Speaker 16. Dr Kieran Tuohy. University of Reading

Transcription

1 Speaker 16 Dr Kieran Tuohy Lecturer in Food Metabonomics Department of Food Biosciences University of Reading

2 Outline Introduction The human gut microflora Functional foods PREBIOTICS Prebiotic discovery and efficacy in vitro and in vivo Case study galactooligosaccharides from lactose; pectic-oligosaccharides from citrus fruit waste Concluding remarks

3 Approaches to food waste Re-design food production process to reduce waste efficacy, impact on product, cost Stabilise, reduce, recover value (material or energy) economic relevance, technological issues Fractionate to added value products identify suitable technologies, high value products, lucrative markets FUNCTIONAL FOODS

4 The Human Gut Microflora In our bodies there are 20 x more bacteria than human cells Human genome 30,000 genes Bacteria 1,600 genes More than 1,000 bacterial species within the gut Redundancy but also genetic, evolutionary and ecological plasticity Microbial mega-genome with metabolic potential many fold greater than human

5 Microbial Interactions: Successional Development Homeostasis Gut Physiology Gut Microflora Diet: infant and adult Immune System

6 PREBIOTICS Non digestible food ingredients that selectively stimulate a limited number of bacteria in the colon, to improve host health 1. Must be non-digestible (after Gibson and Roberfroid, 1995) 2. Selective fermentation in the colon 3. Must improve health

7 Health benefits of prebiotics Immune func in children IBD Diarrhoea/IBS Colon cancer PREBIOTICS Microflora modification Blood glucose Laxation Mineral absorption Lipid metabolism

8 Prebiotic Health Benefits Selective proliferation of beneficial bacteria faecal bulking improved bowel habit prebiotics Bifidobacteria stimulate growth colonisation resistance M cell Pathogens block adhesion reduce exogenous and endogenous intestinal infection eliminate pro-ibd antigens colonocytes dendritic cell Th1 IFNg immunomodulation anti-inflammatory Tr IL-10 TGF-b Th2 allergy prevention suppress IBD inflammation lamina propria Modified from Crittenden 2006

9 Prebiotic Health Benefits Stimulation of beneficial microbial activities reduced cancer risk and IBD inflammation improved bowel habit de novo lipogenesis controlled serum lipids and cholesterol colonocytes trophic and anti-neoplastic effects induce peristalsis SCFA Ca ++ Mg ++ reduced ph increased mineral absorption prebiotics selective fermentation antagonism of pathogens and putrefactive bacteria fewer toxic bacterial metabolites reduced cancer risk Modified from Crittenden 2006

10 Functional Foods market Retail prices of functional foods are typically 30 to 500 times higher than comparable conventional foods Depending on the definition of functional foods, estimated market size is between $30 and $60 billion (Japan, US, EU) Over the past ten years the market has grown at about 10% per annum (World Bank) Prebiotics European market alone is currently worth 59 million and is expected to increase to 121 million in 2010 (Frost & Sullivan) Leading prebiotics fructans (inulin, oligofructose, galactooligosaccharides, lactulose)

11 Determining prebotic potential in vitro and in vivo In vitro gut models (batch, complex) Transverse colon Faecal microbiology (16S rrna) Faecal metabolites (GC, NMR) Ascending colon Caecum Descending colon Biomarkers of disease blood lipids faecal water genotoxicity immunology Metabolite profiles blood, urine, faeces Tuohy et al., Curr.Pharma Design (2005) Rectum

12 31 Volunteers fed Prebiotic Biscuits for 21 days Log10 cells/g faeces Total bacteria Bacteroides spp. Bifidobacterium spp. Clostridium spp. Lactobacillus spp. Active biscuits ( ) Placebo biscuits ( ) Tuohy et al., Brit. J. Nutr. (2001)

13 HYDROLYSIS Raw material Manufacturing technologies Target prebiotic property SYNTHESIS Raw material Bacterial EPS Algal polysaccharides Microbial polysaccharides Plant polysaccharides Phage hydrolysis Controlled hydrolysis Receptor activity Novel structural diversity Species specificity Enhanced persistence Equilibrium synthesis Glycosyl transfer Monosaccharides oligosaccharides Oligosaccharides

14 Synthesis Lactose Sucrose Retentate recycle Acceptors Permeate Enzyme reactor Ultrafiltration Drying β-galactosidase Dextransucrase + dextranase Alternansucrase GOS IMO

