First things first…
Why LAB?
LAB in FOOD 1 Degradation of PROTEINS
• Important for food quality, safety and nutrit
They can transform raw materials into edible food. • Can eliminate food allergens(degradation of
How? • Can help with digestibility of protein in food
• add texture and flavour
Degradation of AMINO ACIDS
• Produce beneficial compounds(e.g., ACE inh
• degrade indigestible components • Catabolism of amino acids(produce flavour)
Various ways:
• degrade harmful compounds Steps of protein degradation
• Deamination – hydrolysis of aa; removes amino group (NH₃)
• increase nutritional value 1. Extracellular/membrane bound proteinase
• Decarboxylation – convert aa to amine; remove CO₂
• increase shelf-life of fermented food peptides
• Transamination – transfer amino group of AA to keto-
Two types of fermented foods: 2. Transport of peptides into cell (Opp, DtpP,
acid(forms aa)
1. Industrial - selected LAB used as starters 3. Peptide degradation via many peptidases
LAB metabolise aa in food→ form proteolytic digests of proteins
• assure production consistency 4. Amino acid catabolism
• Flavour substance
• highly controlled fermentation(reproducible and safe)
• Produce unwanted biogenic amines(health risk)
• perform desired metabolic activities
• Maintain intracellular pH(release NH₃)
• add value to food
▪ Starter culture selected for flavour production
2. Artisanal - spontaneous fermentation/ naturally selected starters
▪ Aromatic compounds unstable
• LAB from sample/environment
• LAB influenced by surroundings
• Back-slopping - part of fermentation matrix used as starter for ACE inhibitors from casein degradation
next one (e.g., sourdough) • Functional food
Prevalence of LAB in fermented food: • Angiotensin 1→ vasoconstrictor
• ecological niches = different food matrices • Used to treat hypertension(high blood pressure) Degradation of CASEIN(protein)
• use HT-NGS for community identification • LAB produce ACE inhibitors via hydrolysis (casein and other
milk products) 1. Cell envelope proteinase (PrtP) degrades casein
• Pre-pro-domain(PP) = signal sequence for secr
removed by autolytic cleavage
Tests to confirm • Serine protease domain(PR) = active centre of
Simple test for proteolytic activity • Insert domain(I) = modulates substrate specific
Degradation of Polysaccharides - STARCH • Clear zones = proteolytic activity • Domain(A) = unknown function
• Fast; simple; cheap • Domain(B) – stabilize enzyme
• identical/different monosaccharides • Helix domain(H) = attached to cell wall to guide
Skim milk agar
• linked by a- or B glycosidic bonds • Domain(W) = hydrophilic; cell wall spacer
• Amylose = linear chains (amylase - break down) 2. Peptide transporters take up smaller peptides in
• Amylopectin = branched chains (amylopullulanase - break down) • Opp system = for casein derived peptides(ATP
• Starch degradation needs a few enzymes • Dpp system = transport hydrophobic di-, tri-, te
• amylolytic strains helps non-amylolytic (LAB) strains break down starch=useable products • DtpT system = hydrophylic di-, tri-peptides(PM
Testing for amylolytic LAB 3. Peptides further degraded by intracellular pepti
Basic testing • Endopeptidases(O; F)
• growth on media containing starch • Aminopeptidases(N; C; L)
• flood plate with iodine • Proline-specific(X; P; I; R; Q)
• iodine-starch = black/blue • Dipeptidases(D; V)
Analytical methods
• halo = amylase activity • Tripeptidases(T)
• Mass spectrophotometry = detect peptide
Molecular testing 4. Negative regulation by CodY(transcriptional regu
fragments
• genome sequencing - ID amylase genes • Branch chains = co-factors
• Complex; expensive
qPCR
• Provides more info • CodY binds to opp-operon(upstream)
• detect expression • BCAAs stimulate binding
, Degradation of phytic acid Conversion of harmful and non-nutritive substances Inhibition of mycotoxins
Phytic acid(Phytate) in plant-based foods LAB can degrade/inhibit/reduce harmful or unwanted substances • Mycotoxins = NB safety concern in food; produced by fungi
• E.g., cereals (lots of phytate) • Inhibit gathering of mycotoxins • Immunosuppression and carcinogenic effects(toxic)
• Phytate = negative charge – high affinity to positive charged • Break down ethyl carbamate in wine • Aflatoxins(AF) – group 1(human carcinogen)
food components • Degrade phytic acid • AFM1(M1) in milk is concerning – biotransformation of
• Inhibits amylases = starch digestibility inhibited • Hydrolyse “bitter” peptides in food • Ochratoxin(OTA) – group 2B(potential human carcinoge
• Anti-nutrient – chelates these nutrients = reduced LAB produce flavour/aroma substances – mask undesirable LAB can solve mycotoxin accumulation problem!
