Bioresource Technology 217 (2016) 129–136

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Valorisation of food waste via fungal hydrolysis and lactic acid fermentation with Lactobacillus casei Shirota Tsz Him Kwan, Yunzi Hu, Carol Sze Ki Lin ⇑ School of Energy and Environment, City University of Hong Kong, Tat Chee Avenue, Kowloon, Hong Kong

h i g h l i g h t s

g r a p h i c a l a b s t r a c t

 We investigated bioconversion of

different types of food waste to lactic acid.  Hydrolysates were rich in glucose, fructose and free amino nitrogen.  Food waste powder produced by commercial food waste machine can be used as feedstock.  The overall yields were 0.23–0.27 g lactic acid g 1 food waste.  We proposed a novel decentralized approach for food waste bioconversion in urban area.

a r t i c l e

i n f o

Article history: Received 29 December 2015 Received in revised form 29 January 2016 Accepted 30 January 2016 Available online 9 February 2016 Keywords: Food waste treatment Lactic acid fermentation Platform chemicals Lactobacillus casei Shirota Hydrolysis

Fungal hydrolysis Mixed food waste

Lacc acid Bakery waste

Cell biomass Food waste powder

Remaining solids

Lipids

a b s t r a c t Food waste recycling via fungal hydrolysis and lactic acid (LA) fermentation has been investigated. Hydrolysates derived from mixed food waste and bakery waste were rich in glucose (80.0– 100.2 g L 1), fructose (7.6 g L 1) and free amino nitrogen (947–1081 mg L 1). In the fermentation with Lactobacillus casei Shirota, 94.0 g L 1 and 82.6 g L 1 of LA were produced with productivity of 2.61 g L 1 h 1 and 2.50 g L 1 h 1 for mixed food waste and bakery waste hydrolysate, respectively. The yield was 0.94 g g 1 for both hydrolysates. Similar results were obtained using food waste powder hydrolysate, in which 90.1 g L 1 of LA was produced with a yield and productivity of 0.92 g g 1 and 2.50 g L 1 h 1. The results demonstrate the feasibility of an efficient bioconversion of food waste to LA and a decentralized approach of food waste recycling in urban area. Ó 2016 Elsevier Ltd. All rights reserved.

1. Introduction There is an increasing interest in recycling food waste due to massive amount of generation and the valuable organic content to be recycled for various applications such as composts, animal feed and biogas (Lin et al., 2014). According to a study commissioned by the United Nations Food and Agriculture Organisation in 2011, roughly one-third of food produced for human consumption was lost or wasted globally, which results in the generation ⇑ Corresponding author. Tel.: +852 3442 7497; fax: +852 3442 0688. E-mail address: [email protected] (C.S.K. Lin). http://dx.doi.org/10.1016/j.biortech.2016.01.134 0960-8524/Ó 2016 Elsevier Ltd. All rights reserved.

Lacc acid fermentaon by Lactobacillus casei Shirota

of 1.3 billion tonnes of food waste per year (FAO, 2011). In Hong Kong, daily food waste generation has reached 3640 tonnes, accounting for 37% of municipal solid waste generation in 2015 (EPD, 2015). Although there are a number of food waste recycling technologies available, none of them can eradicate the food waste problem in highly urbanized city like Hong Kong due to low selling price of regenerated products, scattered food waste generation sources and high transportation costs (Kwan et al., 2014; Pleissner, 2016). Therefore, the development of a practical approach to turn food waste into value-added products is highly desired. Food waste, which is defined as any waste and by-products produced during the food production, processing, wholesale, retail and

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consumption (FAO, 2011), consists of 30–60% starch, 5–10% proteins and 10–40% lipids (w/w) (Pleissner and Lin, 2013). Due to the nutrient-rich composition of food waste, its utilization as feedstock in biorefineries for chemicals, materials and fuels production has been proposed and demonstrated in the recent years in order to reduce the amount of organic waste that needs to be treated and to help alleviating the over-dependence on petroleum (Lin et al., 2013). In our previous study, a process for the production of generic fermentation feedstock from food waste via fungal hydrolysis in submerged fermentation by Aspergillus awamori and Aspergillus oryzae was firstly introduced (Pleissner et al., 2014a). Compared to the traditional food waste recycling technologies, fungal hydrolysis owns a lot of environmental and social advantages including free of air pollutant formation, free of unpleasant smell and not energy intensive. Most importantly, it allows recycling of nutrients in food waste through the use of the hydrolysate as fermentation feedstock for the production of value-added biobased products. For example, we previously demonstrated the utilization of waste bread hydrolysate as fermentation feedstock for succinic acid production by Actinobacillus succinogenes (Leung et al., 2012) and biodegradable polymer polyhydroxybutyrate by Halomonas boliviensis (Pleissner et al., 2014b). Pleissner et al. (2013) reported cultivation of Chlorella pyrenoidosa using mixed food waste hydrolysate as culture medium for the production of algal biomass. Techno-economic analyses have been carried out to investigate the economic feasibility of pilot scale operation by using simulation software. It was noted that the economic profitability was not attractive for upscaling due to prolonged fermentation time and low concentration of products (Lam et al. 2014, Kwan et al. 2015). Nevertheless, Kwan et al. (2015) indicated that fermentative lactic acid production using food waste as raw material could lead to a highly attractive economic profitability since the duration of lactic acid fermentation is relatively short and high concentration of lactic acid can be achieved easily. Lactic acid (LA) was identified as one of the twelve most promising value-added building blocks derived from sugars with a high potential to be a key building block for the production of both commodity and specialty chemicals (DOE, 2004). It has diverse applications including preservative, pH adjusting agent, a starting material in the production of lactate ester, active ingredient in personal care products and monomer in the production of biodegradable polymer polylactic acid. These diverse applications of LA lead to the growing industrial application at a rate of 5– 10% per year (Corbion purac, 2015). There are a number of studies of LA fermentation using different carbon sources as raw materials such as food industry by-products (e.g. kitchen waste, whey), agroindustrial residues and by-products (e.g. cottonseed hulls, corn cob, corn stalks, wheat bran, brewer’s spent grains) and renewable resources (e.g. Jerusalem artichoke hydrolysates) (Venus, 2006; Venus and Richter, 2006; Wang et al., 2011). Different processes including enzymatic hydrolysis and fermentation, simultaneous hydrolysis and fermentation, open fermentation and direct fermentation have been intensively studied with different LA producing microorganisms (Pleissner et al., 2016; Uçkun Kıran et al., 2015). However, the bioconversion of food waste for LA production via fungal hydrolysis and microbial fermentation has not been reported. Furthermore, most of these studies highlighted the cost-efficiency of using waste as raw material and emphasized on the potential to develop economically feasible processes. Techno-economic evaluation has not been carried out due to the lack of the mass balance of the whole process. Starting from July in 2013, Environment and Conservation Fund operated by Environmental Protection Department in Hong Kong has allocated HK$ 50 million as subsidy to encourage the separated

