METHOD TO CREATE VALUABLE PRODUCT

Waste bread converted to succinic acid

A. awamori ATCC 14331 was used for the production of amylolytic enzymes. An industrial
strain of A. oryzae isolated from a soy sauce starter kindly provided by the Amoy Food Ltd.,
Hong Kong.
It was utilized to produce proteolysis enzymes. Their storage and sporulation for inoculum
preparation have been described in a previous publication. A. succinogenes ATCC 55618
were utilized for succinic acid fermentations.

Solid state fermentation (SSF)

Bread was cut into cubes of approximately 1 cm3 in size and autoclaved for 15 min prior to
cultivation. 100 grams of bread cubes were transferred into fermentation tank. Ten millilitre
of cryopreserved spores of A. awamori (2.85107 spores/mL) or A. oryzae (6.31106
spores/mL) were diluted by 10-folds and spread evenly on bread pieces separately. The
prepared samples were incubated at 30 C for 4 days.

Enzymatic hydrolysis
Two 2.5 L bioreactors equipped with automatic temperature control water jacket and stirrers
were used for enzymatic hydrolysis. Various amount of waste bread pieces were blended with
10 L water for 15 min. The resulting mixture was transferred into the bioreactors at 55 C.
Fungal marshes from SSF were added into the vessel. The reaction mixture pH was not
controlled and was stirred at 300 rpm. Samples were taken every hour for 24 h. The resultant
broth was centrifuged at 10,000 rpm for 15 min. The supernatant was subsequently filtered by
vacuum filtration using Whatman No.1 filter paper. All experiments were carried out in
duplicate.

Bacterial fermentation

Two 2.5 L fermentation tank (Biostat, Germany) were used for fermentative SA production
by A. succinogenes. Bacterial fermentation using bread hydrolysate was performed and the
inoculums size was 5%v/v. The initial glucose concentration of the broth was 40 g/L while
the free amino nitrogen (FAN) concentration was 200 mg/L. The latter corresponds to the
FAN content of a 40 g/L yeast extract solution. The hydrolysate was filtered by 2.0 _m PTFE
membrane filter (Sartorius, Germany). Prior to autoclaving, 100 g/L magnesium carbonate
(MgCO3) was added to the fermentation medium as a neutral pH buffer. The pH of the
fermentation broth was automatically controlled within 6.66.8 with the addition of 10 M
NaOH and 0.05 M H2SO4. The broth was sparged with 0.5 vvm CO2 and agitated at 300
rpm. Fermentation samples were taken every 3 h to measure optical density and metabolites
concentration. Fermentations were considered to have ended when glucose was completely
depleted.

ECONOMIC COSTING
1

Waste bread-The waste bread can be collected from bakery or bread factory. The only

costing is on transportation.
Solid waste fermentation- For the solid waste fermentation, a fermentation tank is
required. The price of a fermentation tank depends on its size. The range of price is

from US $500-15000 / Set.

Enzymatic hydrolysis- A bio-reactor with temperature control and stirrer costs around

US $2946-3682. For the whatman filter paper, it costs around US $1-100.

Bacterial fermentation- Another fermentation tank is required. The price for the
fermentation bacteria is around US $40-100 per kg

Process
Solid waste fermentation
Enzymatic hydrolysis

Bacterial fermentation
Operating costing

Item needed
i. Fermentation tank
i. Bio-reactor
ii. Enzyme
iii. Whatman filter paper
i. Fermentation tank
ii. Fermentation bacteria
i. Waste bread
ii. transportation
iii. Electricity
iv. Manpower

Estimated price
US $500-15000 per set
US $2946-3682 per set
US $55-200 per kg
US $1-100
US $500-15000 per set
US $40-100 per kg
US $0-10 per kg
US $2-4 per km
US $ 3751-4055 per month
US$ 775-1085 per month and
per person

Total= : US $ 8570-39236

The exchange rate of RM to US $ is 0.31, therefore

Minimum costing required is RM 27,645.16
Maximum costing required is RM 126,567.74

PRODUCT PROPERTIES AND MARKETABILITY

Product Properties
The properties of succinic acid are listed in the table below (J. T. Baker, 2008):
Table showed the physical properties of succinic acid
CAS Number
Molecular Formula
Molecular Weight
Melting Point

110-15-6
(CH2COOH)2
118.09 g/mol
188 oC

Boiling Point
Specific Gravity
Solubility
pKA1
pKA2

Decomposes at 235oC
1.56
100 g / 100 ml at 100oC
7.69 g / 100 ml at 25oC
4.21
5.64

Succinic acid can be used to create a large array of chemicals including resins and
biodegradable polymers, as well as surfactants, detergents, pharmaceuticals and food
additives [Lee et al., 2002, Lin et al., 2008]. It can also be used as an ingredient in animal
feeds and used to stimulate growth in plants [Wan et al., 2008].

Products Marketability
Succinic acid is a platform chemical that has a broad range of applications, from highvalue niche applications such as personal care products and food additives, to large volume
applications such as plasticizers, polyurethanes, resins and coatings. The possible applications
for succinic acid, which are expected to register strong demand growth in the near future, are
plasticizers, polyurethanes, bio plastics, and chemical intermediates. Currently there are only
a small number of companies which sell succinic acid as a merchant product. The current
price for petrol-based succinic acid oscillates between $ 2,400 / metric ton (MT) - $ 2,600
/MT and for bio-based succinic acid between $ 2,860 / MT - $ 3,000 / MT depending on the
supplier, quality and grade of the chemical.
The high cost of producing succinic acid from petroleum-based feedstock limited its
use to a narrow range of applications such as pharmaceuticals and food ingredients. As a
result, the global succinic acid market was estimated to 40,000 MT in 2011 and over 97% of
the succinic acid production was petrol-based. Currently, the main producers of succinic acid
are DSM, Gadiv Petrochemical Industries, Mitsubishi Chemical, Kawasaki Kasel Chemical,
Nippon Shokubai, along with several Chinese producers such as Anqing Hexing Chemical,
Lixing Chemical and Anhui Sunsing Chemicals, and some producers from India. The main
producers of bio-based succinic acid in 2011 were BioAmber (350 MT), Succinity (500 MT)
and Reverdia (300 MT). Succinic acid market is expected to reach approximately 699,449
MT in 2020, growing at a compound annual growth rate (CAGR) of 32.9% from 2010 to

2020. The market value in 2011 was estimated at $ 63.2 million and is expected to reach $
538.8 million in 2020 at a CAGR of 26% from 2010 to 2020.

ENVIRONMENTAL EFFECT
Definition
An environmental effect is the result of environmental impacts on human health and
welfare. The term is also used synonymously with environmental impact.
http://stats.oecd.org/glossary/detail.asp?ID=822
Good effect:

Fermentation do not pollute the environment.

Can reduce the environmental burden of food waste.
Succinic acid is a sustainable product.