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Polylactic acid is aliphatic polyester made from lactic acid monomers; it was first produced in 1932 by Carothers. The production of lactic acid from renewable raw materials by microbial fermentation has made the production of PLA very economic and made PLA an important member of biodegradable plastics. A mixture of PLA and polyglycolic acid was used in sutures, this was first reported commercial use of PLA. These sutures were available under the brand name Vicryl in the American market in 1974 (Jamshidian et. al., 2010). Along with biodegradability PLA is also biocompatible in human body and hence has been widely used in production of implants, sutures and also used for drug delivery. PLA is first biopolymer that is produced on a commercial scale and it can be biologically degraded fully. PLA is produced in two steps; first lactic acid is produced by microbial fermentation of carbohydrate rich agricultural by products followed by polymerization. PLA is considered as the best and most competitive bio-plastic produced from renewable resources. Nature Works LLC is the market leader in PLA production and its plant in Blair, USA has a production capacity of 300 million pounds per year (Madhavan Nampoothiri et. al., 2010). PURAC Biomaterials is another major producer of PLA and manufactures resorbable homo and hetero polymers (Madhavan Nampoothiri et. al., 2010).
Plastics produced from petrochemicals are synthetic polymers and fail to degrade naturally. About 140 million tons of synthetic polymers are manufactured per year across the globe. The reason for synthetic plastics being used widely can be attributed to low cost of production as they have been used for a long time and has established production line. Synthetic plastics are very durable and strong mechanical properties. Degradability is a serious problem for synthetic plastics (Jamshidian et. al., 2010). Synthetic plastics have a remarkable effect on the eco system making accumulation of these a serious threat to the environment (Madhavan Nampoothiri et. al., 2010). The cost of recycling and management of synthetic plastics is a major one. As result of this there is need for economical and biodegradable substitute for synthetic plastics. Along with the environmental factor, there are concerns with fluctuating petroleum price which adds to increasing demand for biodegradable plastics form easily available raw material.
The structure of the polymer and the raw material used for its production, imparts the property of biodegradability to the polymer. Biopolymers are made from variety of natural resources and are categorised in figure 1. Polyesters are widely used as biodegradable plastics as they can be hydrolysed easily. Today many biodegradable polyesters are available and many are in developmental process, these are: PLA (polylactic acid), PHB (polyhydroxybutyrate), PCL (polycaprolactonate), PHA (polyhydroxyalkanoates), PET (polyethylene terephthalate), PHH (polyhydroxyhexanoate), PHV (polyhydroxyvalerate), PHB (polyhydroxybutyrate), AAC (Aliphatic Aromatic copolyesters), polybutylene adipate and polymethylene adipate (Madhavan Nampoothiri et. al., 2010). PLA could be considered as a substitute for petroleum based plastics as it can be produced on a large scale by microbial fermentation using carbohydrate rich substances as raw materials.
Figure 1: sources of biopolymers
Figure 2: Lactic acid isomers. Top: L(+) lactic acid and bottom: D(-) lactic acid (Source: Polylactic acid)
Lactic acid (PLA monomer) production
Lactic acid was discovered by Carl Wilhelm Scheele in 1780 and was considered as a milk component till 1857, when Louis Pasteur found lactic acid is produced as a result of fermentation by microorganisms (Wee et. al., 2006). Lactic acid is used widely in many industries like food, cosmetics, drink, pharmaceutical, chemical, electronic etc. There are two optical isomeric forms of lactic acid: L(+) lactic acid and D(-) lactic acid (Figure 2). These enantiomers are produced by different enzymes known as Lactate dehydrogenases. L(+) lactic acid is usually produced by microbes, D(-) may be also present with L(+) (Jem et. al., 2010).
Figure 3: (adapted and modified from Wee et. al., 2006) Different manufacturing methods of lactic acid. SSF: simultaneous saccharification and fermentation.
