Description:TECHNICAL FIELD:
The present disclosure generally relates to the field of bioengineering technology and relates to a separation technology of a microbial fermentation liquid, in particular to a method for coupling and separating lactic acid in a fermentation liquid by using an extraction technique.

BACKGROUND:
Over the past few years, there has been an increasing interest shown in the development of bio-based, ‘green’, platform chemicals. A major enabler of their popularity has been the availability of low cost and sustainable feedstocks derived from renewable lignocellulosic sources. The growing awareness and consumer interest in bio-based products, ongoing technological developments, and exposure to a stable feedstock price are some of the added advantages of green products, aside from a reduced carbon footprint. There are ten major categories of bio-based chemicals such as platform chemicals, solvents, polymers for plastics, coating and dyes, surfactants, cosmetics and personal care products, adhesives, lubricants, plasticisers, manmade fibers based on applications, among them platform chemicals have a lot of market. Recent report by "Market Research Future" concluded that bio-based platform chemicals are projected to grow at a compound annual growth rate (CAGR) of 12.5% by 2027.

L-Lactic Acid (LLA) is considered as one of the top second-generation platform chemical, which has a technology readiness level (TRL) of eight or higher. Its chemical name is 2-hydroxy-propionic acid, which has a wide array of applications in food, pharmaceuticals, cosmetics, specialty chemicals, textiles, leather, and biodegradable plastics and polymers. Chemical synthesis and fermentation are two routes to produce commercial lactic acid. The fermentation process has an advantage over the chemical process as it produces optically pure LLA, while the latter makes a racemic mixture of DL-lactic acid. Lactic acid has applications in food, pharma, bioplastic and chemical industries. Unlike conventional polymers such as polyethylene (PE), polypropylene (PP), polyethylene terephthalate (PET) and polystyrene (PS), Poly-L-lactic acid (PLLA) is a linear aliphatic thermoplastic polyester made from renewable resources and readily biodegradable.

LLA purification is an expensive operation, with costs ranging from 0.78 to 1.74 USD per kilogram of lactic acid. The downstream processing of lactic acid might cost about 40–70% of the total price, therefore; an easy and cost-effective downstream method for lactic acid purification is essentially required. Due to the presence of media components, salts, and residual sugars in the fermentation broth, a number of methods for the separation and purification of lactic acid from the fermentation broth have been proposed in the literature. Ultra/nanofiltration, electrodialysis, ion-exchange/adsorption, reactive distillation, and hybrid short path evaporation appeared to be the most commonly known methods thus far. Although, the complexity, high cost of used materials and fouling issues, limited the lactic acid recovery process.

EP2017347A1 discloses a method in which lactic acid component (e.g., lactic acid or oligo (lactic acid)) can be obtained by extraction from lactic acid fermentation liquor with a pH of 4.8 or less, using at least one solvent selected from the group consisting of toluene, xylene, mesitylene, ethylbenzene and mineral spirit.

US6229046B1 discloses a technique for processing mixtures of lactic acid and dissolved lactate salts. The preferred techniques were provided for processing fermentation broths, preferably fermentation broths produced with or adjusted to have a pH of less than about 4.8, typically and preferably less than about 4.5, more preferably less than 4.3 and most preferably within the range of about 3.0 to 4.2 inclusive. The techniques concern processing the mixtures into: (a) a lactic acid stream, component or phase; and, (b) a lactate salt stream component or phase. Preferred techniques were provided so that the lactic acid stream, component or phase can be readily taken on to produce desirable lactate products, such as lactate oligomers, lactate esters, lactate amides and/or polylactate.

US9012685B2 discloses a method for recovery of highly pure alkyl lactate and lactic acid, which includes a step 1 for producing source liquid comprising lactic acid or ammonium lactate; a step 2 for dehydrating the source liquid product of step 1; a step 3 for producing liquid mixture by sequentially adding and stirring alcohol and acid solution to the dehydrated source liquid; a step 4 for separating and removing ammonium salt precipitation from the liquid mixture of step 3; a step 5 for producing alkyl lactate from ammonium salt-free liquid mixture by esterification reaction; and a step 6 for separating alcohol and alkyl lactate by distillation from the mixture of step 5.

