Lactic Acid

•  First organic acid produced by microorganisms
• Commercial production by fermentation or synthetic processes

Uses

• Lactic acid is used as acidulant  (give sour, or acidic flavor to foods)/ flavouring/ pH buffering agent or inhibitor of bacterial spoilage in a wide variety of processed foods. It is non-volatile, odorless, has a mild acidic taste and is classified as GRAS (generally regarded as safe). 
• It is a very good preservative and pickling agent. 
• Addition of lactic acid aqueous solution to the packaging of poultry and fish increases their shelf life.
• The esters of lactic acid are used as emulsifying agents in baking foods (glyceryl lactostearate, glyceryl lactopalmitate). 
• Technical grade lactic acid is used in leather tanning industries. Various textile finishing operations and acid dying of food require low cost technical grade lactic acid for pH adjustment, hardening baths for cellophanes used in food packaging, terminating agent for phenol formaldehyde resins, lithographic and textile printing developers, adhesive formulations etc
• Lactic acid has many pharmaceutical and cosmetic applications and formulations in topical ointments, lotions, anti acne solutions, dialysis applications. 
• Calcium lactate can be used for calcium deficiency therapy and as anti caries agent. 
• Polymers of lactic acids are biodegradable thermoplastics. These polymers are transparent and their degradation can be controlled by adjusting the composition, and the molecular weight. Their properties approach those of petroleum derived plastics. 
• Polymers of lactic acids have medical applications as sutures (stitches), orthopaedic implants, controlled drug release etc.
• Poly L-lactic acid with low degree of polymerization can help in controlled release or degradable mulch films for large-scale agricultural applications.

Biosynthesis

Lactic acid fermentation is a metabolic process by which glucose is converted into pyruvate and then anaerobially, into lactate. Lactic acid fermentation is an anaerobic fermentation reaction that occurs in some bacteria and animal cells, such as muscle cells invoving lactate dehydrogenase enzyme

Theoretically, from I mole of glucose, 2 moles of lactate obtained - 90% yield obtained practically.

• Microorganisms

Commercial production using fermentaion with Lactic acid bacteria

Two kind:

Homofermentative-Maximum substrate conversion into lactic acid. Not much production of byproducts or cell biomass. Used in commercial lactic acid production. Eg. Lactobacillus delbrueckii, L. bulgaricus

Heterofermentative- Produce some lactic acid, and a great amount of byproducts like CO₂, ethanol, acetic acid etc. Not suitable for commercial lactic acid production. Eg. Leuconostoc mesenteroides

 Lactobacillus has complex nutritional requirements, as they have lost their ability to synthesize their own growth factors. They cannot grow solely on carbon source and inorganic nitrogen salts. Organisms such as Rhizopus oryzae have less limiting nutritional requirements and can utilize starch feed stocks. They are able to produce pure L (+) lactic acid. Saccharomyces cerevisiae and Kluyveromyces lactis also can produce of L (+) lactic acid.

The choice of an organism primarily depends on the carbohydrate to be fermented. Lactobacillus delbreuckii are able to ferment sucrose. Lactobacillus bulgaricus is able to use lactose/whey. Lactobacillus helveticus is able to use both lactose and galactose. Lactobacillus amylophylus and Lactobacillus amylovirus are able to ferment starch. Lactobacillus lactis can ferment glucose, sucrose and galactose. Lactobacillus pentosus have been used to ferment sulfite waste liquor.

 L. casei, Rhizopus oryzae and S. lactis are also employed. These are all facultative anaerobes and bioreactors need not be completely anaerobic

Stock culture maintained in skim milk media. Milk used for building up of inoculum since it is an ideal medium. Culture transferred and incubated in increasing amounts to sterile skim milk and finally whey.

Selection of microorganism depends on the carbon source being used

• Media and Conditions

1) Carbon Sources

1. Glucose, maltose, sucrose, lactose etc.
2. Crude substrates like Corn starch, potato starch, molasses, whey, sulfite lyes etc
3. Starch is pretreated with amylase/acid hydrolysis to break down into simple sugars (Glucose, maltose)
4. Sulphite lyes steam stripped to eliminate SO₂ and alkali treatment to remove lignin-pretreatment
5. Sugar conc. Usually adjusted to 5-20%, usually 12%.
6. Whey is a major source since it contains, lactose

Choice of carbon source depends on

Ø  Availability

  Ø Treatments required prior to fermentation

  Ø  Cost

        Whey

• Byproduct of dairy industry, cheese industry
• Value addition by converting whey into lactic acid
• It contains, carbohydrates, nitrogenous substances, vitamins and salts
• Large quantities produced annually as part of dairy industries

2) Nitrogen sources

Ammonium salts like ammonium hydrogen phosphate (0.25%)

3) Growth factors & mineral sources

• Complex nutrients, especially Vit B 12 complexes
• Supplied by enriching the medium with malt sprouts and other crude vegetable sources (care taken to avoid overheating of such materials, during drying)
• Minerals added as per the requirement of microorganisms

4)  pH

• 5.5-6.5 - maintained using CaCO₃
• The acid produced can accumulate and should be neutralized or removed for improved growth and yield
• At low pH values, no other bacterial contamination. High pH values can inhibit the gowth of Lactobacilli and inhibit fermentation

 5)  Temperature

• Depend on the microorganism used
• 45-50⁰C for Lactobacillus delbreuckii or Lactobacillus bulgaricus
• 30⁰C for Lactobacillus pentosus, L. casei, Streptococcus lactis

6) Aeration and Agitation

• anaerobic fermentation; No supply of sterile air
• Facultative anaerobes are used for fermentation so, complete air removal not needed
• Broth usually stirred to keep CaCO₃ in suspension

 7)  Time

• 5-10 days
• Usually completed using 12-13% glucose supply

8) Yield

• L⁺ lactic acid produced commercially.  93-95% of weight of glucose supplied is converted to lactic acid.
• Free lactic acid toxic to the organism- removed continuously by electro dialysis or continuous culture.
• Fermentation carried out in wooden lined tank/stainless steel tank to avoid corrosion.

 Recovery

Method of recovery depends on the type of grade required- food grade, technical grade or plastic grade. Broth heated to dissolve calcium lactate, then filtered. Calcium precipitated by adding H₂SO₄. Lactic acid concentrated and further purified.

 Plastic grade lactic acid by esterification with methanol after concentration

Technical grade lactic acid- Calcium present removed as calcium sulfate dihydrate by precipitation. Filtration to remove the precipitate of CaSO₄ dihydrate and further concentration to 35-40% lactic acid by evaporation.

Food grade lactic acid- Fermented broth filtered followed by acidification of filtrate. Calcium precipitated as CaSO_4, washed_, filtrate treated with activated carbon to remove organic impurities. Concentration of crude lactic acid to 25% solids by evaporation. Refining and evaporation repeated to obtain 65% total acidity. Sodium Ferro cyanide treatment to remove heavy metals (eg., copper) by precipitation to remove discolouration. Ion exchange chromatography finally to remove traces of contamination.

 References

• A Text Book of Industrial Microbiology: (2nd edition By Wulf Crueger & Anneliese Crueger)
• Industrial Microbiology: (By Casida L. E.New Age international (P) ltd publications)