Principle of spectroscopy and applications to quantitative biology
A.A. 2024/2025
Obiettivi formativi
The course consists of classroom lectures, in which the topics are illustrated both with slides and by carrying out exercises, and experimental laboratory activities, in which the students, divided in small groups, carry out different experiments concerning spectroscopy, mass spectrometry and circular dichroism.
Theory modules
The primary objective of the course is to introduce the basic concepts, principles, and techniques of modern analytical chemistry. This will provide the student with the basic knowledge of the theoretical foundations and applications of the main instrumental analytical techniques. Both common protocols and advanced methods for qualitative and quantitative analysis of different classes of biological molecules will be addressed, with an emphasis towards their applications and their importance in the field of biology and life sciences.
Laboratory modules
Within the scope of the laboratory module, experimental procedures/ protocols and calibration methods as well as basic calculations (also using a spreadsheet) related to instrumental analytical chemistry will be described. The laboratory experiments will focus on instrumental atomic and molecular methods for the qualitative and quantitative determination of unknown samples. Furthermore, NMR, MS and Circular Dichroism experiments will be conducted and discussed.
Theory modules
The primary objective of the course is to introduce the basic concepts, principles, and techniques of modern analytical chemistry. This will provide the student with the basic knowledge of the theoretical foundations and applications of the main instrumental analytical techniques. Both common protocols and advanced methods for qualitative and quantitative analysis of different classes of biological molecules will be addressed, with an emphasis towards their applications and their importance in the field of biology and life sciences.
Laboratory modules
Within the scope of the laboratory module, experimental procedures/ protocols and calibration methods as well as basic calculations (also using a spreadsheet) related to instrumental analytical chemistry will be described. The laboratory experiments will focus on instrumental atomic and molecular methods for the qualitative and quantitative determination of unknown samples. Furthermore, NMR, MS and Circular Dichroism experiments will be conducted and discussed.
Risultati apprendimento attesi
Theory modules
At the end of the course the student will have learnt about the main spectroscopic analytical methods as well as several advanced and hyphenated techniques. Furthermore, they will be able to solve some qualitative and quantitative analytical problems by choosing the most suitable instrumental technique. The student will also be able to identify the structure of simple organic compounds through the study of their FTIR, Raman, Nuclear Magnetic Resonance, and mass spectra.
Laboratory modules
The main outcomes will be (a) the ability of choosing and utilising the most suitable instrumental analytical atomic and molecular spectroscopic technique for the determination of unknown samples; (b) the use of the most common calibration methods for quantitative determinations starting from the preparation of standard samples up to the processing of related data (also with spreadsheets). Moreover, the student will be able to analyse and discuss the results of NMR, MS and Circular Dichroism experiments, with a particular attention towards analysis of molecules of biological interest.
Finally, the ability to work with a group of colleagues will be acquired.
At the end of the course the student will have learnt about the main spectroscopic analytical methods as well as several advanced and hyphenated techniques. Furthermore, they will be able to solve some qualitative and quantitative analytical problems by choosing the most suitable instrumental technique. The student will also be able to identify the structure of simple organic compounds through the study of their FTIR, Raman, Nuclear Magnetic Resonance, and mass spectra.
Laboratory modules
The main outcomes will be (a) the ability of choosing and utilising the most suitable instrumental analytical atomic and molecular spectroscopic technique for the determination of unknown samples; (b) the use of the most common calibration methods for quantitative determinations starting from the preparation of standard samples up to the processing of related data (also with spreadsheets). Moreover, the student will be able to analyse and discuss the results of NMR, MS and Circular Dichroism experiments, with a particular attention towards analysis of molecules of biological interest.
Finally, the ability to work with a group of colleagues will be acquired.
Periodo: annuale
Modalità di valutazione: Esame
Giudizio di valutazione: voto verbalizzato in trentesimi
Corso singolo
Questo insegnamento non può essere seguito come corso singolo. Puoi trovare gli insegnamenti disponibili consultando il catalogo corsi singoli.
Programma e organizzazione didattica
Edizione unica
Responsabile
Periodo
annuale
Programma
Course syllabus - Principles of spectroscopy - (Theoretical lessons)
Introduction to spectroscopy
Properties of electromagnetic and light/matter interaction: an overview
-Fundamentals of spectrophotometric methods and instruments.
