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Recent Development of Lactic Acid Production using Membrane

Bioreactors
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3rd MRS-ID meeting 2018 IOP Publishing
IOP Conf. Series: Materials Science and Engineering 622 (2019) 012023 doi:10.1088/1757-899X/622/1/012023

Recent Development of Lactic Acid Production using

Membrane Bioreactors

L. Aliwarga1, A.K. Wardani1, P.T.P. Aryanti2, and I G. Wenten1,3*

1
Chemical Engineering Department, Institut Teknologi Bandung, Jl. Ganesha 10,
Bandung, 40132, Indonesia
2
Chemical Engineering Department, Universitas Jenderal Achmad Yani, Jl. Terusan
Jendral Sudirman, Cimahi, 40285, Indonesia
3
Research Center for Nanosciences and Nanotechnology, Institut Teknologi Bandung,
Jl. Ganesha 10, Bandung, 40132, Indonesia

*Corresponding author: [email protected]

Abstract. Lactic acid has been widely used as flavour and preservative in the food,
pharmaceutical, leather and textile industries. It can be produced by fermentation process of the
substrates with high lactose content, such as cheese whey, soybean milk, corn, and potatoes.
Among various existing technologies, membrane bioreactor is one of the promising methods to
achieve high productivity of lactic acid. In addition, membrane bioreactor allows integration of
fermentation and separation steps, thus it able to simultaneously maintain high cell density,
recycle the cells for further use, and continuously remove lactic acid from the fermenter.

Keywords: lactic acid, membrane bioreactor, production yield

1. Introduction
Lactic acid (2-hydroxypropionic acid), CH3-CHOHCOOH, is a simple organic compound containing
both the hydroxyl and carboxylic acid groups. It was first isolated from sour milk by CW Scheele in
1780 and commercially produced by CE Avery in Littleton, MA, USA in 1881 [1-3]. The production
demand of lactic acid has been increased over years due to due to its high potential of application in a
wide range of fields [4-6]. In 2012, the production of lactic acid was around 259,000 metric tons with
the worldwide growth 12–15% per year [7, 8]. In recent years, lactic acid manufacture is mostly based
on carbohydrate fermentation. The major manufacturers are Archer Daniels Midland Company (USA),
NatureWorks LLC (USA), Purac (The Netherlands), and Galactic S.A. (Belgium).
Lactic acid has been mainly used for food and food-related applications. It is due to the mild acidic
taste of lactic acid. In addition, lactic acid is non-volatile, odourless, and classified as GRAS (generally
recognized as safe) for use as a general purpose food additive [2]. Therefore, many industries choose
lactic acid as a safety flavour and preservative in the food. Lactic acid also has been utilized in the
cosmetic industry such as in the manufacture of hygiene and aesthetic products due to its moisturizing,
antimicrobial and rejuvenating effects on the skin, as well as of oral hygiene products [8]. The other
promising application of lactic acid lies on its polymer, the poly-lactic acid (PLA). It offers tremendous

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3rd MRS-ID meeting 2018 IOP Publishing
IOP Conf. Series: Materials Science and Engineering 622 (2019) 012023 doi:10.1088/1757-899X/622/1/012023

advantages like biodegradability, thermos-plasticity, high strength etc. [9]. PLA is considered as an
environment-friendly alternative to substitute plastics derived from petrochemicals [10]. PLA can be
applied in medical applications for filling the gaps in bones, producing sutures (stitching material), and
joining membranes or thin skins in humans [11].
Another potential growth area for lactic acid derivatives is environmentally friendly solvents,
particularly lactate esters of low molecular weight alcohols such as ethyl, propyl and butyl lactate.
Oxygenated chemicals such as propylene glycol, propylene oxide, acrylic acid, acrylate esters, and other
chemical intermediates such as lactate ester plasticizers also can be made from lactic acid [2, 3]. The
schematic diagram of potential products from lactic acid derivatives can be seen in Figure 1.
The production of lactic acid has been dominated by carbohydrates fermentation processes. Various
studies aimed to improve lactic acid productivity using membrane bioreactors. In membrane bioreactor,
membrane module is integrated with conventional fermenters, thus permit simultaneous production and
purification of lactic acid in the same unit. Membrane bioreactor can be operated in various modes, such
as batch, semi-continuous, continuous, immobilized, and membrane recycle.
Membrane bioreactor offers advantages of great flexibility in scale of production depending on
market demand as well as high levels of separation and purification. In addition, membrane bioreactor
eliminates the requirement of separate purification units and results in compact design with reduced
capital investment [12]. For better understanding, this paper then aims to give a brief review of recent
development in lactic acid production using membrane bioreactors. The configuration and performance
of membrane bioreactors are discussed.