collection and recycling of food waste (EPD, 2011). The subsidy provides funds to support various recycling activities as well as setting up on-site food waste treatment machines. By using the food waste treatment machines, up to 70% volume reduction can be achieved by shredding, grinding and dehydrating at over 100 °C. As one of the most populated cities in the world, the use of food waste treatment machines can facilitate a decentralized process of food waste recycling and also save transportation cost of food waste collection, which can account for more than 70% of the operation cost of recycling food waste according to a local food waste recycling company (HKOWRC, 2015). In view of that, the incorporation of the food waste treatment machines in the bioconversion of food waste could be an advantageous approach for recycling food waste in urban area. This study aims to demonstrate the bioconversion of mixed food waste and bakery waste to LA via fungal hydrolysis and fermentation by Lactobacillus casei Shirota. Feasibility of using food waste powder produced by food waste treatment machine as raw materials for LA production is explored for a decentralized approach for food waste recycling in urban area. The proposed bioprocess can be integrated in a traditional transesterification for the production of biodiesel and an important platform chemical. 2. Methods 2.1. Microorganisms A. awamori ATCC 14331 was purchased from the American Type Culture Collection (Rockville, MD, USA). A. oryzae was isolated from a soy sauce starter provided by the Amoy Food Ltd., Hong Kong (Leung et al., 2012). Spore solutions of both fungi were produced as described earlier (Lam et al., 2013). L. casei Shirota was obtained from Prof. Nagendra Prasad SHAH at School of Biological Sciences in University of Hong Kong. It was cultivated in MRS broth with 20 g L 1 initial glucose concentration in a shaking incubator at 200 rpm for 18 h at 37 °C. 2.2. Food waste handling Mixed food waste was collected from Asia Pacific Catering located in the Hong Kong Science Park and bakery waste was collected from Starbucks outlet located in Sha Tin. The mixed food waste was the leftovers consisting of rice, noodles, meat and vegetables, while the bakery waste was unsold products including cakes, breads and pastries. These wastes were separately blended and stored at 4 °C until use, but for no more than one week. Food waste powder was produced using blended mixed food waste processed by a commercial food waste treatment machine (SPZ-3000D, Spinz, Korea) donated by Eco-Greenergy Limited. The process included shredding and dehydration at 100–150 °C. Two output streams consisted of food waste powder and wastewater effluent generated during the cleaning step of food waste machine. 2.3. Solid state fermentation Solid state fermentation was carried out to produce fungal solid mashes, which were used as enzyme sources in submerged food waste hydrolysis. For 8.5 g (Dry weight) of mixed food waste or bakery waste, 1 mL of spore solution of A. awamori (4.6  105 spores mL 1) or 1 mL of spore solution of A. oryzae (6.3  105 spores mL 1) was added, mixed, and incubated for 7 days at 30 °C. It was performed as described in our previous publication (Lam et al., 2013; Pleissner et al., 2014a).

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Table 1 Compositions of food wastes used in this study.

Carbohydrates (mg g 1) Starch (mg g 1) Proteins (mg g 1) Total nitrogen (mg g 1) Lipids (mg g 1) Ash (mg g 1)

Mixed food waste

Food waste powder

Bakery waste

502.1 ± 10.7 363.2 ± 13.4 138.1 ± 12.1 24.2 ± 2.1 305.0 ± 14.0 22.8 ± 2.7

523.5 ± 2.5 359.6 ± 11.8 127.6 ± 2.8 22.4 ± 0.5 258.2 ± 3.1 23.3 ± 1.0

492.4 ± 0.7 291.4 ± 9.5 84.2 ± 0.7 14.8 ± 0.1 370.6 ± 8.4 26.4 ± 1.8

Table 2 Compositions of the hydrolysates of mixed food waste, food waste powder and bakery waste.