WO2001092555A1 discloses a method for producing lactic acid, comprising producing lactic acid from a sugar-containing fermentation liquid in a fermentor by means of lactic acid-forming bacteria to result in a lactate salt, and isolating lactic acid by subjecting the fermented fermentation liquid to a first ultrafiltration step to result in a substantially polymer-free permeate comprising at least one lactate salt, acidifying the permeate to a pH value of below about 3.9, and performing at least one additional isolation step in which the acidified permeate is subjected to nanofiltration and/or reverse osmosis. Finally, inorganic salts are typically removed by electrodialysis.

US8293940B2 discloses a process for recovery and purification of lactic acid from a fermentation broth containing lactic acid. The process comprises subjecting the fermentation broth to ultrafiltration and/or microfiltration to form a first permeate, concentrating the first permeate to form concentrated broth, subjecting the concentrated broth to supported liquid membrane for extraction of lactic acid into a separate stream, subjecting the extracted lactic acid solution to activated carbon for removal of color, subjecting the extracted lactic acid solution to cation exchange resin for deminerization, subjecting the extracted lactic acid solution to anion exchange resin for removal of anionic impurities and concentrating the extracted lactic acid solution to desired concentration. The supported liquid membrane of the present invention contains an organic layer that comprises an earner, a co-extractant, a diluent and a stabilizer.

Thus, there is a need in the art to provide a lactic acid recovery process which can yield high purity lactic acid and the process should have high efficient recovery

OBJECTIVE:
An objective of the present disclosure is to provide a process to recover L-lactic acid from a filtrate from a bioreactor.

Further an objective of the present disclosure is to provide a process with highly efficient recovery of L-Lactic acid.

Another objective of the present disclosure is to provide a process which recycles used organic solvents and reactive extractant to make the process economical feasible.

SUMMARY:
The present disclosure relates to a process to recover L-lactic acid from a filtrate from a bioreactor, wherein the process comprising: maintaining a pH of the filtrate in a range of 2.5 to 5; adding a mixture of an organic solvents in a range of 50 to 100 v/v% with respect to filtrate and an amine extractant into the filtrate, followed by homogenization at a speed of at least 2500 rpm for at least 10 minutes to obtain a mixture; obtaining an organic phase and an aqueous phase from the mixture; vaporizing the organic phase at a temperature in a range of 45 to 55°C to recover L-lactic acid, wherein the organic solvent is selected from the group consisting of iso-propanol, n-pentanol and n-butanol or combination thereof and the amine extractant is selected from the group consisting of trihexylamine and Bis(2-ethylhexyl)amine or combination thereof.

These and other features, aspects, and advantages of the present subject matter will become 10 better understood with reference to the following description. This summary is provided to introduce a selection of concepts in a simplified form. This summary is not intended to identify key features or essential features of the claimed subject matter, nor is it intended to be used to limit the scope of the claimed subject matter.

DESCRIPTION OF THE ACCOMPANYING DRAWINGS:
These and other features, aspects, and advantages of the present disclosure will become better understood when the following detailed description is read with reference to the accompanying drawings in which like characters represent like parts throughout the drawings, wherein:

Figure 1 illustrates schematic representation showing overall down streaming process by phase partioning method with homogenization and reactive extraction for LLA.

DETAILED DESCRIPTION:
The present disclosure addresses the drawbacks of the art and provides for a process for recovering L-lactic acid from a filtrate from a bioreactor. The invention relates to an extraction method for extracting L-Lactic acid from the fermentation broths using a combination of Bis(2-ethylhexyl)amine based reactive extractant with a synergistic mixture of cheap organic solvents. In some embodiments, the process provides an economical process to recover lactic acid with high purity and high efficiency.

Although any methods and materials similar or equivalent to those described herein can also be used in the practice or testing of the present disclosure, the preferred methods and materials are now described. All publications mentioned herein are incorporated herein by reference to disclose and describe the methods and/or materials in connection with which the publications are cited.

It must be noted that as used herein and in the appended claims, the singular forms "a", "and", and "the" include plural referents unless the context dictates otherwise. Thus, for example, reference to "a compound" includes a plurality of such compounds, and reference to "the step" includes reference to one or more steps and equivalents thereof known to those skilled in the art, and so forth.

The term “some” as used herein is defined as “none, or one, or more than one, or all.” Accordingly, the terms “none,” “one,” “more than one,” “more than one, but not all” or “all” would all fall under the definition of “some.” The term “some embodiments” may refer to no embodiments or to one embodiment or to several embodiments or to all embodiments. Accordingly, the term “some embodiments” is defined as meaning “no embodiment, or one embodiment, or more than one embodiment, or all embodiments.”