15 Prebiotics from lactose Lactose in whey is a major problem for the dairy industry Cost associated with disposal Can convert to high value prebiotics Lactose Enzyme reactor β-galactosidase Drying Manufactured by Friesland Foods (EU) Clasado (UK) Yakult (Japan) GOS Tzortzis et al., Appl. Microbiol. Biotech. (2005)

16 Galb1,4Glc Galacto-oligosaccharides GOS b-galactosidase Glc Galb1,6Galb1,4Glc Galb1,3Galb1,4Glc Galb1,4Galb1,4Glc Galb1,4Glc + Galb1,3Galb1,6Galb1,4Glc Galb1,6Galb1,6Galb1,4Glc Galb1,3Galb1,3Galb1,4Glc Galb1,6Galb1,3Galb1,4Glc Galb1,3Galb1,4Galb1,4Glc Galb1,6Galb1,4Galb1,4Glc + Oligosaccharide % (dry weight) Disaccharides 33 Trisaccharides 39 Tetrasaccharides 18 Pentasaccharides 7 Higher oligosaccharides Time (min)

17 Novel GOS Bi 2 muno GOS/B. bifidum 3-stage gut model counting by FISH with group-specific 16S rdna probes D Log 10 bacteria ml V1 V2 V3 Total bacteria Bifidobacterium spp. Lactobacillus spp. Bacteroides spp. Clostridium histolyticum group Tzortzis et al., J. Nutr. (2005)

18 Novel GOS in vivo Bi 2 muno GOS/B. bifidum Human volunteer trial, 2-weeks feeding counting by FISH with group-specific 16S rdna probes 0.2 Log10 (cell/g) g B2M/d 7g B2M/d 7g VGOS/d 7.6g Sucrose/d Total Bacteria Lactobacillus spp Bifidobacterium spp. Bacteroides spp Clostridium histolyticum group

19 Hydrolysis 50g dextran dm -3 Enzyme reactor U/F Retentate recycle Permeate Refractive Index (mv) U enzyme dm -3, 60 min residence time U enzyme dm -3, 90 min residence time U enzyme dm -3, 60 min residence time 9.5 kda 19.5 kda 1.15 kda 0.18 kda Elution Time (min)

20 Pectin composition

21 Pectic oligosaccharides High methoxy citrus pectin Low methoxy apple pectin Dead-end UF cell NMWCO Amano endo-pectinase High performance anion exchange chromatography POS1 (HM citrus pectin) Pectic oligosaccharides POS2 (LM apple pectin) PGA Hydrolysate Time (min)

22 Total Bifidobacteria Bacteroides Clostridia Lactobacilli Eubacteria Pectic oligosaccharides * 10 * Total Bifidobacteria Bacteroides Clostridia Lactobacilli Eubacteria FOS POS Batch culture, 1% CHO, ph 6.8, bacteriology by FISH Manderson et al., Appl. Environ. Microbiol (2005) Log10 cells/g

23 Antiadhesive saccharides Bacteria Decoy oligosaccharides Toxin

24 Anti-adhesive activity E. coli NCTC * * * * * * * * E. coli NCTC E. coli NCTC E. coli O111:H27 E. coli O119:H4 E. coli O128:H12 Dsv. desulfuricans L. acidophilus L. gasseri Adhesion relative to control (%)

25 Inhibition of verocytotoxin VT1 VT2 Flash-extracted POS * % Cell survivability * * * * * * * 20 0 No toxin Toxin Only Concentration of POS mg ml -1 Olana-Martin et al., FEMS Microbiol. Letts (2003)

26 Prebiotic potential of Bergamot POS Differences in bacterial population size (%) FOS Bergamot Bifidobacteria Bacteroides Lactobacilli Clostridia Eubacteria Mandalari et al., Appl Microbiol Biotech (2007)

27 Biorefining of functional food ingredients Sugar beet pulp Citrus waste Distillers grains Pretreatment Pectinase Fractionation Antioxidants Prebiotics Antiadhesives Hemicellulase Prebiotics Cellulase Fractionation Glucose

28 Concluding remarks Prebiotics appear to be a viable, high value option for food waste management Success dependant on the waste, waste transfer process and cost Cost:benefit ratio set to improve with increasing market growth Market growth dependant upon establishment of prebiotic health benefits

29 Thank you! LTN Profs Glenn Gibson, Bob Rastall (UR) Dr George Tzortzis (Clasado) Dr Giusy Mandalari, Dr Arjan Narbad (IFR)

30 Speaker 17 Tom Berry Principal Sustainability Advisor Business Programme Forum for the Future