Figure 1. schematic diagram of potential products from lactic acid derivatives

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IOP Conf. Series: Materials Science and Engineering 622 (2019) 012023 doi:10.1088/1757-899X/622/1/012023

2. Experimental
In general, lactic acid can be manufactured by either chemical synthesis or carbohydrates fermentation.
The chemical synthesis is mainly based on the hydrolysis of lacto nitrile by a strong acid and produces
only a racemic mixture of lactic acid [13]. Meanwhile, a fermentation process is able to produce a
stereoisomer of lactic acid [7]. Fermentation process is more attractive in terms of its environmental
impact, low production cost, decreased fossil-based feedstock dependency, reduction of CO2 emission,
and high product specificity [14]. Therefore, fermentation process is preferred for the production of
lactic acid, approximately 90% from the total lactic acid production worldwide [13, 15].

Table 1. Comparison of different strains and substrates for lactic acid production

Substrate Organism Yield (g/g) Ref.

Lactobacillus casei 0.91 [16]
Lactobacillus rhamnosus 0.74-0.93 [16-18]
Lactobacillus delbrueckii 0.57-0.83 [19]
Glucose
Lactobacillus salivarius 0.92 [18]
Lactobacillus zeae 0.71-0.98 [17, 20]
Lactobacillus coryniformis 0.98 [20]
Lactobacillus rhamnosus 0.38 [21]
Lactose
Lactococcus Lactis 1.50 [21]
Lactobacillus delbrueckii 0.45-0.58 [22]
Molasses
Lactobacillus rhamnosus 0.40 [22]
Lactobacillus amylovorus 0.52 [23]
Lactobacillus casei 0.67 [23]
Hydrolized barley flour
Lactobacillus delbrueckii 0.85 [24]
Lactobacillus rhamnosus 0.74 [24]
Lactococcus Lactis 0.76 [25]
Hydrolized wheat flour
Lactobacillus delbrueckii 0.11-0.82 [25]
Lactobacillus kefir 0.20 [26]
Paneer whey
Lactobacillus acidophilus 0.17 [26]
Lactobacillus casei 0.32 [27]
Whey
Leuconostoc Lactis 0.22 [27]
Streptococcus Thermophilus 0.35-0.50 [28-30]
Lactobacillus delbrueckii 0.18-0.41 [28-30]
Whey permeate
Lactococcus Lactis 0.20-0.88 [28, 29, 31]
Lactobacillus rhamnosus 0.71 [31]
Lactobacillus plantarum 0.42-0.46 [32]
Solid waste Lactobacillus pentosus 0.43-0.51 [32]
Lactococcus Lactis 0.16 [32]

2.1. Carbohydrate Resources

Lactic acid can be produced from various substrates, either sugar in pure form such as glucose, sucrose,
lactose etc. or sugar-containing materials such as whole-wheat powder [33], starch [13, 34], cucumber
juice [35], cheese whey [36, 37], molasses [38], and sugarcane juice [39]. The selection of carbohydrate
feedstock depends on the price, availability and its purity. The purified sugars are mostly expensive as

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IOP Conf. Series: Materials Science and Engineering 622 (2019) 012023 doi:10.1088/1757-899X/622/1/012023

the feedstock for lactic acid production. Therefore, many researchers utilized agricultural by-products
such as corn starch, cassava starch, cottonseed hulls, wheat bran, sugarcane press mud, barley starch,
carrot processing waste, corn fiber hydrolyzates, and potato starch as potential substrates for lactic acid
production [9]. Meanwhile, proteinaceous and other complex nutrients that required by the organisms
can be provided from corn steep liquor, yeast extract, soy hydrolysate, etc. [3, 39, 40].