bioavailability flavour(fermented food) 1. Inhibition of • Microbial influences can be beneficial/antagon
• Excessive amounts of phytate = cause mineral deficiencies mycotoxin- • Mycotoxin production linked to growth
• Removal of phytate – non-enzyme and enzyme methods producing- • Stress and environmental factors can be linked
• Food processing = cause fractional degradation
• Enzymatic degradation = ↑efficiency
LAB in FOOD 2 fungi by LAB
• LAB influence fungal growth:
• Growth arrest = competition
• Phytases = hydrolyse phytate Bacteriocins • Compete for space/nutrient
• Cleaves P-groups – release minerals in soluble form • Produce antifungal compou
• Modification of external environment
• Reduce phytate concentration in sample Bacteriocins = cationic ribosomally synthesized AMPs
• Produce organic acids = pH↓ - affect m
LAB have been ID with phytase activity • Target cell walls of closely related species
• Produce internal phytase(mostly) AND external • Sometimes include other bioactivities 2. Mycotoxin • Adsorb to functional cell wall components
• Phytase must be secreted – functions outside the cell • Produced by LAB and other bacteria removal via • Polysaccharides and peptidoglycans(m
• Help endogenous phytases due to pH↓(during fermentation) • Lab bacteriocins = beneficial adsorption by • Differ in binding(different LAB); peptid
• Optimal pH for activity = 5.5 • Inhibit pathogens in food – natural “innate immunity” preservative binding sites
LAB
• Binding factors – growth media; bacter
• Can be added as preservative(nisin – commercial)
after heat addition); bacterial strain; in
pH(media)
Vitamins and Riboflavin • Mycotoxin-bacterial complex stability = reversa
• LAB produce various vitamins Products synthesized by LAB in food
3. Mycotoxin • 2nd common method
• E.g., folic acid; riboflavin(B2); vit C • Hydrolysis of OTA(peptidases) to ochra
degradation by
• Vitamin production during fermentation = nutritional • Adsorption = advantageous
advantageous
Gamma-Aminobutyric Acid (GABA) LAB
• Biodegradation = time-consuming
• Riboflavin production • GABA = non-protein amino acid; NB • Can convert toxin→more harmful meta
• NB functions(cellular metabolism) – redox reactions neurotransmitter
• Riboflavin deficiency common, BUT needed for development; • Production of GABA(in situ) by LAB – fermented
lactation; reproduction, etc. products enriched with GABA(functional foods)
Exopolysaccharides (EPS)
• Several LAB can produce riboflavin • EPS = biological polymers secreted by microbes(e.g., LAB)
Structural genes of Rib-operon • Viscous, stabilizing, emulsifying agents; improve texture
Flavour substances produced by LAB
• rigG = riboflavin-specific deaminase and reductase • EPS = homopolysaccharides(HoPS)/heteropolysaccharides(HePS)
• ribB = riboflavin synthase a-subunit Metabolism of Citric acid • HoPS – one type of monosaccharide(d-glucose/d-fructo
• ribA = GTP cyclohydrolase ll/3,4-dihydroxy-2-butanone-4- • LAB – produce various flavours • Simple biosynthesis process(extracellular synt
phosphate synthase • Citric acid metabolism (diacetyl; acetoin; butanediol) • HePS – 3-8 repeating units > 2 monosaccharides each(rh
• ribH = riboflavin synthase B-sununit • Amino acid metabolism (a-keto-acids; aldehydes; • Complex biosynthesis
1. PCR – screen presence of biosynthesis genes alcohols; carboxylic acids) 1. Precursor nucleotide sugars synthesized insid
2. Riboflavin-free assay media(RAM) – check ability to produce 2. Initiation and extension of repeating units
riboflavin(only LAB) 3. Export and polymerization of repeating units
3. RAM – evaluate riboflavin concentration in supernatant Antioxidants produced by LAB • EPS in dairy(cheese; yogurt) - Fats NB for texture
• Natural enrichment of riboflavin in food • LAB can produce antioxidants during fermentation • Demand for reduced-fat products NEED LAB’s EPS for te
• Riboflavin production = strain dependent • Antioxidants – scavenge free radicals(can better water-holding-capacity
• ↑Riboflavin production BY exposure to roseoflavin damage cells) • EPS in bread(sourdough)
• Roseoflavin – structural analog of riboflavin = induce mutations • Fermented blueberry extracts – antioxidant activities • Gluten-free breads = suffer from negative texture(reduc
• Mutations = ↑production capabilities • Anticancer activities(catechol = most significant) volume; freezing stability)
Why LAB?