-Principles and methods of atomic spectroscopy (adsorption, emission, fluorescence). Qualitative and quantitative aspects of atomic spectroscopy.
-Principles and methods of molecular spectroscopy.
-Electronic spectroscopy - Qualitative and quantitative aspects of UV-visible light absorption measurements. The Lambert-Beer equation and its application in quantitative spectroscopic analysis. Analysis of mixtures of compounds.
-Luminescence (fluorescence and phosphorescence). Applications and photoluminescence methods in quantitative analysis. Luminescent probes for cell imaging.
-Vibrational spectroscopy - IR and Raman spectroscopies, functional groups, and molecular structure.
-Scanning electron microscopy (SEM): basic principles, detectors, and applications in chemical-physical analysis of surfaces.
- Transmission electron microscopy (TEM): basic principles, detectors and applications in biology.
NMR (Nuclear Magnetic Resonance)
Theoretical basis of spin and resonance
The NMR observables: chemical shift, coupling constants and integral.
Mono-dimensional spectra for 1H and 13C
Bi-dimensional experiments: COSY, TOCSY and HSQC
Overhauser's effect: theory, examples and applications.
The analysis of 3D structure and conformation of biomolecules and macromolecules by NMR
The analysis of ligand-protein interaction by NMR
Mass Spectrometry (MS)
General principles and fields of application of mass spectrometry: the mass spectrum and its characteristics, isotopic mass and isotopic abundance, resolution and accuracy.
The mass spectrometry instrumentation: sample inlet systems; ion sources and ionization techniques: Electronic Impact (EI), Chemical Ionization (CI), Fast Atom Bombardment (FAB), ElectroSpray Ionization (ESI), Matrix Assisted Laser Desorption Ionization (MALDI). Mass Analysers: Magnetic Field/Sector analyser, Quadrupole(s), Ion Trap, Time Of Flight (TOF), Fourier Transform Ion Cyclotron Resonance (FT-ICR), Orbitrap, Inductively Coupled Plasma Mass Spectrometry (ICP-MS). Notes on detectors
Fragmentation rules and spectra interpretation of small organic molecules and peptides
Mass spectrometry in biology: analysis of peptides, proteins and glycans and cells/tissues (MALDI Imaging). Hyphenated techniques (HPLC-MS and GC-MS) and tandem mass spectrometry (MS-MS). High-performance liquid chromatography: scope of HPLC, components of a HPLC system, columns used in HPLC, types of detectors, difference between HPLC and UPLC, application of HPLC.
Applications of mass spectrometry for the study of protein structures, drug metabolism, metabolic diseases
Circular Dichroism (CD)):
- General principles
- Application of CD in the study of peptide and protein structures
- Analysis of CD spectra of proteins
- Use of free software for the estimation of protein secondary structure percentage
Course syllabus - Principles of Spectroscopy (Laboratory)
Security and lab practices information.
Basic calculations on preparation/dilution of solutions in molar and ppm scales.
Use of spreadsheets for analytical data processing.
Qualitative and quantitative experiments based on the main spectroscopic and hyphenated techniques presented during the lectures in class.
Introduction to spectroscopy
Properties of electromagnetic and light/matter interaction: an overview
-Fundamentals of spectrophotometric methods and instruments.
-Principles and methods of atomic spectroscopy (adsorption, emission, fluorescence). Qualitative and quantitative aspects of atomic spectroscopy.
-Principles and methods of molecular spectroscopy.
-Electronic spectroscopy - Qualitative and quantitative aspects of UV-visible light absorption measurements. The Lambert-Beer equation and its application in quantitative spectroscopic analysis. Analysis of mixtures of compounds.
-Luminescence (fluorescence and phosphorescence). Applications and photoluminescence methods in quantitative analysis. Luminescent probes for cell imaging.
-Vibrational spectroscopy - IR and Raman spectroscopies, functional groups, and molecular structure.
-Scanning electron microscopy (SEM): basic principles, detectors, and applications in chemical-physical analysis of surfaces.
- Transmission electron microscopy (TEM): basic principles, detectors and applications in biology.
NMR (Nuclear Magnetic Resonance)
Theoretical basis of spin and resonance
The NMR observables: chemical shift, coupling constants and integral.