2.2. Microorganisms
The characteristic of the produced lactic acid depends much on the type of organism. In general,
microorganisms for lactic acid production can be classified into homofermentative strain and
heterofermentative strain. A homofermentative strain produces a single product, lactic acid only, while
heterofermentative strain produces other products such as ethanol, diacetyl, formate, acetoin or acetic
acid, and carbon dioxide along with lactic acid [9]. The comparisons of homofermentative strain and
heterofermentative strain organisms for lactic acid production are shown in Table 2. The existing
commercial lactic acid production processes mainly use homofermentative organisms, especially from
the genera Lactobacillus, Streptococcus, Enterococcus, and Pediococcus. These organisms exhibit
maximum productivity only within a very narrow pH range, preferably in the range of 5.5 to 6.5 [41].
The choice of microorganisms is mainly determined by the resource of carbohydrate. For substrate
from glucose, any homofermentative strains of the genus Lactobacillus can be used, however the
preferred organism is Lactobacillus delbrueckii [42]. For lignocellulosic substrates, Lactobacillus
pentosus is required to maximize the yield [32]. Meanwhile, starch is able to be both hydrolyzed and
fermented by certain amylase-producing Lactobacillus strains, such as Lactobacillus amylovorus [43].
Lactobacillus bulgaricus is the most used organism for fermentation of whey. This organism is able to
produce lactic acid with yields in the range of 80-90% [44, 45]. Besides, fungal strains such as Mucor,
Monilia, and Rhizopus can also be utilized to produce lactic acid. The best-known fungal source of lactic
acid is Rhizopus oryzae that able to obtain 63–69% yields of lactic acid from chemically defined media
containing 15% glucose [46, 47].

2.3. Methods
Lactic acid is a metabolic product of simple carbohydrates produced by many species of organisms
mainly through the fermentative metabolic pathway. After supplementation of nutrients, sugar contained
substrates are inoculated with the selected microorganism, and the fermentation takes place [8]. The
stoichiometry for homofermentative fermentation of lactic acid from hexose and pentose can be
expressed as equation (1) and (2), respectively [48].

C6H12O6  2 C3H6O3 (1)

C5H10O5  C3H6O3 + C2H4O2 (2)

The carbohydrate fermentation operates most efficiently and effectively at near neutral pH, which
requires neutralization and produces the salt of the acid instead of the acid itself. Meanwhile, pH of the
fermentation broth goes on lowering as lactic acid is formed and accumulated. Therefore, pH of the
fermentation must be maintained, usually by addition of lime, thus lactic acid (partly) is converted to be
calcium lactate. The precipitation and acidification of calcium lactate then generates huge quantity of
calcium sulphate (gypsum). Approximately one ton of gypsum by-product is produced for every ton of
lactic acid produced by the conventional fermentation, which poses an environmental problem [2]. In
addition, the fermentation broths of lactic acid also contain impurities such as a residual sugars,
nutrients, and microorganisms, and other organic acids [7, 41]. Therefore, the complex separation steps
to recover and purify the lactic acid from the fermentation broths become one of the major economic
hurdle and process cost of conventional carbohydrate fermentation process.

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IOP Conf. Series: Materials Science and Engineering 622 (2019) 012023 doi:10.1088/1757-899X/622/1/012023

Table 2. Comparison of different strains and substrates for lactic acid production

Characteristic Homofermentative strain Heterofermentative strain

 Lactic acid  Lactic acid
 Ethanol
 Diacetyl
Products
 Formate
 Acetic acid
 Carbon dioxide
 Lactococcus  Leuconostoc
 Streptococcus  Oemococcus
 Pediococcus  Bifidobacterium
Genera
 Enterococcus  Lactobacillus
 Lactobacillus
 Sporolactobacillus
Low selectivity (high by-product
Selectivity of lactic acid High selectivity
formation)
Availability for
Available Not available
commercial production

Various technologies have been introduced for lactic acids recovery from the fermentation broth,
such as precipitation, extraction, adsorption, ion-exchange system, membrane, etc. [51-59]. However,
those existing processes still have many obstacles that need to be addressed. Precipitation and liquid-
liquid extraction need a large number of chemical agents and energy. They produce a large amount of
water effluent as well as solid residue and involve phase changes that lead to quality degradation of
citric acid [60-62]. Meanwhile, adsorption has a short lifetime of adsorbents and low capacity [54, 60].
In addition, fouling still becomes the most significant hurdle in membrane processes [63, 64].

