Chemistry and Biochemistry of Agri-Food Molecules
A.Y. 2023/2024
Learning objectives
This teaching aims to give the student an advanced knowledge in the chemical and biotechnological fields about molecules (mainly non-informational) of agri-food interest.
Part I (Chemistry), dedicated to the description of such molecules in terms of structure and chemical reactivity, will be followed by Part II (Biochemistry) which has the main objectives to highlight the structure / function relationships of these molecules either in organisms that produce them and in the end users and to stimulate the interest towards potential molecular targets of biotechnological interventions.
The laboratory practice included in this course have the aim to make familiar the main extraction techniques of some of the molecules faced in the teaching and some methods for the in vitro evaluation of their bioactivity.
Part I (Chemistry), dedicated to the description of such molecules in terms of structure and chemical reactivity, will be followed by Part II (Biochemistry) which has the main objectives to highlight the structure / function relationships of these molecules either in organisms that produce them and in the end users and to stimulate the interest towards potential molecular targets of biotechnological interventions.
The laboratory practice included in this course have the aim to make familiar the main extraction techniques of some of the molecules faced in the teaching and some methods for the in vitro evaluation of their bioactivity.
Expected learning outcomes
The knowledge of the potential for intervention on these molecules, through chemical, enzymatic or process modifications aimed at improving their overall quality (chemical and physical stability, solubility, interactions and biological activity) and the basic ability to independently undertake studies and applications in the research contexts, and in the wide and interdisciplinary professional contexts.
Lesson period: Second semester
Single course
This course can be attended as a single course.
Course syllabus and organization
Single session
Responsible
Lesson period
Second semester
Course syllabus
The Course is organized in two Teaching Units:
- Unit 1: Chemistry of Agri-Food Molecules (3 CFU)
- Unit 2: Biochemistry of Agri-Food Molecules (5 CFU)
The description of each Teaching Unit is reported below.
CHEMISTRY OF AGRI-FOOD MOLECULES
Lessons: Introduction. Role of the secondary metabolites in the interactions between plants and environment. Allelopatic substances and elicitors; Repellent and attractors. Biosynthetic building blocks for secondary metabolism. AcetylCoA, shikimic acid, mevalonic acid and methyleritritol phosphate.
Reaction mechanisms in the biosynthesis of natural substances. Alkylation reactions: nucleophilic substitution (SAM), electrophilic addition, Wagner-Meerwein transpositions; Reactions based on the formation of the enolate anion: aldol reactions and Claisen reactions. Formation of the C-N bond: imines, Mannich reaction, transaminations. Decarboxylation reactions: α-amino acids, α-keto acids, β-keto acids; Oxidation and reduction reactions. Dehydrogenase (NAD and FAD), Oxidase, Oxygenase, Baeyer-Villiger monooxygenase; Oxidative coupling of phenols; O-glycosidation reactions (formation and hydrolysis); C-glycosidation reactions.
The acetate pathway: fatty acids and polyketides. Saturated and unsaturated fatty acids, prostaglandins, thromboxanes, leukotrienes; Macrolides; Linear and aromatic polyketides. Post-polyketide synthetase modifications. Mixed biogenetic pathway of THC.
The shikimate pathway: aromatic amino acids and phenylpropanoids. Route of shikimic acid; Benzoic acids; Tannins: classification. Hydrolyzable tannins; Phenylpropanoids; Aromatic amino acids; Cinnamic acids and cinnamic alcohols; Lignans and lignin; Phenylpropenes; Coumarins; Aromatic polyketides; MIXED biogenesis (acetate + shikimate pathways); Flavonoids; Stilbenoids; Terpenoid quinones.