LAB in FOOD 1 Degradation of PROTEINS
• Important for food quality, safety and nutrit
They can transform raw materials into edible food. • Can eliminate food allergens(degradation of
How? • Can help with digestibility of protein in food
• add texture and flavour
Degradation of AMINO ACIDS
• Produce beneficial compounds(e.g., ACE inh
• degrade indigestible components • Catabolism of amino acids(produce flavour)
Various ways:
• degrade harmful compounds Steps of protein degradation
• Deamination – hydrolysis of aa; removes amino group (NH₃)
• increase nutritional value 1. Extracellular/membrane bound proteinase
• Decarboxylation – convert aa to amine; remove CO₂
• increase shelf-life of fermented food peptides
• Transamination – transfer amino group of AA to keto-
Two types of fermented foods: 2. Transport of peptides into cell (Opp, DtpP,
acid(forms aa)
1. Industrial - selected LAB used as starters 3. Peptide degradation via many peptidases
LAB metabolise aa in food→ form proteolytic digests of proteins
• assure production consistency 4. Amino acid catabolism
• Flavour substance
• highly controlled fermentation(reproducible and safe)
• Produce unwanted biogenic amines(health risk)
• perform desired metabolic activities
• Maintain intracellular pH(release NH₃)
• add value to food
▪ Starter culture selected for flavour production
2. Artisanal - spontaneous fermentation/ naturally selected starters
▪ Aromatic compounds unstable
• LAB from sample/environment
• LAB influenced by surroundings
• Back-slopping - part of fermentation matrix used as starter for ACE inhibitors from casein degradation
next one (e.g., sourdough) • Functional food
Prevalence of LAB in fermented food: • Angiotensin 1→ vasoconstrictor
• ecological niches = different food matrices • Used to treat hypertension(high blood pressure) Degradation of CASEIN(protein)
• use HT-NGS for community identification • LAB produce ACE inhibitors via hydrolysis (casein and other
milk products) 1. Cell envelope proteinase (PrtP) degrades casein
• Pre-pro-domain(PP) = signal sequence for secr
removed by autolytic cleavage
Tests to confirm • Serine protease domain(PR) = active centre of
Simple test for proteolytic activity • Insert domain(I) = modulates substrate specific
Degradation of Polysaccharides - STARCH • Clear zones = proteolytic activity • Domain(A) = unknown function
• Fast; simple; cheap • Domain(B) – stabilize enzyme
• identical/different monosaccharides • Helix domain(H) = attached to cell wall to guide
Skim milk agar
• linked by a- or B glycosidic bonds • Domain(W) = hydrophilic; cell wall spacer
• Amylose = linear chains (amylase - break down) 2. Peptide transporters take up smaller peptides in
• Amylopectin = branched chains (amylopullulanase - break down) • Opp system = for casein derived peptides(ATP
• Starch degradation needs a few enzymes • Dpp system = transport hydrophobic di-, tri-, te
• amylolytic strains helps non-amylolytic (LAB) strains break down starch=useable products • DtpT system = hydrophylic di-, tri-peptides(PM
Testing for amylolytic LAB 3. Peptides further degraded by intracellular pepti
Basic testing • Endopeptidases(O; F)
• growth on media containing starch • Aminopeptidases(N; C; L)
• flood plate with iodine • Proline-specific(X; P; I; R; Q)
• iodine-starch = black/blue • Dipeptidases(D; V)
Analytical methods
• halo = amylase activity • Tripeptidases(T)
• Mass spectrophotometry = detect peptide
Molecular testing 4. Negative regulation by CodY(transcriptional regu
fragments
• genome sequencing - ID amylase genes • Branch chains = co-factors
• Complex; expensive
qPCR
• Provides more info • CodY binds to opp-operon(upstream)
• detect expression • BCAAs stimulate binding
, Degradation of phytic acid Conversion of harmful and non-nutritive substances Inhibition of mycotoxins
Phytic acid(Phytate) in plant-based foods LAB can degrade/inhibit/reduce harmful or unwanted substances • Mycotoxins = NB safety concern in food; produced by fungi
• E.g., cereals (lots of phytate) • Inhibit gathering of mycotoxins • Immunosuppression and carcinogenic effects(toxic)
• Phytate = negative charge – high affinity to positive charged • Break down ethyl carbamate in wine • Aflatoxins(AF) – group 1(human carcinogen)
food components • Degrade phytic acid • AFM1(M1) in milk is concerning – biotransformation of
• Inhibits amylases = starch digestibility inhibited • Hydrolyse “bitter” peptides in food • Ochratoxin(OTA) – group 2B(potential human carcinoge
• Anti-nutrient – chelates these nutrients = reduced LAB produce flavour/aroma substances – mask undesirable LAB can solve mycotoxin accumulation problem!