Glucose (g L 1) Fructose (g L 1) Free amino nitrogen (mg L

1

)

Mixed food waste hydrolysate

Food waste powder hydrolysate

Bakery waste hydrolysate

100.2 ± 2.4 Nil 1081.0 ± 70.2

97.2 ± 1.0 Nil 946.5 ± 41.7

80.0 ± 1.5 7.6 ± 0.5 864.6 ± 54.3

2.4. Fungal hydrolysis in submerged fermentation Fungal hydrolysis was carried out in a 2.5 L bioreactor (BioFlo/ CelliGen 115, New Brunswick Scientific, NJ, USA) at 55 °C for 48 h in duplicate (Pleissner et al., 2014a). No control of pH was needed as the pH value remained between 4.0 and 4.5, which was found to be appropriate for enzymes secreted by A. awamori and A. oryzae to be active (Lam et al., 2013). The broth was stirred at 1200 rpm at the initial 10 h and decreased to 400 rpm when most of the food waste was dissolved and hydrolyzed. Blended food waste was mixed with deionised water at a solid-to-liquid ratio at 30% (w/v) that 300 g (dry weight) of food waste was mixed with 1 L of deionised water. Fungal mashes of A. awamori (14 g) and A. oryzae (14 g) were successively added at the beginning and after 24 h, respectively. The broth was harvested, centrifuged at 11500g for 30 min and stored in 4 °C for 5 h. Lipid solidified at the top was firstly collected and used for other study of biodiesel production via transesterification (Karmee et al., 2015). The hydrolysate was then separated from the remaining solid fraction by vacuum filtration using Whatman No. 1 filter paper. The hydrolysate and solid residues were kept frozen at 20 °C until use. 2.5. Utilization of glucose and fructose for LA production by L. casei Shirota Experiments were carried out in 250 mL conical flasks containing 100 mL of MRS broth, which consisted of 2 g L 1 K2HPO4, 20 g L 1 glucose, 0.2 g L 1 H14MgO11S, 0.05 g L 1 H8MnO8S, 8 g L 1 meat extract, 10 g L 1 peptone, 5 g L 1 CH3COONa  3H2O, 2 g L 1 C6H17N3O7 and 4 g L 1 yeast extract. Glucose and fructose solutions were separately autoclaved and added to the MRS broth to adjust the sugar concentrations. Phosphate buffer (10 mM) was employed to maintain the pH between 5.5 and 6.5, and 5 M NaOH

was used to adjust the pH if necessary. Seed culture was cultivated using MRS broth in 37 °C for 24 h. The inoculum size was 2% (v/v). Samples were aseptically taken at every 3 h to measure the pH, cell dry weight (DCW), sugars and organic acids concentrations. Cell growth, sugar consumption, and LA production were studied at different glucose and fructose concentrations. Experiment was carried out in duplicate for sugar concentration ranged from 0 to 100 g L 1. 2.6. Bench-top scale fermentation Batch fermentations were carried out in two 2.5 L fermentors (Biostat, Sartorius stedim, Germany) for lactic acid production using hydrolysates and defined medium, respectively. Hydrolysate supplemented with 10 g L 1 yeast extract (Angel Yeast Co., Ltd, China) was filter-sterilized by a 0.2 lm PTFE membrane filter (Sartorius, Germany) before adding to the fermentor. The temperature was set as 37 °C and pH was automatically controlled at 6.0 using 10 M NaOH solution during the fermentation. The broth was sparged with 2 vvm air and agitated at 200 rpm. Seed culture was cultivated using MRS broth in 37 °C for 24 h. The inoculum size was 2% (v/v). Glucose and fructose solutions were separately autoclaved and added to the MRS broth to adjust the sugar concentrations. Samples were aseptically taken to measure the cell dry weight, sugars and lactic acid concentrations. Fermentations were ended when sugars were completely depleted, or when no change in total sugar concentration for a period of 5 h. 2.7. Determinations of cell dry weight, sugars, free amino nitrogen and lactic acids concentrations After sampling, cells and supernatant were separated by centrifugation at 13,000 rpm for 3 min. The cell biomass was washed

Table 3 L. casei Shirota fermentation performance using glucose and fructose separately as carbon source. Initial sugar concentrations (g L 1)

Fermentation time (h)

Specific growth rate (h 1)

DCW (g L 1)

Lactic acid concentration (g L 1)

Yield (g g 1)

Productivity (g L 1 h 1)

Glucose 20 g L 1 40 g L 1 60 g L 1 80 g L 1 100 g L 1

27 33 47 59 75

0.21 ± 0.00 0.23 ± 0.00 0.24 ± 0.01 0.23 ± 0.00 0.23 ± 0.00

2.6 ± 0.0 5.0 ± 0.1 6.2 ± 0.2 7.4 ± 0.0 7.6 ± 0.1

15.5 ± 0.5 32.7 ± 0.2 49.2 ± 0.6 62.9 ± 1.0 75.9 ± 0.0

0.77 ± 0.1 0.81 ± 0.0 0.81 ± 0.1 0.78 ± 0.0 0.76 ± 0.0

0.57 ± 0.02 1.00 ± 0.07 1.05 ± 0.01 1.07 ± 0.02 1.00 ± 0.02

Fructose 20 g L 1

21

0.24 ± 0.00

3.8 ± 0.1

16.9 ± 1.6

0.83 ± 0.1

0.81 ± 0.03

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Fig. 2. LA fermentation profile using (a) bakery waste hydrolysate and (b) defined medium as feedstock. (j) Glucose; (r) fructose; (N) lactic acid; (s) cell biomass.

