Lysine production

INDUSTRIAL LYSINE PRODUCTION

1. LYSINE PRODUCTION BY FERMENTATION PROCESS 1 SOURAV MONDAL MSc MICROBIOLOGY

2. INTRODUCTION AN OVERWIEW OF AMINO ACIDS AND LYSINE

3. 3 AMINO ACIDS ARE THE MONOMERIC SUBUNITS BUILDING BLOCKS OF PROTEINS . ALL PROTEINS IN OUR WORLD ARE MADE FROM THE UBIQUITOUS SET OF 20 AMINO ACIDS AMINO ACIDS ARE ORGANIC COMPOUNDS, THEY HAVE TWO FUNCTIONAL GROUPS, A CARBOXYL GROUP (-COOH) AND AN AMINE GROUP (-NH2) BONDED TO THE SAME CARBON ATOM (THE α CARBON) THE AMINO GROUP IS BASI C IN NATURE AND THE CARBOXYL GROUP IS ACIDIC IN NATURE THEY ALSO CONTAIN A HYDROGEN ATOM AND A SIDE CHAIN (R GROUP) WHICH IS SPECIFIC TO EACH AMINO ACID. THE KEY ELEMENTS OF AN AMINO ACID ARE CARBON (C) , HYDROGEN (H) , OXYGEN (O) , AND NITROGEN (N) . ACCORDING TO THE REQUIREMENTS OF THE AMINO ACIDS IN DIET , THEY ARE DIVIDED INTO TWO CLASSES - ESSENTIAL AMINO ACIDS AND NON ESSENTIAL AMINO ACIDS . WHAT IS AN AMINO ACID ?

4. 4 AMINO ACID PRODUCED IN THIS PHASE

5. WHAT IS LYSINE ?? 5 H I H 2 N—C—COOH I CH 2 I CH 2 I CH 2 I CH 2 I NH 2 LYSINE (Lys, K) L- LYSINE (2,6-DIAMINOHEXANOIC ACID) IS AN ESSENTIAL AMINO ACID FOR THE NUTRITION OF HUMANS AND OTHER ANIMALS. OTHER NAMES OF LYSINE ARE Lys , K AND HAVING MOLECULAR FORMULA (C 6 H 14 N 2 O 2 ) IT IS USED FOR SUPPLEMENTING THE CEREAL PROTEINS LACKING THIS AMINO ACID. THIS AMINO ACID IS USED MEDICALLY AS A NUTRIENT IN FORM OF SUPPLEMENTS AND MEDICINES LYSINE IS PRESENT IN TWO FORMS , L - FORM AND D - FORM

6. HISTORY… 6

7. 7 PRODUCTION OF LYSINE OVERVIEW OF LYSINE PRODUCTION NATURALLY AND INDUSTRIALLY

8. 8 PRODUCTION OF LYSINE NATURALLY , TAKES PLACES BY TWO BIOSYNTHETIC PATHWAYS THE DAP ( DIAMINOPIMELIC ACID ) PATHWAY FOUND IN BACTERIA (e.g. Corynebactrium glutamicum), ALGAE , HIGHER PLANTS AND OOMYCOTA THE AAA ( α – AMINO – ADIPATE) PATHWAY FOUND IN FUNGI AND EUGLENOIDS NATURALLY (LYSINE BIOSYNTHESIS)

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10. 10 I U F D O INBOUND LOGISTICS UP STREAM PROCESSING FERMENTATION DOWN STREAM PROCESSING OUTBOUND LOGIDTICS RAW MATERIALS MICROBIAL CULTURE MEDIA CULTURE OPTIMIZATION MEDIA FORMULATION MEDIA UTILIZATION & LYSINE PRODUCTION SEPERATION EXTRACTION PURIFICATION PACKAGING OF L- LYSINE

11. 11 INBOUND LOGISTICS

12. 12 FERMENTATION MEDIA CARBON SOURCE  CANE MOLASSES NITROGEN SOURCE  CORN STEEP LIQUOR / SOYABEAN MEAL MINERAL AND SALTS  KH 2 PO 4 / K 2 HPO 4 / CaCO 3 TRACE ELEMENTS  CORN STEEP LIQUOR ANTI FOAMING AGENT  PEG-2000, SILICONE BASED OIL. RAW MATERIALS INSTRUMENTS