3. Results and discussion

The increasing demand of lactic acid and increasing concern over environmental impact of gypsum
accumulation as a by-product leads to development of alternative technologies for lactic acid production.
Batch, fed-batch, repeated batch, and continuous fermentations have been developed to increase
production efficiency of lactic acid. Various studies showed that higher lactic acid concentration could
be obtained in batch and fed-batch fermentations, however higher productivity was achieved by the use
of continuous fermentations [15]. It is due to the low concentration of microbes in the batch system.
To solve the problem of production efficiency as well as gypsum formation, membrane bioreactor
can be a promising alternative. Membrane bioreactor integrates the fermentation and separation step
using membrane such as microfiltration, ultrafiltration, Nano filtration, and electro dialysis, thus
simultaneously maintaining high cell density, recycling the cells for further use, and continuously
removing lactic acid from the fermenter [65-68]. In continuous fermentation processes using membrane
bioreactor, components like microbial cells, proteins, nutrients (yeast extract, salts of ammonium,
potassium, phosphorus, etc.), unconverted carbon sources, and water are separated by a filtration unit
and returned to the fermenter , while the lactic acid is concentrated in the permeate [4, 12, 69]. Therefore,
the use of membrane bioreactor in lactic acid production does not produce any harmful bio-product. The
detail comparison of membrane bioreactor and conventional technologies for lactic acid production are
shown in Table 3.
In general, both polymer (i.e. polysulfone [65] and polyacrylonitrile [70]) and ceramic (alumina [71])
can be used as membrane bioreactor materials to produce lactic acid. Ceramic membrane tends to have
better mechanical and chemical resistance than polymer membrane. Meanwhile, polymer membrane
offers advantages of flexible configuration and low production cost [72, 73]. There are only a few
researchers who mention the material of membrane bioreactor for lactic acid production. They mostly
focused on the operation mode and product efficiency.

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IOP Conf. Series: Materials Science and Engineering 622 (2019) 012023 doi:10.1088/1757-899X/622/1/012023

Table 3. Comparison of membrane bioreactor and conventional technologies for lactic acid
production [12, 50, 74]

Characteristic Membrane bioreactor Conventional technology

Unit operations  Fermentation tank  Fermentation tank
 Membrane module  pH adjustment through hydrated lime
 Rotary drum filtration
 Carbon bleaching
 Plate and frame filtration
 Calcium lactate evaporation
 Crystallization
 Extraction
 Distillation unit
 Dilution
Flexibility parameters
 Modules Flexible in size and numbers Fixed configuration
 Production capacity Flexible Fixed
 Steps of operations Few steps Large number of steps
Effect to the environment  Does not produce any harmful  Generates million tons of
and economic aspects bio-product gypsum/year
 Free from pre- and post-treatment  Heat generation due to exothermic
 Low energy consumption reaction
 Use no harsh chemicals  Highly energy intensive
 Use harsh chemicals, especially acids
and alkalis

In lactic acid production, the operation mode of membrane bioreactor can be divided into batch,
semi-continuous, continuous, immobilized, and membrane recycle. Among those modes, membrane
recycle bioreactor has the highest efficiency in production of lactic acid, as shown in Table 4. However,
the high productivity is not the only requirement for the economic feasibility of the process. Timmer
and Kromkamp [44] investigated that the process might be primarily influenced by production capacity
and product concentration and to a lesser extent by the volumetric productivity when annual lactic acid
production capacity rose to as high as 4540 metric tons. When lactic acid concentration is significantly
low, the energy cost for water removal in the downstream process offsets the benefits of the increased
productivity.
In 1986, Mehaia and Cheryan [37] produced lactic acid from acid whey permeate in a membrane
recycle bioreactor. Whey permeate was obtained by ultrafiltration of cottage cheese whey and
supplemented with yeast extract. The lactose in the permeate was converted into lactic acid by
Lactobacillus bulgaricus in a high-performance membrane bioreactor configured in the cell recycle
mode. At a cell concentration of 10 g/L, optimum productivity of lactic acid was only 35 g/L.h.
Meanwhile, lactic acid productivity of 80 g/L.h only could be obtained at cell concentration of 60 g/L.
Other studies [89-93] also showed that lactic acid concentration produced by membrane bioreactor was
still below 60 g/L.
To enhance the economic advantage of the membrane bioreactor process, methods that increase the
lactic acid concentration along with the high-cell density are required. Ohashi et al. [84] studied
continuous production of lactic acid by retaining cells at a high density of Lactococcus lactis in a stirred
ceramic membrane reactor (SCMR). The mass concentration and productivity of lactic acid reached 40
g/L and 10.6 g/L.h, respectively. In 1998, Kamoshita et al. [94] developed the improved SCMR system.
Using the improved SCMR system, a cell mass concentration of 178 g/L and viability of 98 % were
obtained after 198 h of fermentation. They demonstrated that the improved permeability of the SCMR
with the use of a membrane cleaning system influenced a rapid increase in the concentration and viability