2.8. Compositions of mixed food waste and bakery waste Quantification of carbohydrate, proteins and lipids in bakery and mixed food wastes was performed in triplicate as described in Pleissner et al. (2013). All specific contents of waste constituents reported in this study are based on dry weight. Starch quantification was carried out in triplicate using starch test kit (Megazyme, Ireland) (Koutinas et al., 2007). Fig. 1. LA fermentation profile using (a) mixed food waste hydrolysate (b) mixed food waste powder hydrolysate produced by food waste treatment machine and (c) defined medium as feedstock. (j) Glucose; (N) lactic acid; (s) cell biomass.

3. Results and discussion 3.1. Food waste composition and fungal hydrolysis

with 10 mL 0.9% NaCl twice and centrifuged to remove the supernatant. The residual biomass was then washed with deionised water and dried at 90 °C until the weight was stable. Sugars and organic acids were analysed using high performance liquid chromatography (HPLC, Waters, UK) equipped with Aminex HPX-87H column (Bio-Rad, CA, USA). The supernatants were filtered by 0.22 lm 13 mm Nylon membranes filter (Jin Teng, China) prior to the analysis. In each analysis, 10 lL was injected into the column and eluted isocratically with 0.4 mL/min with 5 mM H2SO4 at 65 °C. Detection was performed by a RI detector (Waters, UK) at 35 °C and photodiode array (PDA) analyser (Waters, UK) at 210 nm. Free amino nitrogen (FAN) concentration was measured by using nynhidrin reaction method (Lie, 1973). All quantifications were carried out in duplicate.

The composition of mixed food waste and bakery waste are shown in Table 1. Result shows that both mixed food waste and bakery waste were rich in carbon and nitrogen sources of around 50% carbohydrate, 29–36% starch, 8–14% protein and 30–37% lipids. The mixed food waste could be hydrolysed into generic medium with higher concentration of glucose since the starch content in mixed food waste (363.2 ± 13.4 mg g 1) was higher than that in bakery waste (291.4 ± 9.5 mg g 1). Although a high variability between different kinds of food waste has been reported (Sayeki et al., 2001), there is a consistency of the composition of food waste from similar sources. Our previous studies have reported that mixed food and bakery waste generally consist of 30–60% starch, 5–10% proteins and 10–40% lipids (Leung et al., 2012; Pleissner et al., 2014a, 2013; Zhang et al., 2013), which is close to the values reported in this study. On the other hand, food

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T.H. Kwan et al. / Bioresource Technology 217 (2016) 129–136 Table 4 Comparison of fermentation performance using defined medium and hydrolysates as feedstock. Mixed food waste

Initial glucose concentration (g L 1) Initial fructose concentration (g L 1) Specific growth rate (h 1) Cell dry weight (g L 1) Glucose uptake rate (g L 1 h 1) Fructose uptake rate (g L 1 h 1) LA concentration (g L 1) Yield (g LA g 1 sugar) Productivity (g L 1 h 1) *

Bakery waste

Defined medium

Hydrolysate

Powder hydrolysate

100.0

100.2

97.5

0.22 4.94 2.79

0.23 6.60 4.55

0.22 6.45 4.67

93.2 0.93 1.55

94.0 0.94 2.61

90.1 0.92 2.50

*

Defined medium

Hydrolysate

80.0 7.6 0.23 4.79 2.43 0.66 81.6 0.93 1.51

80.2 7.5 0.23 6.54 4.14 0.70 82.6 0.94 2.50

Powder hydrolysate was prepared from mixed food waste processed by food waste treatment machine.

Table 5 Summary of LA production using different kinds of food waste as raw materials. Microorganism

Substrate

Process

LA Conc. (g L 1)

Yield (g g 1)

Productivity (g L 1 h 1)

Refs.

Lactobacillus casei Shirota Lactobacillus casei Shirota Thermoanaerobacterium aotearoense LA1002 Bacillus coagulans NBRC12583 Mixed culture

Mixed food waste Bakery waste Bakery waste

Fungal hydrolysis & fermentation Fungal hydrolysis & fermentation Fungal hydrolysis & fermentation

94.0 82.6 78.4

0.94 0.94 0.85

2.61 2.50 1.63

This study This study Yang et al. (2015)

Kitchen waste

Enzymatic hydrolysis & fermentation Enzymatic hydrolysis & fermentation Proteolytic pretreatment & fermentation Direct fermentation Direct fermentation

86.0

0.98

0.72

34.5

0.54

0.21

37.0

0.84

36.3 48.7

0.70

1.01 0.75

33.8 45.5

0.73

0.47 0.75

Sakai and Ezaki, (2006) Tashiro et al. (2013) Pleissner et al. (2015) Wang et al. (2011) Ohkouchi and Inoue, (2006) Wang et al. (2005) Wang et al. (2009)