bioavailability flavour(fermented food) 1. Inhibition of • Microbial influences can be beneficial/antagon
• Excessive amounts of phytate = cause mineral deficiencies mycotoxin- • Mycotoxin production linked to growth
• Removal of phytate – non-enzyme and enzyme methods producing- • Stress and environmental factors can be linked
• Food processing = cause fractional degradation
• Enzymatic degradation = ↑efficiency
LAB in FOOD 2 fungi by LAB
• LAB influence fungal growth:
• Growth arrest = competition
• Phytases = hydrolyse phytate Bacteriocins • Compete for space/nutrient
• Cleaves P-groups – release minerals in soluble form • Produce antifungal compou
• Modification of external environment
• Reduce phytate concentration in sample Bacteriocins = cationic ribosomally synthesized AMPs
• Produce organic acids = pH↓ - affect m
LAB have been ID with phytase activity • Target cell walls of closely related species
• Produce internal phytase(mostly) AND external • Sometimes include other bioactivities 2. Mycotoxin • Adsorb to functional cell wall components
• Phytase must be secreted – functions outside the cell • Produced by LAB and other bacteria removal via • Polysaccharides and peptidoglycans(m
• Help endogenous phytases due to pH↓(during fermentation) • Lab bacteriocins = beneficial adsorption by • Differ in binding(different LAB); peptid
• Optimal pH for activity = 5.5 • Inhibit pathogens in food – natural “innate immunity” preservative binding sites
LAB
• Binding factors – growth media; bacter
• Can be added as preservative(nisin – commercial)
after heat addition); bacterial strain; in
pH(media)
Vitamins and Riboflavin • Mycotoxin-bacterial complex stability = reversa
• LAB produce various vitamins Products synthesized by LAB in food
3. Mycotoxin • 2nd common method
• E.g., folic acid; riboflavin(B2); vit C • Hydrolysis of OTA(peptidases) to ochra
degradation by
• Vitamin production during fermentation = nutritional • Adsorption = advantageous
advantageous
Gamma-Aminobutyric Acid (GABA) LAB
• Biodegradation = time-consuming
• Riboflavin production • GABA = non-protein amino acid; NB • Can convert toxin→more harmful meta
• NB functions(cellular metabolism) – redox reactions neurotransmitter
• Riboflavin deficiency common, BUT needed for development; • Production of GABA(in situ) by LAB – fermented
lactation; reproduction, etc. products enriched with GABA(functional foods)
Exopolysaccharides (EPS)
• Several LAB can produce riboflavin • EPS = biological polymers secreted by microbes(e.g., LAB)
Structural genes of Rib-operon • Viscous, stabilizing, emulsifying agents; improve texture
Flavour substances produced by LAB
• rigG = riboflavin-specific deaminase and reductase • EPS = homopolysaccharides(HoPS)/heteropolysaccharides(HePS)
• ribB = riboflavin synthase a-subunit Metabolism of Citric acid • HoPS – one type of monosaccharide(d-glucose/d-fructo
• ribA = GTP cyclohydrolase ll/3,4-dihydroxy-2-butanone-4- • LAB – produce various flavours • Simple biosynthesis process(extracellular synt
phosphate synthase • Citric acid metabolism (diacetyl; acetoin; butanediol) • HePS – 3-8 repeating units > 2 monosaccharides each(rh
• ribH = riboflavin synthase B-sununit • Amino acid metabolism (a-keto-acids; aldehydes; • Complex biosynthesis
1. PCR – screen presence of biosynthesis genes alcohols; carboxylic acids) 1. Precursor nucleotide sugars synthesized insid
2. Riboflavin-free assay media(RAM) – check ability to produce 2. Initiation and extension of repeating units
riboflavin(only LAB) 3. Export and polymerization of repeating units
3. RAM – evaluate riboflavin concentration in supernatant Antioxidants produced by LAB • EPS in dairy(cheese; yogurt) - Fats NB for texture
• Natural enrichment of riboflavin in food • LAB can produce antioxidants during fermentation • Demand for reduced-fat products NEED LAB’s EPS for te
• Riboflavin production = strain dependent • Antioxidants – scavenge free radicals(can better water-holding-capacity
• ↑Riboflavin production BY exposure to roseoflavin damage cells) • EPS in bread(sourdough)
• Roseoflavin – structural analog of riboflavin = induce mutations • Fermented blueberry extracts – antioxidant activities • Gluten-free breads = suffer from negative texture(reduc
• Mutations = ↑production capabilities • Anticancer activities(catechol = most significant) volume; freezing stability)
