agent. This work represents a first step towards the simultaneous multicomponent enantiospecific analysis of complex mixtures, a capability that will have substantial impact on metabolism studies, metabolic phenotyping, chemical reaction monitoring, and many other fields where complex mixtures containing chiral molecules require efficient characterization.
Workshop limited to 4 participants (first come, first served)
Contact person:
Silvia Lope-Piedrafita, PhD ()
This course combines a comprehensive series of lectures on the technology of Magnetic resonance spectroscopy and imaging (MRS/MRI) with hands-on laboratory sessions to provide practical demonstrations of key concepts and procedures for preclinical studies.
Whether you are considering MRI as a research tool in your lab or just would like to learn more about MRI, this workshop addresses practical aspects of experimental MRI with laboratory animals and provide valuable hands-on experience on a 7 Tesla Bruker BioSpec spectrometer.
To improve the obsolescence of the oldest equipments, the following new NMR spectrometers have been installed/updated at the SeRMN-UAB Facility in May 2021:
A new AVANCE NEO console for the existing 500 MHz NMR spectrometer, equipped with an automated ATMA accessory and a compatible cryoplatform for the current TCI cryoprobe.
A New 400 MHz NMR spectrometer including a ultrashielded ASCEND magnet, automated SAMPLE-CASE sample changer, high-resolution liquid (iProbe) and solid-state (CP-MAS 4.0mm) NMR probes, and a BCU-II unit for automated sample refrigeration until 233K.
3. A New 300 MHz NMR spectrometer including a Nanobay AVANCE nanoNEO console, ultra shielded ASCEND magnet and a BBFO probe head.
4. A cooled SAMPLE-CASE sample changer and a BCU-II unit for automated sample refrigeration until 233K for the existing 600MHz NMR spectrometer
More info about the equipments and accesories can be found at the Bruker WWW site.
The purchase of these equipments has been co-financed by the Ministry of Science, Innovation, and Universities to the 2019 infrastructure call (project EQC2019-005396-P) co-financed by the European Fund for Economic and Regional Development (FEDER) through the plurirregional operating program of Spain (POPE) period 2014-2020.