Recovery & purification
HCN and catalyst additionLactic Acid can be manufactured either by chemical synthesis or by microbial fermentation (Figure 3). Fermentative production of lactic acid is generally method choice for production of lactic acid over chemical synthesis. Petrochemicals are starting material for chemical synthesis and there is price fluctuation as well as availability issues with petrochemicals. Only racemic mixture of DL-lactic acid can be produced by chemical synthesis. From the top 15 lactic acid producers in the world only Musashino, Japan uses chemical synthesis method for producing DL-lactic acid (Jem et. al., 2010). Microbial fermentation has many advantages over chemical synthesis: low cost of raw materials, process runs at low temperature and lower pressure than chemical synthesis, less energy consumption and it is environmental friendly (Madhavan Nampoothiri et. al., 2010). Optically pure L(+) or D(-) lactic acid is produced by microbial fermentation. Optical purity is very essential for imparting physical properties of PLA. Bacteria and fungi are the two categories of microbes used in production of lactic acid. The type of enantiomer produced depends on the organism used for fermentation. The fermentation process can be classified on the type of bacteria used as Homo-fermentative and hetero-fermentative. In homo-fermentative method 2 molecules of lactic acid are produced per molecule of glucose, it has higher yield and less by products; so
is widely used in industrial production of lactic acid. The hetero-fermentation produces lower level of lactic acid along with other metabolites like acetic acid, ethanol, carbon dioxide etc. (Hartman, 1998). Fermentation is carried out at acidic PH (about 6), around 40 degree temperature and anaerobic or very low oxygen level. Racemization may occur at high temperature, thus it is vital to avoid high temperature to obtain optically pure lactic acid (Jem et. al., 2010).
Typically, batch, fed batch or continuous fermentations are used for producing lactic acid. Continuous cultures provide high productivity whereas batch and fed batch assure higher concentrations (Hofvendahl et. al., 2000). Kwon et. al. demonstrated a novel and a successful attempt to produce lactic acid using continuous fermentation using Lactobacillus rhamnosus. They produced 92 g/L of lactic acid and productivity of 57 g/(L.h), by using membrane cell cycle bioreactors connected in series (Kwon et. al., 2001). Wee et. al. have reviewed different fermentation approaches used for lactic acid production in greater detail . End product inhibition is an major issue during lactic acid production and thus lactic acid must be continuously removed from fermentation broth. Downstreaming process for product recovery involves biomass separation by filtration, centrifugation and decantation; followed by chromatography and esterification / distillation, to separate the impurities (Jem et. al., 2010).
Lactic acid producing microbes
An ideal microorganism suitable for industrial or commercial production of lactic acid should have the following characteristics: 1. should be capable to use raw materials fully and rapidly 2. Should utilize minimum amount for nitrogenous substances and growth factors 3. Should yield maximum amount for required lactic acid stereo type 4.should be able to ferment at low PH and low temperature 5. Production of biomass and by-products should be very low (Narayanan et. al., 2004). Lactic acid producing microorganisms can be categorised as bacteria and fungi (Wee et. al., 2006). Lactic acid producing bacteria have been widely research than filamentous fungi as fungal fermentation has lower product yield and it produces by products like ethanol and fumaric acid (Tay et. al., 2002).
Rhizopus, are filamentous fungi capable of producing lactic acid through aerobic fermentation of glucose. Rhizopus oryzae and Rhizopus arrhizus have the ability to convert starch directly to L(+) lactic acid eliminating the need for pre-treatment of raw material due to its amlolytic enzyme activity (Oda et. al., 2002). The medium requirements for fungal fermentation are relatively simple but it requires extensive aeration which may increase cost of production. Many researchers have attempted to maximize productivity and yield of lactic acid using fungal fermentation, but no significant results were obtained. Tay and Yang used fibrous bed immobilized Rhizopus oryzae to enhance production of lactic acid form starch (Tay et. al., 2002). Park et. al. used Rhizopus oryzae NRRL 395 strain in an air lift bioreactor and obtained lactic acid yield of 104.6 g/L. They also suggested that mycelial floc formation improved lactic acid formation (Park et. al., 2008). Despite of all attempts made to improve production of lactic acid; bacterial fermentative production of lactic acid has an upper hand and is used to produce lactic acid on an industrial scale. Porro et. al. studied production of L(+) lactic acid from, Saccharomyces cerevisiae and Kluyveromyces lactis, for their desirable property to tolerate high concentration of hydrogen ions (refer page 171, (Narayanan et. al., 2004) .