13. 13 IT IS A GRAM POSITIVE BACTERIUM IT IS ROD SHAPED BACTERIUM THEY LIVES IN CHAINS OR IN CLUSTERS THE BACTERIUM IS USUALLY PRESENT IN SOIL IT IS NON MOTILE THE BACTERIUM IS A NON SPORE FORMER THE BACTERIUM IS A NON PATHOGENIC CULTURE ( Corynebacterium glutamicum ATCC 21526)

14. 14 UP STREAM PROCESS

15. 15 MEDIA FORMULATION INGREDIENTS AMOUNT ( GRAM / LITRE ) GLUCOSE 100 g UREA 5 g BIOTIN 0.01 g LEUCINE 0.4 g MgSO 4 2.85 g KH 2 PO4 / K 2 HPO4 0.5 g MnSO 4 0.016 g CaCO 3 20 g (NH 4 ) 2 SO 4 46 g DISTILLED WATER 1000 L MEDIA PREPRATION FOR GROWTH OF CULTURE AND INOCULLUM DEVELOPEMENT

16. 16 DONE BY GENOME SHUFFLING RIBOZOME ENGINEERING PROTOPLAST FUSION CULTURE OPTIMIZATION DONE TO OVEREXPRESS THE EXPRESSION OF INDIVIDUAL BIOSYNTHETIC GENE Lys C AND Dap A

17. INOCULLUM DEVELOPEMENT 17 500ml 1 litre L . F 5 L SEED FERMENTER 50 L PILOT FERMENTER 100 L 100ml PRE CULTURE SLANT TO PRODUCTION FERMENTER

18. 18 FERMENTATION

19. CHARACTERISTICS OF THE FERMENTATION 19

20. 20 CULTURE MEDIA PREPATED BY MIXING PROCESSED WATER, GLUCOSE & NUTRIENTS FERMENTATION PERFORMED IN FED-BATCH MODE AND UNDER AEROBIC CONDITION IN BATCH PHASE SEED CULTURE IS FED INTO THE FERMENTER PREVIOUSLY FILLED WITH FERMENTATION BATCH MEDIUM WHEN GLUCOSE IS EXHAUSTED, THE BATCH FERMENTATION PHASE ENDS AND FED-BATCH FERMENTATION PHASE STARTS THE FERMENTATION PROCESS

21. 21 DOWN STREAM PROCESSING

22. 22 D S P - SCHEMATIC

23. SEPERATION OF L-LYSINE 23 LIQUID L-LYSINE L-LYSINE SEPERATED FROM CELLS BY ULTRAFILTRATION L-LYSINE SELECTIVELY ABSORBED BY ION-EXCHANGE CHROMATOGRAPHY L-LYSINE ELUTED FROM CHROMATOGRAPHY RESINS BY WASHING WITH AQUEOUS AMMONIA SOLUTION

24. 24 LIQUID L - LYSINE MOTHER LIQUOR (CANE MOLASSES) HCL EXTRACTION OF L-LYSINE

25. PURIFICATION OF L-LYSINE 25 ACIDIFIED L - LYSINE PURE L – LYSINE LIQUID OR POWDERED L- LYSINE OR CRYSTALINE L-LYSINE

26. 26 OUTBOUND LOGISTICS

27. 27 PACKAGING OF L-LYSINE 25 KG BAG 50 g BOTTLE PURIFIED L-LYSINE

28. 28 USES OF L-LYSINE FIELD OF APPLICATION USED IN IN FOOD NUTRITIONAL SUPPLEMENTS, FOOD DECORANTS IN BEVERAGES NUTRITIONAL SPORTS DRINK IN PHARMACEUTICAL PREVENTING AND TREATING COLD SORES CAUSED BY HERPES SIMPLEX LABIALIS VIRUS IN HEALTH AND PERSONAL CARE COSMETICS, PERSONAL CARE PRODUCTS, BABY PRODUCTS, BATH PRODUCTS, EYE MAKEUP, SAVING KITS IN AGRICULTURE / ANIMAL FEED / POULTRY ANIMAL FEED AND USED IN FERTILIZERS IN OTHER INDUSTRIES AS INTERMEDIATE TO PRODUCE OTHER ORGANIC CHEMICALS

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