The terminology and structure employed herein is for describing, teaching and illuminating some embodiments and their specific features and elements and does not limit, restrict or reduce the spirit and scope of the claims or their equivalents.

More specifically, any terms used herein such as but not limited to “includes”, “comprises”, “has”, “consists” and grammatical variants thereof is inclusive or open-ended and does not exclude additional, unrecited elements or method steps. The specification will be understood to also include embodiments which have the transitional phrase “consisting of” or “consisting essentially of” in place of the transitional phrase “comprising.” The transitional phrase “consisting of” excludes any element, step, or ingredient not specified in the claim, except for impurities associated therewith. The transitional phrase “consisting essentially of” limits the scope of a claim to the specified materials or steps “and those that do not materially affect the basic and novel characteristic(s)” of the claimed disclosure.

Whether or not a certain feature or element was limited to being used only once, either way it may still be referred to as “one or more features” or “one or more elements” or “at least one feature” or “at least one element.” Furthermore, the use of the terms “one or more” or “at least one” feature or element do NOT preclude there being none of that feature or element, unless otherwise specified by limiting language such as “there NEEDS to be one or more. ” or “one or more element is REQUIRED.”

As used herein, the term “about” is used to indicate a degree of variation or tolerance in a numerical or quantitative value. It indicates that the disclosed value is not intended to be strictly limiting, and may vary by plus or minus 5%, without departing from the scope of the invention.
Unless otherwise defined, all terms, and especially any technical and/or scientific terms, used herein may be taken to have the same meaning as commonly understood by one having an ordinary skill in the art.

Reference is made herein to some “embodiments.” It should be understood that an embodiment is an example of a possible implementation of any features and/or elements presented in the attached claims. Some embodiments have been described for the purpose of illuminating one or more of the potential ways in which the specific features and/or elements of the attached claims fulfil the requirements of uniqueness, utility and non-obviousness.

Use of the phrases and/or terms such as but not limited to “a first embodiment,” “a further embodiment,” “an alternate embodiment,” “one embodiment,” “an embodiment,” “multiple embodiments,” “some embodiments,” “other embodiments,” “further embodiment”, “furthermore embodiment”, “additional embodiment” or variants thereof do NOT necessarily refer to the same embodiments. Unless otherwise specified, one or more particular features and/or elements described in connection with one or more embodiments may be found in one embodiment, or may be found in more than one embodiment, or may be found in all embodiments, or may be found in no embodiments. Although one or more features and/or elements may be described herein in the context of only a single embodiment, or alternatively in the context of more than one embodiment, or further alternatively in the context of all embodiments, the features and/or elements may instead be provided separately or in any appropriate combination or not at all. Conversely, any features and/or elements described in the context of separate embodiments may alternatively be realized as existing together in the context of a single embodiment.

As used herein, the term "bioreactor" or “fermentor” refers to a vessel in which a chemical process is carried out which involves organisms or biochemically active substances derived from such organisms.
As used herein, the term “homogenize” refers to distributing a substance uniformly throughout a fluid.

As used herein the terms “method” and “process” have been used interchangeably.

The present disclosure discloses a process for recovering L-lactic acid from a filtrate from a bioreactor.

In some embodiments, the process comprises adding a mixture of an organic solvent and an amine extractant into the filtrate, followed by homogenization.

In some embodiments, a process to recover L-lactic acid from a filtrate from a bioreactor, wherein the process comprising: maintaining a pH of the filtrate in a range of 2.5 to 5; adding a mixture of an organic solvent in a range of 50 to 100 v/v% with respect to filtrate and an amine extractant into the filtrate, followed by homogenization at a speed of at least 2500 rpm for at least 10 minutes to obtain a mixture; obtaining an organic phase and an aqueous phase from the mixture; vaporizing the organic phase at a temperature in a range of 45 to 55°C to recover L-lactic acid, wherein the organic solvent is selected from the group consisting of iso-propanol, n-pentanol and n-butanol or combination thereof and the amine extractant is selected from the group consisting of trihexylamine and Bis(2-ethylhexyl)amine or combination thereof.

In a preferred embodiment, there is provided the process, wherein the pH of the filtrate is in a range of 3.0 to 4.0.

In some embodiments, there is provided the process, wherein the L- lactic acid was produced in bioreactor using Lactobacillus Sp.. The Lactobacillus Sp. is selected from Lactobacillus helveticus, Lactobacillus amylophilus, and Lactobacillus delbrueckii.

