Lactic acid is an important product in food industry and is used as a preservative, acidulant and flavouring agent. It also finds applications in the textile and pharmaceutical industries and in the chemical industry as a raw material for the production of lactate ester, propylene glycol, 2,3-pentanedione, propanoic acid, acrylic acid, acetaldehyde and dilactide (1,2). In recent times, lactic acid has gained considerable importance because it acts as a starting material in the production of poly lactic acid (PLA), which is biodegradable and a sustainable bioplastic material (3, 4). Both fermentation and chemical synthesis routes are used for lactic acid production. The biotechnological production of lactic acid has received a significant amount of importance over the chemical route because of the optical purity of products obtained and a practical alternative to environmental pollution caused by the petrochemical industry. Moreover, the limited supply of petrochemical resources is also overcome (5). Hence, fermentation processes are being developed which are cost effective and can survive at an industrial scale.
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There have been numerous investigations on the development of biotechnological processes for lactic acid production, with the ultimate objectives of enabling the process to be more efficient and economical (5). Lactic acid fermentation is usually followed by product recovery and/or purification. Lactic acid production through free cell fermentation provides about 50% of the world supply, but the productivity is very low in conventional batch processes due to low concentration of the cells in the broth (6). To overcome this, immobilization method is employed, which can maintain high cell concentration and improve production rates of lactic acid while reducing medium requirements and inhibitions (7). Fermentations with immobilized cells also provide relative ease of product separation, reuse of biocatalysts, high volumetric productivity, improved process control and reduced susceptibility of cells to contamination (Göksungur, 1999)
Entrapment of whole cells in calcium alginate gel beads is the most widely used method for lactic acid bacteria immobilization due to its simplicity, nontoxicity, mild gelation conditions and ease of use (Boyaval, 1988, 11-17). However, the Ca2+ ions in alginate gels are susceptible to cation chelating agents such as phosphate and lactate due to which beads loose their stability. During lactic acid production, calcium ions which stabilize this type of gel are displaced by lactate ions produced by lactic acid bacteria leading to disruption or dissolution of the beads (Li, 1996). Further the cells start leaking out from the beads into the medium, and subsequently grow very rapidly in the medium than in the beads (Tanaka, 1989). Hence attempts have been made to improve the stability of Ca-alginate beads by covering the beads with poly-L-lysine (Champagne, 1992) or treating them with polyethyleneimine, glutaraldehyde and hexamethylenediamine (Bódalo, 1997). Coating of Ca-alginate beads with chitosan is another method to increase the stability of beads (Göksungur, 2004). Other types of immobilization supports and matrices include polyvinyl alcohol containing sodium alginate (9), calcium pectate gels and chemically modified chitosan beads (10). Lactobacillus cells have also been immobilized on fruit pieces for food grade lactic acid production (8).
Batch fermentation of liquid pineapple waste to lactic acid was performed using Lactobacillus delbrueckii subsp. delbrueckii ATCC 9646 immobilized with calcium alginate. Various parameters like sodium alginate concentration, bead diameter, initial pH and temperature on cell growth, glucose consumption and lactic acid production were investigated (Ani Idris, Process Biochemistry 41 (2006) 1117-1123). Lactobacillus casei subsp. casei ATCC 39392 was entrapped in barium alginate beads for lactic acid production using whey and corn steep liquor supplemented with glucose. The volumetric productivity obtained in batch mode was 0.625g/L.hr (Saeed Mirdamadi, Iranian journal of Biotechnology, Vol. 6, No. 1,2008). Lactobacillus casei cells immobilized with calcium pectate gel were utilized for lactic acid production using whey medium. This fermentation gave a yield of 94.37% and the lactic acid concentration achieved was 32.95 g/l in batch mode (P. S. Panesar, Appl Microbiol Biotechnol (2007) 74:35-42).
Lactobacillus helveticus cells were entrapped in κ-carrageenan and locust bean gum matrix and were used in a continuous two-stage process wherein immobilization was combined with free cell system for lactic acid production from whey permeate/yeast extract medium. A high productivity of 19-22g/L.hr was obtained in continuous mode with controlled pH (Christophe Lacroix, Enzyme and Microbial Technology 38 (2006) 324-337). Continuous lactic acid production was attained using unique plastic composite support (PCS) by immobilizing Lactobacillus casei subsp. rhamnosus (ATCC 11443). The productivity and yield obtained were 5.8g/L.hr and 70% respectively ( A. L. Pometto III, Appl Microbiol Biotechnol (2001) 57:626-630). Lactobacillus casei cells immobilized in chitosan-coated Ba-alginate capsule gave stable production of lactic acid in batch mode with relatively low cell leakage during repeated batch fermentation. Here the productivity obtained was 2.7g/L.hr (Ho Nam Chang, Enzyme and Microbial Technology 19:426-433, 1996). The growth cultivation pattern of Lactobacillus rhamnosus cells entrapped in alginate/starch liquid-core capsules was compared with free cells system in a growth medium containing whey supplemented with yeast extract. The concentration of viable cells in encapsulated matrix was 4.8-1010 cfu/ml of capsule volume, and that of free cells cultured in was found to be 5.7 -109 cfu/ml. The cell biomass productivity in capsule was found low as compared to free cell cultivation because lactic acid inhibition on cell growth in the capsules limited the mass transfer rate (R. Dembczynski, Enzyme and Microbial Technology 31 (2002) 111-115). A pH controlled batch system, with glucose and lactate as substrate, was employed for the production of propionic and acetic acid using immobilized Propionibacterium shermenii with calcium alginate (Bonita A. Glatz, Enzyme and Microbial Technology 22:409-414, 1998).
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Lactobacillus delbrueckii was immobilized on polyurethane foam (PUF) and operated in dual reactor system with pH control. Lactic acid was produced with a productivity of 5g/L.hr (V. Rangaswamy.2008. Letters in Applied Microbiology ISSN 0266-8254). Lactobacillus delbrueckii ZU-S2 immobilized in calcium alginate was utilized for conversion of cellulosic hydrolysate into lactic acid in a continuos system. The yield and productivity obtained were 92.4% and 5.746g/L.hr. respectively (Xueliang Shen, World J Microbiol Biotechnol (2006) 22:1109-1114). Calcium alginate immobilized Lactobacillus bifermentans was used for lactic acid production from wheat bran hemicellulosic hydrolysate in batch process. The yield and productivity obtained were 83% and 1.17g/L.hr respectively (Sebastien Givry, World J Microbiol Biotechnol (2008) 24:745-752). Calcium alginate immobilized Lactobacillus delbrueckii IFO 3202 were used for lactic acid production from beet molasses which gave 90% effective yield and productivity of 13.92g/L.hr in continuous system (Göksungur, Y. Journal of Chemical Technology and Biotechnology. Vol. 74, pp. 131-136). Lactococcus lactis IO-1 cells encapsulated in microcapsules of alginate membrane were used for lactic acid production from glucose in batch mode, which gave a productivity of 1.8g/L.hr. Continuous system using packed bed reactor with pH control and recycling of broth gave a productivity of 4.5g/L.hr (Sarote Sirisansaneeyakul, J Ind Microbiol Biotechnol (2007) 34:381-391).