Mono-dimensional spectra for 1H and 13C
Bi-dimensional experiments: COSY, TOCSY and HSQC
Overhauser's effect: theory, examples and applications.
The analysis of 3D structure and conformation of biomolecules and macromolecules by NMR
The analysis of ligand-protein interaction by NMR
Mass Spectrometry (MS)
General principles and fields of application of mass spectrometry: the mass spectrum and its characteristics, isotopic mass and isotopic abundance, resolution and accuracy.
The mass spectrometry instrumentation: sample inlet systems; ion sources and ionization techniques: Electronic Impact (EI), Chemical Ionization (CI), Fast Atom Bombardment (FAB), ElectroSpray Ionization (ESI), Matrix Assisted Laser Desorption Ionization (MALDI). Mass Analysers: Magnetic Field/Sector analyser, Quadrupole(s), Ion Trap, Time Of Flight (TOF), Fourier Transform Ion Cyclotron Resonance (FT-ICR), Orbitrap, Inductively Coupled Plasma Mass Spectrometry (ICP-MS). Notes on detectors
Fragmentation rules and spectra interpretation of small organic molecules and peptides
Mass spectrometry in biology: analysis of peptides, proteins and glycans and cells/tissues (MALDI Imaging). Hyphenated techniques (HPLC-MS and GC-MS) and tandem mass spectrometry (MS-MS). High-performance liquid chromatography: scope of HPLC, components of a HPLC system, columns used in HPLC, types of detectors, difference between HPLC and UPLC, application of HPLC.
Applications of mass spectrometry for the study of protein structures, drug metabolism, metabolic diseases
Circular Dichroism (CD)):
- General principles
- Application of CD in the study of peptide and protein structures
- Analysis of CD spectra of proteins
- Use of free software for the estimation of protein secondary structure percentage
Course syllabus - Principles of Spectroscopy (Laboratory)
Security and lab practices information.
Basic calculations on preparation/dilution of solutions in molar and ppm scales.
Use of spreadsheets for analytical data processing.
Qualitative and quantitative experiments based on the main spectroscopic and hyphenated techniques presented during the lectures in class.
Prerequisiti
Knowledge of the fundamentals of general and inorganic chemistry, organic chemistry, physics and mathematics.
Metodi didattici
The course consists of classroom lectures, in which the topics are illustrated both with slides and by carrying out exercises, and experimental laboratory activities, in which the students, divided into small groups, carry out different experiments concerning spectroscopy, mass spectrometry and circular dichroism.
Materiale di riferimento
Skoog, West, Holler, Crouch, Principles of Instrumental Analysis, Seventh Edition
A. Randazzo. Guide to NMR spectral interpretation. A problem-based approach to determining the structures of small organic molecules. Loghia Publishing.
Slides of the lessons available on Ariel website of the course
Additional scientific papers will be presented and provided to students during the course lectures.
A. Randazzo. Guide to NMR spectral interpretation. A problem-based approach to determining the structures of small organic molecules. Loghia Publishing.
Slides of the lessons available on Ariel website of the course
Additional scientific papers will be presented and provided to students during the course lectures.
Modalità di verifica dell’apprendimento e criteri di valutazione
The exam will consist of two written final tests (one for each semester) with a maximum score of 30/30 cum laude. The tests will contain exercises and questions about the topics of both theoretical lessons and laboratory experiments, including spectra interpretation.
The final mark will be calculated as the average of the two tests.
Any additional information and details on the assessment methods will be explained during the course.
The final mark will be calculated as the average of the two tests.
Any additional information and details on the assessment methods will be explained during the course.
CHIM/01 - CHIMICA ANALITICA - CFU: 2
CHIM/02 - CHIMICA FISICA - CFU: 2
CHIM/03 - CHIMICA GENERALE ED INORGANICA - CFU: 1
CHIM/06 - CHIMICA ORGANICA - CFU: 5
CHIM/02 - CHIMICA FISICA - CFU: 2
CHIM/03 - CHIMICA GENERALE ED INORGANICA - CFU: 1
CHIM/06 - CHIMICA ORGANICA - CFU: 5
Esercitazioni: 48 ore
Lezioni: 56 ore
Lezioni: 56 ore
Docente/i
Ricevimento:
Su appuntamento
Dipartimento di Chimica, via Golgi 19 - Milano