The mevalonate and methyleritritol-phosphate pathways: terpenoids and steroids. Terpenes. Isoprenic rule. Head-tail coupling. Mevalonic acid and methyleritritol phosphate; Hemiterpenes: isoprene, DMAPP and IPP; Linear and cyclic monoterpenes; Irregular monoterpenes; Iridoid monoterpenes; Linear and cyclic sesquiterpenes; Linear and cyclic diterpenes; Linear and cyclic triterpenes; Phytosterols; Triterpenoid saponins; Vitamins of group D; Cardiac glycosides; Tetraterpenes; Natural rubber and gutta-percha
Alkaloids. Anti-nutritional principles: Notes on the biosynthesis of alkaloids starting from the corresponding amino acids. Structure and function of imines and Mannich reaction; Alkaloids classification; Alkaloids deriving from ornithine; Biosynthesis of tropane and pyridine nuclei; Alkaloids derived from aromatic amino acids.
Laboratory tutorials: steam extraction of lavender essential oil, TLC chromatography of essential oil; Hydrolysis of triglycerides-saponification.
BIOCHEMISTRY OF AGRI-FOOD MOLECULES
The meaning of molecular diversity. General aspect of metabolism. Typologies of enzyme reactions. General aspects of secondary metabolism.
Amino acids as precursors of secondary metabolites. Biosynthesis of amino acids (general aspects). Features of aromatic amino acids. Biosynthesis of aromatic amino acids. Bacteria-fungi and plants specificity of the prephenate branch. Biosynthesis of tyrosine from phenylalanine: phenylalanine hydroxylase (phenylketonuria). Shikimate pathway regulation in bacteria and plants (hints). EPSP synthase, the action site of glyphosate. Mechanism of the glyphosate activity. Key amino acid residues in the interaction glyphosate/EPSP synthase. Glyphosate-tolerant EPSP synthases. Glyphosate-resistant plants. Advantages and disadvantages in the use of glyphosate (hints).
Phenylpropane: basic structure of phenylpropanoids and tyrosine and phenylalanine. Phenylpropanoid pathway. Phenylalanine ammonia lyase and tyrosine ammonia lyase. Cinnamate hydroxylase. The typical reactions of phenylpropanoid biosynthesis. Acetyl-CoA, precursor of the polyketide moiety of flavonoids. Acetyl-CoA carboxylase. Chalcone synthase. From chalcone to the other flavonoids (hints). The other type-III polyketide synthases. The transcriptional regulation and the promoter of the chalcone synthase gene. Natural roles of flavonoids: vicenin-2; involvement in the nodulation process; the anthocyans and the colours.
Metabolic precursors of terpenoids. Mevalonate pathway. Acetoacetyl-CoA thiolase and HMG-CoA synthase: cholesterogenic and ketogenic forms. HMG-CoA reductase: statins sensitivity. The methylerythritol phosphate pathway. The DXP reductoisomerase: the fosmidomycin target. Statins and fosmidomycin in the study of the metabolic source of terpenoids. Subcellular localization and taxonomy specificity of MVA and MEP pathways. The elongation phase of terpenoid biosynthesis: prenyltransferases and their subcellular localization; primary and secondary refinement (hints).
Taxol (paclitaxel): discovering, source, biosynthesis (hints), subcellular localization, cytotoxicity, physiological role, effect of tubulin structure on cytotoxicity. Taxol production and its improvement perspectives. Strategies of metabolic engineering. Holistic approach of metabolic engineering: the case of catharanthine. Pros and cons in the use of plant cell coltures for the production of molecules of interest. Plant cell coltures for the production of complex recombinant proteins. The case of glucocerebrosidase and the Gaucher illness. Other cell systems for metabolite production: bioreactors and biofuels (hints). The case of biohydrogen (dark fermentation, photofermentation). Potential biotechnological use of lignocellulosic wastes (hints).