In some embodiments, there is provided the process, wherein the organic phase and the aqueous phase is obtained by allowing the mixture to stand-still for the time period in a range of 30 to 45 minutes.

In some embodiments, there is provided the process, wherein the organic solvent is a mixture of iso-propanol and n-pentanol or a mixture of iso-propanol and n-butanol.

In a preferred embodiment, the organic solvent is a mixture of iso-propanol and n-pentanol.

In some embodiments, there is provided the process, wherein iso-propanol is present in a range of 50 to 100v/v% and n-pentanol is present in a range of 50 to 100v/v%. In some embodiments, iso-propanol is 50v/v% and n-pentanol 50 v/v%. In some embodiments, iso-propanol is 100 v/v% and n-pentanol is absent. In some embodiments, n-pentanol is 100 v/v% and iso-propanol is absent.

In some embodiments, there is provided the process, wherein the amine extractant is 0.1-0.5 M Bis(2-ethylhexyl)amine.

In some embodiments, there is provided the process, wherein the organic solvent is in range of 50 to 100 v/v% and the amine extractant is in range of 1 to 20 wt/vol%. In a preferred embodiment, the amine extractant is in range of 2-15 wt/vol%. In 50 ml volume, 1.20 g of amine extractant is added to make the 0.1 M and 6.03 g of amine extractant is added to make the 0.5 M concentration.

In some embodiments, there is provided the process, wherein the homogenization speed is in a range of 2000 to 5000 rpm.

In some embodiments, there is provided the process, wherein the homogenization time is in a range of 5 to 30 minutes.

In some embodiments, there is provided the process, wherein the recovery of L-lactic acid is 92% and the purity of the L-lactic acid is 98%.

In an exemplary and non-limiting embodiment, advantages of the present invention over processes known in prior art includes the recovery of L-Lactic acid (LLA) from the fermentation broths using a Bis(2-ethylhexyl)amine based reactive extractant with a synergistic mixture of cheap organic solvents via phase partitioning method and homogenization. At optimized process conditions 92% lactic acid is recovered with 98% purity as confirmed by 1H-NMR. Recycling of amines extractant and organic solvents is additional advantage of this process for its application at commercial scale.

The present disclosure is further illustrated by reference to the following examples which is for illustrative purpose only and does not limit the scope of the disclosure in any way. These examples are not intended to be inclusive of all aspects of the subject matter disclosed herein, but rather to illustrate representative features, methods, compositions, and results. These examples are not intended to exclude equivalents and variations of the present disclosure, which are apparent to one skilled in the art.

EXAMPLES:
Source of Biological Material: The biological material Lactobacillus delbrueckii LD2365 was obtained from National Collection of Industrial Microorganisms (NCIM), Pune, India.

Example 1: Microorganism cultivation and growth conditions
The LLA was produced by Lactobacillus delbrueckii LD2365 was obtained from National Collection of Industrial Microorganisms (NCIM), Pune, India. The culture was routinely maintained on DeMan, Rogosa, and Sharpe (MRS) agar slants at 4?C and was sub cultured fortnightly. For lactic acid production in bioreactor, the freshly grown mother culture (O.D. ˜ 0.7) was used to inoculate the synthetic medium comprise yeast extract 10 gL-1, di-ammonium hydrogen phosphate 0.1 gL-1, calcium carbonate 4.5 gL-1, Dextrose (120 gL-1) at a 10% (v/v) inoculum level. The inoculated broth was incubated at 40-42? at 150 rpm for 48-72 hours. The cells were harvested by centrifugation (10,000 rpm, 10 min) and the fermentation broth was treated with concentrated sulfuric acid to convert calcium lactate to free LLA. The generated gypsum was separated by repeating the centrifugation step and the solution was clarified using activated carbon in order to remove all the organic impurities. For this activated carbon (20 gL-1) was added in gypsum free LLA solution and vigorously vortexed for proper mixing before keeping it shaking for 2 hours at 40?. The next step involved filtration in order to remove the activated carbon and the filtrate was used for later step.

