1. Ethanol Production

2. Feedstock 1.Biomass 2.Starch

3. Definitions Biomass Lignocellulosic biomass the relatively large amounts of heterogeneous matter produced by living organisms. It includes residues originating from plants, animals, and microorganisms Biomaterials whose composition is dominated by lignified cell walls from vegetative plants.

4. Polysaccharide and lignin content of representative lignocellulosic feedstocks a,b ComponentLignocellulosic Material (Straw) Glucan31.9 Xylan18.9 Arabinan2.1 Mannan0.2 Galactan0.6 Lignin22.8 Sum of above76.5 a values are percentages on a dry weight basis b data taken from Puls and Schuseil (1992) c measured as Klason lignin

5. Biomass (cellulose, Hemicellulose, lignin) Milled Biomass Ethanol Prehydrolysate Liquid (xylose, 2-furaldehyde) Pretreated solid (cellulose, lignin) Mechanical chipping/grinding Pretreatment (Dilute acid, 180 o C) Fermentation Enzymatic saccharification (fungal cellulases) Hydrolyzed Solid (lignin) Hydrolysate Liquid (glucose) Ethanol fermentation

6. CompositionsRangeAverage Starch61.0 – 78.071.7 Protein6.0 – 12.09.5 Fat3.1 – 5.74.3 Ash1.1 – 3.91.4 Cellulose a 3.3 – 4.33.3 Pentosans b 5.8 – 6.66.2 Sugars c 1.0 – 3.02. other1.0 Chemical Composition of Corn (percent of dry matter) a plus lignin b as xylose c as glucose

7. Corn Wet Milling Clean Corn Steeps Steep water (6.5%) Mill and Screen System Germ System Wash Centrifugal Separator Losses (1%) Adapter from C.R. Keim, 1999 Germ (8%) Fiber (10.2%) Prime Starch (68%) Gluten (6.3%)

8. Enzymatic Starch Conversion Starch Slurry Liquefaction Maltodextrin Purification Saccharification Refining Isomerization Adapter from H.S. Olsen, 1995 To Fuel Ethanol Maltose Syrups Fructose Syrups  -amylase Glucoamylase/ Pullulanase Glucose Syrups Mixed Syrups Glucose Isomerase

9. Feeder Pathways for Glycolysis

13. Entry of Fructose into Glycolysis

14. Fate of Pyruvate Lactate dehydrogenase –during exercise –Reversible in liver –Location: cytoplasm Pyruvate dehydrogenase –Source of AcetylCoA –Irreversible reaction –Location: Mitochondria Ethanol synthesis –In yeast, some bacteria –Location cytoplasm

16. Reduction of pyruvate to ethanol (microorganism) It occurs by the 2 reactions shown below: The overall reaction of alcohol fermentation: Glc+2ADP +2P---------> 2 Ethanol + 2CO2 + 2ATP + 2 H2O

17. Pyruvate decarboxylase mechanism

18. There is no net oxidation-reduction in the conversion of Glc into Ethanol, NAD+ is used first and made it later!

19. Active site of Alcohol dehydrogenase

20. Pyruvate decarboxylase is present in brewer’s and baker’s yeast. CO2 produced during alcohol fermentation is responsible for the characteristic carbonation of champagne. In baking, CO2 fermentation by pyruvate decarboxylase during fermentation of dough due to CO2, dough rises. Alcohol dehydrogenase metabolizes alcohol. TPP carries “active aldehyde” groups The pyruvate decarboxylase reaction is the first reaction we see that TPP is involved. TPP------> Vit B1. If B1 is not enough

21. More about TPP TPP plays an important role in the cleavage of bonds adjacent to a carbonyl group such as the decarboxylation of alpha-ketoacids and in chemical rearrangements involving transfer of an activated aldehyde group from one C to another. The functional part of TPP is the thiazolium ring. The proton at C-2 of the ring is relatively acidic, loss of this proton, produces an active site in TPP. TPP is involved in the following reactions –1. Pyruvate decarboxylase –2. Pyruvate dehydrogenase –3. Alpha-Ketoglutaratedehyrogenase –4. Transketolase

22. Microbial fermentation yield other end products of commercial value: Lactate and ethanol are the common products of microbial fermentation Clostridium acetobutyricum, ferments starch to butanol and acetone. Here comes industrial fermentation, purpose is to make important products from readily available material (like starch) by using microorganism.