Vitamin K, role and biosynthetic origin. The phylloquinone role in the plant. The menaquinone role in bacteria. Functional distinction between ubiquinone and menaquinones. Vitamin D (cholecalciferol, ergocalciferol, calcitriol, ercalcitriol). Metabolic source of cholecalciferol. Vitamin A (retinol and aldehyde and acid derivatives). Retinal role in the vision system (rhodopsin and photopsins). Biosynthetic source of retinal and beta-carotene. Vitamin A deficit. Food fortification and biofortification. Golden Rice. Golden Rice DNA constructs (GR1 and GR2 prototypes).
Reactive oxygen and other radical species biologically significant. Biological sources of radicals. Mechanism and ROS reactivity. Oxidative damages in the cell. Oxidative stress defence mechanisms. Glutathione and glutathione peroxidase. Reduced glutathione regeneration: glutathione reductase. Glucose 6-phosphate dehydrogenase and the relation with favism and malaria. Isouramil and divicine induced-redox-cycle. Thiolic functional homologues of glutathione. Meaning and evaluation of antioxidant power in biological/experimental and food systems. Vitamin E (tococromanols). Biosynthetic source of tocopherols. Chloroplasts tocopherols (plastoglobules). Antioxidant activity of tocopherols. Antioxidant triad. Ascorbic acid (vitamin C) and redox activity. Biosynthetic source of ascorbic acid (animals, plants). Ascorbic acid role in plants and animals. Ascorbic acid as co-factor of alpha-chetoglutarate-dependent hydroxylases. The ascorbic acid, the collagen and the scurvy. Essential fatty acids.
Storage seed tissues in dicots and monocots and its molecular macro composition. Osborne's protein fractionation. 2S albumins. 7S and 11S globulins. Prolamines: types and structures. Maize prolamines: zeins. Subcellular protein sorting pathways. Prolamines and protein bodies. Cupin and prolamine protein superfamilies. Functional-technological properties of wheat gluten. Gluten proteins: glutenins and gliadins. Electrophoretic analysis of gliadins. Main chemical and enzyme flour-additives and mechanism of action. Food and biotechnological use of transglutaminase. Flour allergens (hints). General and molecular aspects of celiac sprue. Applications to reduce celiac pathogenesis (chemical and enzyme modifications of gluten, "glutenases", etc.).
Plant "toxic" proteins. Ribosome-inactivating proteins (RIPs). The N-glycosidase activity of RIPs. The ricin cytotoxicity mechanism. Antiviral RIP activity. General structure of antibodies. The immunotoxins.
The following topics will be deepened and/or refreshed to allow the full comprehension of topics faced during the course: transamination reactions; recombinant expression of proteins (hints); protein mutagenesis (hints); general canonical procedure to produce GMO plants (hints); the Double Strand Breaks and genome editing through CRISPR/Cas9 system (hints); GS/GOGAT system; cytochrome P450 superfamily; hints on general structure of promoters, biotechnological importance of promoters and promoter analysis; functional omics (metabolomics); immunolabeling; photo-affinity labeling; general procedure for plant cell colture (hints); protein transmembrane domains through primary structure analysis; molecular docking (hints); epitopes mapping (hints); protein allergen identification approach.
Laboratory practices: enzyme reaction inhibition; the IC50 value meaning and its extrapolation from dose-response plots; the colorimetric assay for phosphatase; the phosphatases and the intestinal alkaline phosphatase; organization of a laboratory report.
- Unit 1: Chemistry of Agri-Food Molecules (3 CFU)
- Unit 2: Biochemistry of Agri-Food Molecules (5 CFU)
The description of each Teaching Unit is reported below.
CHEMISTRY OF AGRI-FOOD MOLECULES
Lessons: Introduction. Role of the secondary metabolites in the interactions between plants and environment. Allelopatic substances and elicitors; Repellent and attractors. Biosynthetic building blocks for secondary metabolism. AcetylCoA, shikimic acid, mevalonic acid and methyleritritol phosphate.