Example 2: Downstream processing of lactic acid
A) Screening of diluents
Different diluents such as acetone, ethanol, iso-propanol, 1-butanol, 1-pentanol and their combinations (1:1) were screened to identify the best diluent or their combination which facilitated the LLA extraction. For this equal amount of LLA rich aqueous phase and organic solvent was added and allowed to homogenize the solution at 2000 rpm for 5 minutes. The mixture was then transferred to separating funnel and allowed to stand till organic and aqueous phase to reach the distribution equilibrium. After 30 minutes, the upper organic phase was separated from the aqueous phase. Thereafter, the organic phase was vaporized at 50?. The obtained LLA was dissolved in 1 ml water and quantified on the Water HPLC system equipped with Aminex HPX-87H ion exclusion column (7.8 × 300 mm) and refractive index detector (RID) using 5 mM H2SO4 as a mobile phase at a flow rate of 0.6 mL minute-1. The extraction efficiency was evaluated by the distribution coefficient (KD) of LLA in the organic phase [LA]org and in aqueous phase [LA]aq after sufficient mixing (Equation 2). The extraction yield was defined by Equation 3.

When different solvents, including polar aprotic solvents, polar, non-polar alone or their combinations were tested, iso-propanol combination with n-butanol and n-pentanol emerged as the best organic solvents, as shown in Table 1. In presence of iso-propanol alone the extraction efficiency was 38 ± 2%. However, its combination with n-butanol and n-pentanol efficiency enhanced significantly to 63 ± 4% and 64 ± 3% respectively. The order of organic solvents which showed high compatibility for LLA are as follows: n-pentanol + iso-propanol > n-butanol + iso-propanol > n-pentanol > n-butanol > iso-propanol > ethanol > acetone. LLA has a good solubility in lower alcohols, the medium components contained in the lactic acid fermentation liquor and the agents for use in neutralization or acidification have a poor solubility in these solvents. The solvent is recycled for the next round of LLA purification for example, in the case of a standard medium M17 for lactic acid fermentation, 100% of the medium components are dissolved in water, but 33.8% of the medium components are soluble in methanol, 5.1% in ethanol, 3.8% in isopropanol, and 4.5% in butanol, that is, the proportion of those that are soluble in these alcohols is small. Furthermore, lactic acid solubility in iso-propanol is higher but if they use alone, lactic acid in the aqueous phase was fractionally transferred into the organic phase, and vice versa leading to incorporation of both phases and, in turn incomplete solvent recovery and complicated lactic acid purification in subsequent steps. In contrast to this, lactic acid solubility in long chain aliphatic alcohols namely butanol and pentanol is low but they helpful to make distinct large organic to aqueous ratio because of low solubility in water. Therefore, Iso-propanol, in combination with n-pentanol, likely facilitated better extraction of lactic acid. However, it was essential to improvise the LLA extraction efficiency further. Therefore, optimization of pH, homogenization speed, time and reactive amine concentration were looked upon, which can synergistically enhance the distribution of lactic acid in the organic phase by reducing water availability for its dissolution.
Table 1: Effect of organic solvents alone or in combination on the extraction efficiency of LLA.

B) Optimization of physicochemical parameters
Effect of various physicochemical parameters such as the effect of initial pH (3.0, 3.5, 4.0, 4.5, 5.0) of the cell free supernatant, homogenization speed (1000 - 5000 rpm) and time (5, 10, 15, 20, 30 minute) was studied using one variable at a time (OVAT) approach. Therefore, we screened the two different amines in combinations with optimized diluents and they were experimentally evaluated for reactive extraction of LA. In comparison to only diluents, reactive extractants based on amine-diluent and homogenization combinations demonstrated higher extractive yields. Extractants with the highest extractive capacity were selected for further research aimed to optimize the concentration of amines for maximum extraction yield. Thereafter each component (acid/diluent) from the organic phase is separated by fractional distillation and selected solvent was recycled. The process is very suitable to remove any color soluble tar, which would otherwise form a problem in subsequent process step, particularly in distillation. Overall, the process is simple, cost effective, economically feasible and results in higher yields of lactic acid. The complete fractionation scheme is shown in figure 1.