36.6

0.88

2.78

Bacillus coagulans Lactobacillus TY50 Lactobacillus manihotivorans LMG18011 Lactobacillus sp. TH165 &175 Lactobacillus rhamnosus 6003 Lactobacillus rhamnosus CECT-288

Kitchen waste Defatted algal biomass and defatted food waste Kitchen waste Kitchen waste Kitchen waste Kitchen waste Apple pomace

Direct fermentation Simultaneous enzymatic hydrolysis and fermentation Simultaneous enzymatic hydrolysis and fermentation

waste powder produced by food waste treatment machine did not have significant compositional change (Table 1), except a 15% decrease of lipid content resulted from the degradation of lipids at high temperature (Bougrier et al., 2008). Furthermore, the powder was dark brown in color, which indicated that the Maillard reaction may take place during the dehydration at high temperature. Fungal hydrolyses were performed in order to recover nutrients such as glucose, fructose and FAN, which are essentially required for the subsequent LA fermentation by L. casei Shirota. As shown in Table 2, glucose (100.2 ± 2.4 g L 1) and FAN concentrations (1,081 ± 70.2 mg L 1) were obtained at 30% (w/v) mixed food waste hydrolysate. However, small reduction in glucose (97.2 ± 1.0 g L 1) and FAN (946.5 ± 41.7 mg L 1) presented in the hydrolysate derived from food waste powder produced by food waste treatment machine, mainly due to the Maillard reaction mentioned earlier. On the other hand, glucose (80.0 ± 1.5 g L 1), fructose (7.6 ± 0.5 g L 1) and FAN concentrations (864.6 ± 54.3 mg L 1) were resulted at 30% (w/v) bakery waste hydrolysate. The composition of these hydrolysates was consistent with the earlier research carried out by our group (Pleissner et al., 2014a; Zhang et al., 2013). 3.2. Utilization of glucose and fructose for LA production by L. casei Shirota Lactic acid production, cell growth and sugar consumption of L. casei Shirota has been investigated at 20–100 g L 1 glucose and 20 g L 1 fructose. The L. casei Shirota strain used in this study was isolated from food by Minoru Shirota in 1930 (Shirota et al., 1966). Since then, it has received a great attention of research on

Gullón et al. (2007)

safety for human consumption as it is widely used to manufacture fermented milk products. However, none of the study has reported the influence of glucose and fructose concentrations on L. casei Shirota fermentation for LA production. Table 3 summarises the cell growth and LA production under different glucose and fructose concentrations. A similar specific growth rate (0.21–0.24 h 1) was found among all the concentrations and maximal DCW (7.6 ± 0.1 g L 1) was achieved at 100 g L 1 of glucose concentration. Sugars were depleted at the end of all fermentations. The maximal LA production (75.9 g L 1) was found at 100 g L 1 of glucose concentration with a yield and productivity of 0.76 g g 1 and 1.00 g L 1 h 1, respectively. These results indicated that L. casei Shirota can convert both glucose and fructose to LA and the sugars presented in the hydrolysates could be fully utilized for LA production. 3.3. Batch fermentation of lactic acid 3.3.1. Mixed food waste hydrolysate Fig. 1a shows the profile of fermentation using mixed food waste hydrolysate as feedstock. It was ended at 36 h when 94.0 g L 1 of LA was produced with a yield of 0.94 g g 1 and a productivity of 2.61 g L 1 h 1 (Table 4). The specific growth rate and DCW were found to be 0.23 h 1 and 6.60 g L 1 respectively. Compare to defined medium, fermentation using mixed food waste hydrolysate achieved higher glucose uptake rate (4.55 g L 1 h 1) and productivity of LA production, which could attribute to the higher cell biomass obtained in hydrolysate than that of defined medium (4.94 g L 1). This indicated that mixed food waste hydrolysate contained sufficient nutrients to support the cell growth and

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no other inhibitory compounds which inhibit the cell growth and LA production by L. casei Shirota. 3.3.2. Food waste powder hydrolysate The hydrolysate derived from the food waste powder produced by food waste treatment machine was used as feedstock for LA fermentation. After 36 h, the fermentation ended with LA concentration, yield and productivity of 90.1 g L 1, 0.92 g g 1 and 2.50 g L 1 h 1, respectively (Fig. 1b). Table 4 shows the comparison of fermentation performance using mixed food waste hydrolysate and defined medium. It was found that the cell growth, sugar consumption and LA production were similar, compared to the one without processed by food waste treatment machine as reported in the previous Section (3.3.1). The results show that the treatment of food waste machine did not significantly affect the LA fermentation performance of L. casei Shirota using food waste powder hydrolysate. 3.3.3. Bakery waste hydrolysate The profile of fermentation using bakery waste hydrolysate as feedstock is shown in Fig. 2a. At 33 h, 82.6 g L 1 of LA was produced which corresponds to a yield and productivity of