Reaction mechanisms in the biosynthesis of natural substances. Alkylation reactions: nucleophilic substitution (SAM), electrophilic addition, Wagner-Meerwein transpositions; Reactions based on the formation of the enolate anion: aldol reactions and Claisen reactions. Formation of the C-N bond: imines, Mannich reaction, transaminations. Decarboxylation reactions: α-amino acids, α-keto acids, β-keto acids; Oxidation and reduction reactions. Dehydrogenase (NAD and FAD), Oxidase, Oxygenase, Baeyer-Villiger monooxygenase; Oxidative coupling of phenols; O-glycosidation reactions (formation and hydrolysis); C-glycosidation reactions.
The acetate pathway: fatty acids and polyketides. Saturated and unsaturated fatty acids, prostaglandins, thromboxanes, leukotrienes; Macrolides; Linear and aromatic polyketides. Post-polyketide synthetase modifications. Mixed biogenetic pathway of THC.
The shikimate pathway: aromatic amino acids and phenylpropanoids. Route of shikimic acid; Benzoic acids; Tannins: classification. Hydrolyzable tannins; Phenylpropanoids; Aromatic amino acids; Cinnamic acids and cinnamic alcohols; Lignans and lignin; Phenylpropenes; Coumarins; Aromatic polyketides; MIXED biogenesis (acetate + shikimate pathways); Flavonoids; Stilbenoids; Terpenoid quinones.
The mevalonate and methyleritritol-phosphate pathways: terpenoids and steroids. Terpenes. Isoprenic rule. Head-tail coupling. Mevalonic acid and methyleritritol phosphate; Hemiterpenes: isoprene, DMAPP and IPP; Linear and cyclic monoterpenes; Irregular monoterpenes; Iridoid monoterpenes; Linear and cyclic sesquiterpenes; Linear and cyclic diterpenes; Linear and cyclic triterpenes; Phytosterols; Triterpenoid saponins; Vitamins of group D; Cardiac glycosides; Tetraterpenes; Natural rubber and gutta-percha
Alkaloids. Anti-nutritional principles: Notes on the biosynthesis of alkaloids starting from the corresponding amino acids. Structure and function of imines and Mannich reaction; Alkaloids classification; Alkaloids deriving from ornithine; Biosynthesis of tropane and pyridine nuclei; Alkaloids derived from aromatic amino acids.
Laboratory tutorials: steam extraction of lavender essential oil, TLC chromatography of essential oil; Hydrolysis of triglycerides-saponification.
BIOCHEMISTRY OF AGRI-FOOD MOLECULES
The meaning of molecular diversity. General aspect of metabolism. Typologies of enzyme reactions. General aspects of secondary metabolism.
Amino acids as precursors of secondary metabolites. Biosynthesis of amino acids (general aspects). Features of aromatic amino acids. Biosynthesis of aromatic amino acids. Bacteria-fungi and plants specificity of the prephenate branch. Biosynthesis of tyrosine from phenylalanine: phenylalanine hydroxylase (phenylketonuria). Shikimate pathway regulation in bacteria and plants (hints). EPSP synthase, the action site of glyphosate. Mechanism of the glyphosate activity. Key amino acid residues in the interaction glyphosate/EPSP synthase. Glyphosate-tolerant EPSP synthases. Glyphosate-resistant plants. Advantages and disadvantages in the use of glyphosate (hints).
Phenylpropane: basic structure of phenylpropanoids and tyrosine and phenylalanine. Phenylpropanoid pathway. Phenylalanine ammonia lyase and tyrosine ammonia lyase. Cinnamate hydroxylase. The typical reactions of phenylpropanoid biosynthesis. Acetyl-CoA, precursor of the polyketide moiety of flavonoids. Acetyl-CoA carboxylase. Chalcone synthase. From chalcone to the other flavonoids (hints). The other type-III polyketide synthases. The transcriptional regulation and the promoter of the chalcone synthase gene. Natural roles of flavonoids: vicenin-2; involvement in the nodulation process; the anthocyans and the colours.