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IOP Conf. Series: Materials Science and Engineering 622 (2019) 012023 doi:10.1088/1757-899X/622/1/012023

of cells, and accordingly, the increased production rate of lactic acid in proportion to the concentration
of viable cells.
Table 4. Comparison of bioreactors for lactic acid production

Lactic Productivity
Bioreactor Substrate Organism Ref.
acid (g/L) (g/L.h)
Banana wastes Lactobacillus casei - 0.13 [75]
Cassava bagasse Lactobacillus 81.9 1.36 [76]
Batch delbrueckii
Sugar cane baggage Lactococcus lactis 10.9 0.17 [77]
Cheese whey Lactobacillus bulgaricus 9.6 4.8 [78]
Semi- Whey permeate Lactobacillus helveticus 55 3.54 [69]
continuous
Whey permeate Lactobacillus helveticus 64 22 [79]
Continuous Glucose Lactobacillus lactis 210 2.2 [80]
Cheese whey Lactobacillus helveticus 38 19-22 [81]
Glucose Lactobacillus lactis 115 2.25 [82]
Immobilized Liquid distillery Lactococcus rhamnosus 42.2 1.22 [83]
stillage
Whey permeate Lactobacillus bulgaricus 43 85 [37]
Molasses Lactococcus lactis 40 10.6 [84]
Glucose Lactococcus rhamnosus 88 35.2 [85]
Membrane
Glucose Lactococcus paracasei 91 31.5 [86]
recycle
Glucose Lactococcus paracasei 120 150 [87]
Glucose Lactobacillus 35 76 [88]
delbrueckii

Another effort to increase lactic acid productivity was developed by combining the advantage of both
membrane bioreactor and multi-staged bioreactor. Kulozik et al. [95] investigated the performance of a
seven-staged cascade reactor with cell recycle. In comparison with a single-stage membrane bioreactor,
the cascade reactor resulted 4 times higher productivity, 28 g/L.h, in which the cell concentrations were
maintained at 20 g/L and the lactic acid concentrations were around 72 g/L. In 2001, Kwon et al. [96]
produced lactic acid by a two-stage cell-recycle culture of Lactococcus rhamnosus. The membrane cell-
recycle bioreactors were arranged in a series and successfully obtained 92 g/L of lactic acid with a
productivity of 57 g/L.h. Meanwhile, Xu et al. [86] developed a membrane cell-recycle bioreactor that
equipped with a diaphragm pump and tangential flow-rate controller to produce lactic acid by
Lactococcus paracasei. The maximum productivity of this system was 31.5 g/L.h.
Later, Danner et al. [97] developed UF membrane bioreactor coupled with on-line monopolar electro
dialysis to recover, pre-purify, and concentrate lactic acid. The results showed that the volumetric
productivity was low (1.38 g/L h). The lactic acid concentration was 35 g/L with lactic acid yield on
consumed glucose appeared stable at around 80%.

4. Conclusion
Fermentative production of lactic acid has roused interest among researchers in the recent years due to
its high potential of application in a wide range of fields. High purity lactic acid could be produced and
separated continuously in a fully membrane-integrated fermentation process using a cheap and
renewable carbon source. Various types of substrate such as whole-wheat powder, starch, cheese whey,
molasses, and sugarcane juice have been investigated to produce lactic acid with the help of organisms,
especially from the genera Lactobacillus.

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IOP Conf. Series: Materials Science and Engineering 622 (2019) 012023 doi:10.1088/1757-899X/622/1/012023

In general, the efficiency of lactic acid production by membrane bioreactor is higher than
conventional technologies. However, several studies showed that membrane bioreactor was only able
to produce lactic acid with concentration below 60 g/L. Therefore, further development of methods that
increase the lactic acid concentration was required. The improvement in continuous fermentation could
be done by increasing the cell density in the bioreactor, thus enhances the substrate-to-product
conversion rate and resulting in higher productivity. The increase of cell densities in the fermentation
process can be conducted either by the use of cell immobilization or by cell-recycle using membrane

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3rd MRS-ID meeting 2018 IOP Publishing
IOP Conf. Series: Materials Science and Engineering 622 (2019) 012023 doi:10.1088/1757-899X/622/1/012023

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