0.94 g g 1 and 2.50 g L 1 h 1, respectively. It was noted that L. casei Shirota converted both glucose and fructose to LA, and it began to utilize fructose when glucose was about to be depleted. Compared to the fermentation using defined medium as feedstock, more cell biomass (6.54 g L 1) was produced in the fermentation using bakery waste hydrolysate, leading to a higher glucose uptake rate (4.14 g L 1 h 1) and productivity of LA. However, similar amount of LA and yield were obtained. This suggests that bakery waste hydrolysate contains sufficient nutrients to facilitate the cell growth and the sugars recovered from bakery waste can be efficiently utilized for LA production. Various processes and microorganisms have been reported for the bioconversion of food wastes to LA (Uçkun Kıran et al., 2015). Table 5 summarises the LA fermentation results using various modes of fermentation processes and microorganisms. Ohkouchi and Inoue (2006) reported direct fermentation of starch to lactic acid by an amylolytic lactic acid bacterium Lactobacillus manihotivorans LMG18011 for a shortened processing time. Wang et al. (2009) applied the crude enzyme produced from Aspergillus niger for simultaneous enzymatic hydrolysis and lactic acid fermentation with Lactobacillus rhamnosus 6003 in order to reduce the processing time and the cost of enzymes. Although these studies

Fig. 3. Process flow diagram of the proposed bioconversion process (a) mixed food waste and bakery waste to lactic acid (b) food waste powder to lactic acid.

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T.H. Kwan et al. / Bioresource Technology 217 (2016) 129–136 Table 6 Mass balance of bioconversion of mixed food waste, bakery waste and food waste powder produced by food waste treatment machine to lactic acid (unit: kg). Stream Substrate: mixed Component Carbohydrate Protein Lipids Ash Yeast extract NaOH Cell biomass Lactic acid H2O Total

Food waste

Yeast extract

Water

NaOH

Remaining solids

Lipids

Cell biomass

Effluent

Lactic acid

food waste 459.4 126.4 279.1 20.9

42.6 11.8 26.0 2.0

74.7 54.5 123.2 4.8

177.0

33.3 142.0 22.0

915.0

Substrate: food waste powder Component Carbohydrate 479.0 Protein 116.8 Lipids 236.3 Ash 21.3 Yeast extract NaOH Cell biomass Lactic acid H2O Total 915.0 Substrate: bakery Component Carbohydrate Protein Lipids Ash Yeast extract NaOH Cell biomass Lactic acid H2O Total

Fungal mashes

85.0

33.3

3333.0 3333.0

355.0 497.0

44.5 21.9 10.8 2.0

263.0

77.9 51.3 112.2 4.5

177.0

22.0

0.0

268.5 268.5 537.0

480.0 480.0

252.3 252.3 504.6

0.0

230.5 230.5 461.0

146.0

33.3 133.5 21.5

85.0

33.3

3333.0 3333.0

332.5 497.0

263.0

177.0

21.0

waste 450.5 77.0 339.1 24.2

41.8 31.6 7.2 2.2

129.9 33.7 117.5 5.2

247.0

33.3 124.8 21.8

915.0

85.0

33.3

3333.0 3333.0

indicated the improved process efficiency with less time and cost requirements, most of them did not achieve high LA concentrations since plenty of carbon sources was not recovered and utilized for LA production, and the yield of LA fermentation was not high. Inefficient recovery of nutrients could lead to a low conversion yield from substrate to LA. Also, low LA concentration leads to an inefficient LA recovery with large volume of waste broth generated during the recovery process. The advantages of the proposed bioprocess in this study clearly showed that 85% of carbohydrate recovery from mixed food and bakery waste through fungal hydrolysis was achieved. Furthermore, L. casei Shirota can produce up to 94.0 g L 1 LA with a yield of 0.94 g g 1 and a productivity of 2.61 g L 1 h 1, and no acid by-products were found in the L. casei Shirota fermentation. 3.4. Mass balance from wastes to products In order to estimate the amount of raw materials needed and products that can be produced in the bioconversion process proposed in this study, a mass balance was calculated based on the experimental data of material input and output. The process flow diagram is presented in Fig. 3, in which all the input and output streams were identified throughout the process. The mass balance is shown in Table 6. Since the water content of food waste can be highly variable, so the weight of raw materials is presented on dry basis. In order to simulate the operation of pilot scale, 1000 kg (dry weight) of waste was considered as raw material. As shown in Table 6, 3333 L of water was added to 1000 kg of food waste and fungal solid mashes at a solid-to-liquid ratio of

312.0 436.8

294.0

247.0

21.8

30.0% in fungal hydrolysis. In fact, the amount of water could be reduced if wet food waste is used and the solid-to-liquid ratio is increased. After 48 h, a total of 334 kg glucose and 3.6 kg FAN were recovered from mixed food waste; 324 kg glucose and 3.2 kg FAN from food waste powder produced by food waste treatment machine; 267 kg glucose, 25 kg fructose and 2.8 kg FAN from bakery waste. According to the yield given in Table 3, 314, 300 and 274 kg of LA could be produced from mixed food waste, food waste powder and bakery waste, respectively. It was assumed that 84% of LA was recovered from fermentation broth by a novel LA recovery process using ethyl acetate (Hu et al., 2016). The overall yields were 0.27 g g 1 mixed food waste, 0.25 g g 1 food waste powder and 0.23 g g 1 bakery waste. The difference of the yields between different kinds of food waste indicate that the variation in nutrient composition have a significant impact on the amount of lactic acid produced in the process, which suggests further economic study should address the effect of composition on the economic performance of the overall process. Remaining solids and lipids were the by-products streams identified in Fig. 3. After the fungal hydrolysis, 263–294 kg of remaining solids and 177–247 kg of lipids were produced (Table 6). The utilization of these by-products streams has been explored to achieve a zero waste approach for the proposed bioprocess. Our previous study successfully extracted the lipids from the remaining solids and turned the crude lipids extract into plasticizer and the defatted solid was hydrolyzed by proteolytic enzymes as a nitrogen source for LA production (Pleissner et al., 2015). Karmee et al. (2015) reported 100% conversion of the lipids recovered from fungal hydrolysis to biodiesel through alkali-catalyzed transesterification

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using KOH. Further study should address the environmental and economic aspects of the bioprocess in order to achieve a circular biobased economy.