B.1) Effect of pH
Partitioning of LLA was affected by the pH of the medium. At lower pH lesser amount of lactic acid was retained in the aqueous phase, and better recovery was obtained in the organic phase. The best recovery of lactic acid (70 ± 3.0%) was achieved in medium with pH 3.0 (Table 2). The pH significantly influenced lactic acid extraction by the solvent because of its ionization behavior. LLA is the primary form below its pKa of 3.86, and at pH above it, lactate exists predominantly in solution. A comparison between the molecular polarities of lactate ion and lactic acid shows that lactic acid has a lower polarity than lactate ion, and thus can be more easily dissolved in a solvent other than water at lower pH. LLA extraction by the organic solvents is best reported around a pH of 3.0-4.0.
Table 2: Effect of pH on the extraction efficiency of LLA
pH Extraction yield (%)
3.0 70 ± 3.0
3.5 61 ± 4.0
4.0 56 ± 5.0
4.5 45 ± 3.0
5.0 40 ± 2.0

B.2) Effect of homogenization speed and time
The effect of homogenization speed and time on the lactic acid extraction was given in Table 3. It was observed that yield of lactic acid increased (74.0 ± 4.0%) with increasing homogenization speed until 3000 rpm, and then a stagnant. Increasing homogenization speed leads to higher mechanical force, caviation and turbulence effect generate in the medium, and violent collapse of bubbles induces mass-transfer. This results in an increase in the dissolution of lactic acid. In order to explore the time to reach the distribution equilibrium of lactic acid between aqueous phase and organic solvent via homogenization process, the mixing systems were analyzed at different time points. From the trend in Table 3, the yield of lactic acid increased with increasing duration of homogenization treatment during the first 10 minutes (76.0 ± 3.0%) and then no more lactic acid was extracted in solvent. It can be concluded that the radiation pressure and cavitation effect generated by the homogenization can be introduced into liquid-liquid extraction system to transfer the matrix in extractant with good efficiency.
Table 3: Effect of homogenization speed and time on the extraction efficiency of LLA
Homogenization speed (rpm) Extraction yield (%)
1000 62 ± 2.0
2000 67 ± 5.0
3000 75 ± 2.0
4000 76 ± 3.0
5000 76 ± 4.0
Homogenization time (minutes) Extraction yield (%)
5 64 ± 2.0
10 76 ± 3.0
15 77 ± 2.0
20 78 ± 2.0
30 78 ± 3.0

B.3) Reactive extraction of LLA using amines with organic solvents/diluents
Reactive extraction involves a reversible reaction between the acid in the aqueous phase and the extractant in the organic phase (diluent phase) which results in the formation of extractant-acid complex having high affinity for the organic phase. Two amines extractants namely trihexylamine and Bis(2-ethylhexyl)amine were investigated in their increasing concentration with n-pentanol + iso-propanol diluent for the purification of LLA (Table 4). The resulted extraction yield (E%) are summarized in Table 4 for various amine-diluent systems.

As mentioned above, not only the nature of amine and its association with diluent but also the concentration of amine can influence the equilibrium extraction characteristics. The influence of varying amine concentration (0.1-0.5 M) on the reactive extraction of LA was investigated in Table 4. As shown in Table 4, Bis(2-ethylhexyl)amine is more effective in comparison to Trihexylamine for the extraction of LLA. These long chain aliphatic amines especially secondary amines are effective extractants for the separation of carboxylic acids from the dilute aqueous solution. The specific chemical interactions between the amines and the acid molecules to form acid-amine complexes in the extractant phase allow more acid to be extracted from the aqueous phase. These amines extractants are also dissolved in a diluent such as alcohols that is, an organic solvent that dilutes the extractant to the desired concentration and controls the viscosity and density of the solvent phase. An increase in extraction yield (E %) of LLA (92%) was observed with Bis(2-ethylhexyl)amine in their concentration of 0.2 M. Further increase in amine concentration did not help to improve the extraction yield (E%).

Table 4: Extraction yield (E%) of LLA at 25°C for selected organic solvents (n-pentanol + iso-propanol) in the presence of increasing concentration of different amines.

C) Characterization Proton Nuclear Magnetic Resonance
All Proton Nuclear Magnetic Resonance (1H-NMR) spectra were recorded either on a Agilent DD2 NMR spectrometer operating at a proton frequency of 500 MHz, spectral width of 8012.8 Hz (-2.0 to 14.0 ppm), 90° pulse = 11.8 µs, relaxation delay = 20 s, and digital resolution of 0.49 Hz/point or on a JEOL ECA-500 spectrometer operating at a proton frequency of 500 MHz and the same specified parameters as above (90° pulse = 10.7 µs). Sixteen repetitions were averaged with 32K data points and 6.24- and 6.38-min experimental time, respectively, for DD2 and ECA machines. All the NMR spectra were integrated after baseline correction, and a mean of minimum three integration values has been taken for each calculation.