As mentioned earlier lactic acid producing bacteria are divided into homo-fermentative and hetero-fermentative. Homo-fermentative lactic acid bacteria give better yield of lactic acid per molecule of glucose, which is more than 0.90 g/g and have no major by products associated. For this reason homo-fermentative bacteria are used for commercial production of lactic acid (Wee et. al., 2006). As different lactic acid bacteria have specificity for different substrates thus an organism should be selected based on carbohydrate to be fermented (Narayanan et. al., 2004). Table 1 lists lactic acid producing bacteria and the carbohydrate they can utilize. In addition to carbon and nitrogen sources lactic acid bacteria also require growth factors as they cannot synthesize amino acids, vitamin B and minerals (Wee et. al., 2006).
Table 1: Carbohydrate utilization by lactic acid bacteria (made from review by Narayanan et. al.)
Lactic acid bacteria
Lactose and galactose
Sulphite waste liquor
Commercially used strains of microorganisms are proprietary and much information is not available on them. Most of the current investigations involve use of Lactobacillus for production of lactic acid. By using the strain Lactobacillus delbrueckii NCIMB 8130 Kotzanmanidis et. al. produced lactic acid from beet molasses, they produced 90 g/L of lactic acid with yield of 0.97 g/g and productivity of 3.8 g/(L.h) (Kotzanmanidis et. al., 2002). Robel et. al. performed coculturing of Lactococcus lactis ssp. lactis cells and Aspergillus awamori to produce lactic acid from cassava starch. They produced 90 g/L of lactic acid with yield of 0.76 g/g and with slightly low productivity of 1.6 g/(L.h) (Robel et. al., 2003). Similarly, Hujanen and Linko obtained high of 0.91 g/g with 82 g/L of lactic acid production. They used Lactobacillus casei NRRL B-441 strain in tested effects of different temperature and nitrogen sources (Hujanen and Linko, 1996). Various different studies have been carried out to study lactic acid production using Lactobacillus rhamnosus, Lactobacillus helveticus, Lactobacillus bulgaricus, Lactobacillus plantaru and Lactobacillus pentosus (Wee et. al., 2006).
2.2 Genetically Engineered Microbes
Many researchers have genetically modified to improve or induce production of L(+) lactic acid and D(-) lactic acid.
Lactobacillus helveticus (Kylä-Nikkilä et. al., 2000)
Kylä-Nikkilä et. al. used Lactobacillus helveticus CNRZ32 strain and attempted to increase production on L(+) lactic acid. They constructed two strains negative for ldhD (lactate dehydrogenase D gene) expression. In one strain ldhD was replaced by ldhL and in the second strain the promoter for ldhD was deleted thus blocking the transcription. Their results show that there was 2 fold increase in production of L(+) lactic acid and it was equivalent to the total lactic acid produced by wild type strain.
Lactobacillus plantarum (Ferain et. al., 1994)
They cloned the lactate dehydrogenase L gene form Lactobacillus plantarum and cloned it into E.coli. After sequencing this gene Ferain et. al. strains of Lactobacillus plantarum were constructed either over expressing or lacking expression of ldhL gene. This resulted in 13 fold increase in expression of gene but had negligible effect on L(+) lactic acid production.
Chang et. al. produced recombinant strain of E.coli by introducing ldhL gene from Lactobacillus casei into E.coli mutant lacking phosphotransacetylase and ldhD gene. They also induced IdhD production by introducing ldhD gene. They were successful in showing that E.coli can be genetically modified to produce optically pure L(+) or D(-) Lactic acid. Dien et. al. genetically modified E.coli to produce L(+) lactic acid from hexose sugar and also pentose sugar (Narayanan et. al., 2004 and Wee et. al., 2006). Jung et. al. constructed a metabolically engineered strain of E.coli with improved propionate CoA-transferase PHA synthase activity. They successfully produces PLA and P(3HB-co-LA) using glucose in single step without use of any external inducer. The strain of E.coli developed in this study was E. coli JLXF5 (Jung et. al., 2011).