Metabolic precursors of terpenoids. Mevalonate pathway. Acetoacetyl-CoA thiolase and HMG-CoA synthase: cholesterogenic and ketogenic forms. HMG-CoA reductase: statins sensitivity. The methylerythritol phosphate pathway. The DXP reductoisomerase: the fosmidomycin target. Statins and fosmidomycin in the study of the metabolic source of terpenoids. Subcellular localization and taxonomy specificity of MVA and MEP pathways. The elongation phase of terpenoid biosynthesis: prenyltransferases and their subcellular localization; primary and secondary refinement (hints).
Taxol (paclitaxel): discovering, source, biosynthesis (hints), subcellular localization, cytotoxicity, physiological role, effect of tubulin structure on cytotoxicity. Taxol production and its improvement perspectives. Strategies of metabolic engineering. Holistic approach of metabolic engineering: the case of catharanthine. Pros and cons in the use of plant cell coltures for the production of molecules of interest. Plant cell coltures for the production of complex recombinant proteins. The case of glucocerebrosidase and the Gaucher illness. Other cell systems for metabolite production: bioreactors and biofuels (hints). The case of biohydrogen (dark fermentation, photofermentation). Potential biotechnological use of lignocellulosic wastes (hints).
Vitamin K, role and biosynthetic origin. The phylloquinone role in the plant. The menaquinone role in bacteria. Functional distinction between ubiquinone and menaquinones. Vitamin D (cholecalciferol, ergocalciferol, calcitriol, ercalcitriol). Metabolic source of cholecalciferol. Vitamin A (retinol and aldehyde and acid derivatives). Retinal role in the vision system (rhodopsin and photopsins). Biosynthetic source of retinal and beta-carotene. Vitamin A deficit. Food fortification and biofortification. Golden Rice. Golden Rice DNA constructs (GR1 and GR2 prototypes).
Reactive oxygen and other radical species biologically significant. Biological sources of radicals. Mechanism and ROS reactivity. Oxidative damages in the cell. Oxidative stress defence mechanisms. Glutathione and glutathione peroxidase. Reduced glutathione regeneration: glutathione reductase. Glucose 6-phosphate dehydrogenase and the relation with favism and malaria. Isouramil and divicine induced-redox-cycle. Thiolic functional homologues of glutathione. Meaning and evaluation of antioxidant power in biological/experimental and food systems. Vitamin E (tococromanols). Biosynthetic source of tocopherols. Chloroplasts tocopherols (plastoglobules). Antioxidant activity of tocopherols. Antioxidant triad. Ascorbic acid (vitamin C) and redox activity. Biosynthetic source of ascorbic acid (animals, plants). Ascorbic acid role in plants and animals. Ascorbic acid as co-factor of alpha-chetoglutarate-dependent hydroxylases. The ascorbic acid, the collagen and the scurvy. Essential fatty acids.
Storage seed tissues in dicots and monocots and its molecular macro composition. Osborne's protein fractionation. 2S albumins. 7S and 11S globulins. Prolamines: types and structures. Maize prolamines: zeins. Subcellular protein sorting pathways. Prolamines and protein bodies. Cupin and prolamine protein superfamilies. Functional-technological properties of wheat gluten. Gluten proteins: glutenins and gliadins. Electrophoretic analysis of gliadins. Main chemical and enzyme flour-additives and mechanism of action. Food and biotechnological use of transglutaminase. Flour allergens (hints). General and molecular aspects of celiac sprue. Applications to reduce celiac pathogenesis (chemical and enzyme modifications of gluten, "glutenases", etc.).
Plant "toxic" proteins. Ribosome-inactivating proteins (RIPs). The N-glycosidase activity of RIPs. The ricin cytotoxicity mechanism. Antiviral RIP activity. General structure of antibodies. The immunotoxins.