4. Conclusions Utilization of mixed food waste, mixed food waste powder produced by food waste treatment machine and bakery waste for LA production has been demonstrated with the overall conversion yields of 0.27, 0.25 and 0.23 g g 1, respectively. Results suggest that fungal hydrolysis and fermentation with L. casei Shirota is an efficient approach for the bioconversion of food waste to LA. Also, the use of food waste treatment machine could facilitate a decentralized approach for food waste recycling. Further study on techno-economic evaluation will be carried out to investigate the economic feasibility of the proposed process. Acknowledgements The authors acknowledge the Innovation and Technology Funding (ITP/002/14TP) from the Innovation and Technology Commission in Hong Kong. We are also grateful for the industrial partners and industrial sponsors, which include Asia Pacific Catering, Hong Kong Organic Recycling Waste Ltd., Central Textiles (Hong Kong) Ltd. and Rizhao Kaishun Tire Co., Ltd. for providing food waste and industrial sponsorships. Special thank is dedicated to Eco-Greenergy Limited for their in-kind donation of food waste processing machine. We also thank Prof. Nagendra Prasad SHAH from the Department of Food Sciences and Technology in The University of Hong Kong for giving lactic acid bacterium L. casei Shirota used in this study. References Bougrier, C., Delgenes, J.P., Carrère, H., 2008. Effects of thermal treatments on five different waste activated sludge samples solubilisation, physical properties and anaerobic digestion. Chem. Eng. J. 139 (2), 236–244. Corbion purac, 2015. Personal communication. In: Jem, K.J., (Ed.), Integrating Green Chemistry Principles Into the Development of Food Waste Valorisation for Sustainable Chemistry. 2015 Annual Meeting of Association of Bio-based Materials Industry. Hsinchu, Tai Wan. DOE, 2004. Top Value Added Chemicals from Biomass Volume I: Results of Screening for Potential Candidates from Sugars and Synthesis Gas. EPD, 2011. Help-desk Service for Food Waste Recycling Projects in Housing Estates. EPD, 2015. Monitoring of Solid Waste in Hong Kong 2014. FAO, 2011. Global Food Losses and Food Waste – Extent, Causes and Prevention, Rome. Gullón, B., Garrote, G., Alonso, J.L., Parajó, J.C., 2007. Production of L-lactic Acid and oligomeric compounds from apple pomace by simultaneous saccharification and fermentation: a response surface methodology assessment. J. Agric. Chem. 55 (14), 5580–5587. HKOWRC, 2015. Personal Communication. Hu, Y., Kwan, T.H., Walid, D., Lin, C.S.K., 2016. Continuous ultrasonic solvent extraction of lactic acid from fermentation broth, submitted for publication. Karmee, S.K., Linardi, D., Lee, J., Lin, C.S.K., 2015. Conversion of lipid from food waste to biodiesel. Waste Manage. 41, 169–173. Koutinas, A., Arifeen, N., Wang, R., Webb, C., 2007. Cereal-based biorefinery development: integrated enzyme production for cereal flour hydrolysis. Biotechnol. Bioeng. 97 (1), 61–72. Kwan, T.H., Lin, C.S.K., Chan, K.M., 2014. Application of food waste valorization technology in Hong Kong. In: Lin, C.S.K., Luque, R. (Eds.), Renewable Resources for Biorefineries. Royal Society of Chemistry, UK, pp. 93–116. Kwan, T.H., Pleissner, D., Lau, K.Y., Venus, J., Pommeret, A., Lin, C.S.K., 2015. Technoeconomic analysis of a food waste valorization process via microalgae cultivation and co-production of plasticizer, lactic acid and animal feed from algal biomass and food waste. Bioresource Technology 198, 292–299.