A mutant with minimum (5%) alcohol dehydrogenase activity produced lactic acid from pyruvate in oxygen limiting condition. Normally the fungus grows only for short period in absence of oxygen due to presence alcohol dehydrogenase (Narayanan et. al., 2004).
Sweet sorghum, Wheat, Corn, Cassava, Potato, Rice, Rye, barley
Corn cob & stover, Waste paper, Wood, Alfalfa fibre, Wheat barn & straw
Whey and Molasses
Figure 4: Classification of raw materials used for lactic acid production (made from review by Wee et. al., 2006)
2.3 Raw materials
Different raw materials used in production of lactic acid have been classified in figure 4. Generally starchy materials, cellulosic materials and industrial waste products are used as raw materials for production of lactic acid by microbial fermentations. Starchy and cellulosic materials have intrigued researchers recently as they are affordable, present in large quantities in the environment and most importantly they are easily renewable. These characteristics are vital for commercial producers of lactic acid and PLA.
Production of lactic acid from starchy materials requires a hydrolysis step to convert them into fermentable sugars as glucose is linked by Î±-(1,4) and Î±-(1,6) bonds because all microbes cannot hydrolyse these bonds. Lactobacillus amylophylus, Lactobacillus amylovirus, Rhizopus oryzae and Rhizopus arrhizus, have amlolytic properties hence they can directly convert starchy materials into lactic acid (Wee et. al., 2006). Similar to starchy materials cellulosic material need a pre-treatment to convert lignocellulose to cellulose. During the pre-treatment step several compounds are generated which inhibit the hydrolysis of lignocellulose; these are acetic acid, furfural etc. Wee et. al. suggested that this effect can be reduced to an certain extent by direct adaptation of lactic acid bacteria to wood hydrolysate medium (Wee et. al., 2006). Industrial waste products like molasses and whey contain sugars which act as substrate for lactic acid bacteria. Table 2 lists raw materials and microorganisms used to produce lactic acid.
3 Polylactic acid production
Lactic acid is polymerized to produce PLA. The PLA industry requires lactic acid which has high optical purity with L(+) lactic acid concentration more than 98 to 99 %. There are two widely used ways to produce PLA from lactic acid. First one is direct polycondensation of lactic acid and the other being via formation of lactide (cyclic dimer of lactic acid) followed by ring opening reaction of lactide. Both these processes are commercially used to produce PLA (Jem et. al., 2010).
Table 2: Production of lactic acid form cheap raw materials (Narayanan et. al., 2004 and Wee et. al., 2006)
Lactic acid yield (g/L)
Wheat and rice barn
Lactobacillus casei NRRL B-441
Lactobacillus paracasei No. 8
84.5 - 81.5
Wood and pretreated wood
48 - 108
Rhizopus sp. MK-96-1196
24 - 90
Rhizopus oryzae NRRL 395
Industrial waste material
Lactobacillus delbrueckii NCIMB 8130
Lactobacillus helveticus R211
The direct polycondensation process involves formation of oligomers by dehydrating lactic acid which is then followed by polymerization of the oligomers to obtain high molecular weight PLA by simultaneous dehydration. The major drawback of this method is removal of moisture from the molten viscous polymer. The trapping of water molecule can result in low molecular weight PLA. Hence this process is not used widely. Mitsui Toatsu chemicals is one of the few companies that uses direct condensation and organic solvent based azeotropic distillation to remove water continuously (Ajioka et. al., 1995).
The most commonly used method to obtain high molecular weight PLA is known as ring opening polymerization (ROP) of lactide. The first step in this process is polycondensation of lactic acid to form oligomers. This is followed by depolymeriztion to remove water, which results in formation cyclic dimer of lactic acid called lactide (Narayanan et. al., 2004) . The lactide in then polymerised my ring opening polymerization into polyester. The ROP process is performed at temperature more than melting temperature of lactide but below degradation temperature (Jem et. al., 2010). ROP process is catalysed by stannous octoate catalyst. Cargill Dow LLC has patented processes of ROP in which synthesis of PLA is performed in melt and ROP is catalysed by tin catalyst which is environment friendly (Narayanan et. al., 2004).