The following topics will be deepened and/or refreshed to allow the full comprehension of topics faced during the course: transamination reactions; recombinant expression of proteins (hints); protein mutagenesis (hints); general canonical procedure to produce GMO plants (hints); the Double Strand Breaks and genome editing through CRISPR/Cas9 system (hints); GS/GOGAT system; cytochrome P450 superfamily; hints on general structure of promoters, biotechnological importance of promoters and promoter analysis; functional omics (metabolomics); immunolabeling; photo-affinity labeling; general procedure for plant cell colture (hints); protein transmembrane domains through primary structure analysis; molecular docking (hints); epitopes mapping (hints); protein allergen identification approach.
Laboratory practices: enzyme reaction inhibition; the IC50 value meaning and its extrapolation from dose-response plots; the colorimetric assay for phosphatase; the phosphatases and the intestinal alkaline phosphatase; organization of a laboratory report.
Prerequisites for admission
Principles of Organic Chemistry.
Principles of Biochemistry and Molecular Biology.
Principles of Biochemistry and Molecular Biology.
Teaching methods
Frontal lectures and laboratory practices.
Teaching Resources
The teaching material will be suggested and/or will be available on the ARIEL sites of the course (https://eraggcmia.ariel.ctu.unimi.it ; http://fforlanibmia.ariel.ctu.unimi.it ).
Assessment methods and Criteria
The examination will be composed of a written part ("Chemistry of Agri-Food molecules") and an oral part ("Biochemistry of Agri-Food molecules").
The written part consists in three questions: a) on the structural analysis of several natural organic substances and discussion of their possible biosynthetic pathway; b) one general question over specific topics discussed during the course; c) a detailed biosynthesis, with mechanisms of reaction, of a particular secondary metabolite, to be chosen over those cited in the unit program.
In the interview, the report of laboratory practices, a presentation of an in-depth topic of choice, and the answer to 1-2 questions about the program of the unit will be evaluated.
The modalities of the single parts of the examination will be furtherly detailed during the lessons by the teachers and will be described in the ARIEL sites of the course (https://eraggcmia.ariel.ctu.unimi.it ; http://fforlanibmia.ariel.ctu.unimi.it ).
The final score will be expressed in thirtieths, and will be calculated weighting (on the basis of the CFU) the score of the single parts of the examination.
The written part consists in three questions: a) on the structural analysis of several natural organic substances and discussion of their possible biosynthetic pathway; b) one general question over specific topics discussed during the course; c) a detailed biosynthesis, with mechanisms of reaction, of a particular secondary metabolite, to be chosen over those cited in the unit program.
In the interview, the report of laboratory practices, a presentation of an in-depth topic of choice, and the answer to 1-2 questions about the program of the unit will be evaluated.
The modalities of the single parts of the examination will be furtherly detailed during the lessons by the teachers and will be described in the ARIEL sites of the course (https://eraggcmia.ariel.ctu.unimi.it ; http://fforlanibmia.ariel.ctu.unimi.it ).
The final score will be expressed in thirtieths, and will be calculated weighting (on the basis of the CFU) the score of the single parts of the examination.
Biochemistry of Agri-Food Molecules
BIO/10 - BIOCHEMISTRY
CHIM/06 - ORGANIC CHEMISTRY
CHIM/06 - ORGANIC CHEMISTRY
Single bench laboratory practical: 8 hours
Lessons: 36 hours
Lessons: 36 hours
Professor:
Forlani Fabio
Chemistry of Agri-Food Molecules
BIO/10 - BIOCHEMISTRY
CHIM/06 - ORGANIC CHEMISTRY
CHIM/06 - ORGANIC CHEMISTRY
Single bench laboratory practical: 8 hours
Lessons: 20 hours
Lessons: 20 hours
Professors:
Contente Martina Letizia, Pinto Andrea
Educational website(s)
Professor(s)
Reception:
By appointment (request by email)
DeFENS - Sezione di Scienze Chimiche e Biomolecolari (ex DISMA; bldg 21040 Facoltà di Scienze Agrarie e Alimentari)