NMR spectra showed that LLA obtained after phase extraction was of purified form as it matched with commercial lactic acid. It was characterized by a doublet centered at 1.3 ppm and a quartet at 4.3 ppm corresponding to the methyl (CH3) and carboxyl group (-COOH) protons of lactic acid, respectively. These results indicated that all other impurities were successfully removed so that the final product was obtained in high purity around >98%. The simple purification process and high-quality product further guaranteed the remarkable efficiency and potential of this unique method.

Advantages:
The process disclosed herein has following benefits over prior art:
1. Prior art is applicable to purify the LLA by utilizing ultra/nanofiltration, electrodialysis, ion-exchange/adsorption, reverse osmosis, there are concerns about the life and fouling of membranes whereas, the present invention disclosed a method to isolate high purity lactic acid (98%), using combination of readily available cost-effective organic solvents assisted with reactive extractant amine and homogenization.
2. The present invention does not require costly chemicals/reaction reagents, or consumables such as separation membranes.
3. The disclosed method is simple and easy to use involving with minimized steps for processing.
4. The cavitation and turbulence effect generated by homogenization is capable for efficient mixing of liquid-liquid blend and to adequately extract lactic acid in organic solvent from aqueous solution.
5. Lactic acid is preferentially extracted with the synergistically active mixture of solvents whereas; impurities were left behind in aqueous phase of fermentation liquor.
6. This process is very suitable to remove any color and impurities, which would otherwise form a problem in subsequent process step, particularly in vacuum distillation.
7. The process is very simple which leads to higher extraction yields of lactic acid (92%) which was easily separated by distillation.
8. Here, used solvents were recovered by distillation and reused many times without any substantial loss in the efficiency.
9. The used amine based extractant is recycled back to the reaction step multiple times, which reduce the additional use and improve the cost-efficiency.
10. None of the prior art explored the homogenization and Bis(2-ethylhexyl)amine based reactive extractant for the purification of lactic acid.
Although the subject matter has been described in considerable detail with reference to certain preferred embodiments thereof, other embodiments are possible. As such, the spirit and scope of the subject matter should not be limited to the description of the preferred embodiment 10 contained therein. , Claims:1. A process to recover L-lactic acid from a filtrate from a bioreactor, wherein the process comprising:
a. maintaining a pH of the filtrate in a range of 2.5 to 5;
b. adding a mixture of an organic solvents in a range of 50 to100 v/v% with respect to filtrate and an amine extractant into the filtrate, followed by homogenization at a speed of at least 2000 rpm for at least 5 minutes to obtain a mixture;
c. obtaining an organic phase and an aqueous phase from the mixture;
d. vaporizing the organic phase at a temperature in a range of 45 to 55°C to recover L-lactic acid,
wherein the organic solvent is selected from the group consisting of iso-propanol, n-pentanol and n-butanol or combination thereof and the amine extractant is selected from the group consisting of trihexylamine and Bis(2-ethylhexyl)amine or combination thereof.
2. The process as claimed in claim 1, wherein the L- lactic acid was produced in bioreactor using Lactobacillus Sp..
3. The process as claimed in claim 1, wherein the organic phase and the aqueous phase is obtained by allowing the mixture to stand-still for the time period in a range of 30 to 45 minutes.
4. The process as claimed in claim 1, wherein the pH of the filtrate is preferably in a range of 3.0 to 4.0.
5. The process as claimed in claim 1, wherein the organic solvent is a mixture of iso-propanol and n-pentanol or a mixture of iso-propanol and n-butanol.
6. The process as claimed in claim 1 and 4, wherein the organic solvent is a mixture of iso-propanol and n-pentanol.
7. The process as claimed in claim 6, wherein iso-propanol is present in a range of 50 to 100 v/v% and n-pentanol is present in a range of 50 to 100 v/v%.
8. The process as claimed in claim 1, wherein the amine extractant is 0.1 to 0.5 M Bis(2-ethylhexyl)amine.
9. The process as claimed in claim 1, wherein the organic solvent is in range of 50 to 100 v/v% and the amine extractant is in range of 1 to 20 (wt/vol)%.
10. The process as claimed in claim 1, wherein the homogenization speed is in a range of 2000 to 5000 rpm.
11. The process as claimed in claim 1, wherein the homogenization time is in a range of 5 to 30 minutes.
12. The process as claimed in claim 1, wherein the recovery of L-lactic acid is 92% and the purity of the L-lactic acid is 98%.