Lam, K.F., Leung, C.C.J., Lei, H.M., Lin, C.S.K., 2014. Economic feasibility of a pilotscale fermentative succinic acid production from bakery wastes. Food and Bioproducts Processing 92 (3), 282–290. Lam, W.C., Pleissner, D., Lin, C.S.K., 2013. Production of fungal glucoamylase for glucose production from food waste. Biomolecules 3, 651–661. Leung, C.C.J., Cheung, A.S.Y., Zhang, A.Y., Lam, K.F., Lin, C.S.K., 2012. Utilisation of waste bread for fermentative succinic acid production. Biochem. Eng. J. 65, 10– 15. Lie, S., 1973. The EBC-ninhydrin method for determination of free alpha amino nitrogen. J. Inst. Brew. 79 (1), 37–41. Lin, C.S.K., Pfaltzgraff, L.A., Herrero-Davila, L., Mubofu, E.B., Abderrahim, S., Clark, J. H., Koutinas, A.A., Kopsahelis, N., Stamatelatou, K., Dickson, F., Thankappan, S., Mohamed, Z., Brocklesby, R., Luque, R., 2013. Food waste as a valuable resource for the production of chemicals, materials and fuels. Current situation and global perspective. Energy Environ. Sci. 6 (2), 426–464. Lin, C.S.K., Koutinas, A.A., Stamatelatou, K., Mubofu, E.B., Matharu, A.S., Kopsahelis, N., Pfaltzgraff, L.A., Clark, J.H., Papanikolaou, S., Kwan, T.H., Luque, R., 2014. Current and future trends in food waste valorization for the production of chemicals, materials and fuels: a global perspective. Biofuels Bioprod. Biorefin. 8 (5), 686–715. Ohkouchi, Y., Inoue, Y., 2006. Direct production of L(+)-lactic acid from starch and food wastes using Lactobacillus manihotivorans LMG18011. Bioresour. Technol. 97 (13), 1554–1562. Pleissner, D., 2016. Decentralized utilization of wasted organic material in urban areas: a case study in Hong Kong. Ecol. Eng. 86, 120–125. Pleissner, D., Lin, C.S., 2013. Valorisation of food waste in biotechnological processes. Sustainable Chem. Processes 1 (1), 21. Pleissner, D., Lam, W.C., Sun, Z., Lin, C.S., 2013. Food waste as nutrient source in heterotrophic microalgae cultivation. Bioresour. Technol. 137, 139–146. Pleissner, D., Kwan, T.H., Lin, C.S., 2014a. Fungal hydrolysis in submerged fermentation for food waste treatment and fermentation feedstock preparation. Bioresour. Technol. 158, 48–54. Pleissner, D., Lam, W.C., Han, W., Lau, K.Y., Cheung, L.C., Lee, M.W., Lei, H.M., Lo, K.Y., Ng, W.Y., Sun, Z., Melikoglu, M., Lin, C.S.K., 2014b. Fermentative polyhydroxybutyrate production from a novel feedstock derived from bakery waste. Biomed. Res. Int. 2014, 8. Pleissner, D., Lau, K.Y., Schneider, R., Venus, J., Lin, C.S.K., 2015. Fatty acid feedstock preparation and lactic acid production as integrated processes in mixed restaurant food and bakery wastes treatment. Food Res. Int. 73, 52–61. Pleissner, D., Qi, Q., Gao, C., Rivero, C.P., Webb, C., Lin, Carol Sze Ki, Venus, J., 2016. Valorization of organic residues for the production of added value chemicals: a contribution to the bio-based economy. Biochem. Eng. J. (accepted manuscript) Sakai, K., Ezaki, Y., 2006. Open L-lactic acid fermentation of food refuse using thermophilic Bacillus coagulans and fluorescence in situ hybridization analysis of microflora. J. Biosci. Bioeng. 101 (6), 457–463. Sayeki, M., Kitagawa, T., Matsumoto, M., Nishiyama, A., Miyoshi, K., Mochizuki, M., Takasu, A., Abe, A., 2001. Chemical composition and energy value of dried meal from food waste as feedstuff in swine and cattle. Anim. Sci. J. Shirota, M., Aso, K., Iwabuchi, A., 1966. Study on the intestinal microflora: effect of the administration of Lactobacillus acidophilus strain Shirota on the composition of the intestinal microflora of healthy children (in Japanese). Jpn. J. Bacteriol. 21 (5), 274–283. Tashiro, Y., Matsumoto, H., Miyamoto, H., Okugawa, Y., Pramod, P., Miyamoto, H., Sakai, K., 2013. A novel production process for optically pure L-lactic acid from kitchen refuse using a bacterial consortium at high temperatures. Bioresour. Technol. 146, 672–681. Uçkun Kıran, E., Trzcinski, A.P., Liu, Y., 2015. Platform chemical production from food wastes using a biorefinery concept. J. Chem. Technol. Biotechnol. 90 (8), 1364–1379. Venus, J., 2006. Utilization of renewables for lactic acid fermentation. Biotechnol. J. 1 (12), 1428–1432. Venus, J., Richter, K., 2006. Production of lactic acid from barley: strain selection, phenotypic and medium optimization. Eng. Life Sci. 6 (5), 492–500. Wang, Q., Wang, X., Wang, X., Ma, H., Ren, N., 2005. Bioconversion of kitchen garbage to lactic acid by two wild strains of Lactobacillus species. J. Environ. Sci. Health Part A 40 (10), 1951–1962. Wang, X.Q., Wang, Q.H., Zhi Ma, H., Yin, W., 2009. Lactic acid fermentation of food waste using integrated glucoamylase production. J. Chem. Technol. Biotechnol. 84 (1), 139–143. Wang, X., Wang, Q., Wang, X., Ma, H., 2011. Effect of different fermentation parameters on lactic acid production from kitchen waste by Lactobacillus TY50. Chem. Biochem. Eng. Q. 25 (4), 433–438. Yang, X., Zhu, M., Huang, X., Lin, C.S.K., Wang, J., Li, S., 2015. Valorisation of mixed bakery waste in non-sterilized fermentation for L-lactic acid production by an evolved Thermoanaerobacterium sp. strain. Bioresour. Technol. 198, 47–54. Zhang, A.Y., Sun, Z., Leung, C.C.J., Han, W., Lau, K.Y., Li, M., Lin, C.S.K., 2013. Valorisation of bakery waste for succinic acid production. Green Chem. 15 (3), 690–695.