Castor bean cake as raw material for biodegradable material
Roseli Sengling Lacerdaa, Ana Mônica Quinta Barbosa Bittanteb, Hulda Chambib, Catarina Abdala Gomidea,
Izabel Cristina Freitas Moraesb, Rosemary Aparecida de Carvalhob, Paulo José do Amaral Sobralb
a
Animal Science Department, Faculty of Animal Science and Food Engineering, University of São Paulo,
Pirassununga, Brazil ([email protected])
b
Food Engineering Department, Faculty of Animal Science and Food Engineering, University of São Paulo,
Pirassununga, Brazil ([email protected])
ABSTRACT
The valorization of castor bean cake could contribute to the success of the production of biodiesel from
castor bean. Thus, the objective of this work was to study the potential of castor bean cake as a protein source
of interest for the production of biodegradable material. For the characterization of the raw material, the
castor bean was submitted to classical chemical analyses. And, the protein extractions tests were conducted at
different conditions of pH, temperature, concentration of the cake in the extraction solution and agitation
speed. The protein extracts (supernatant) were separated from the solid residues (decanting) by centrifugation
(4000rpm for 20 minutes) and analyzed for protein, dry matter and ash. The castor bean cake had high
protein content (40%) and crude fiber (29%) compared to other components (ash, lignin, cellulose,
hemicelluloses and lipids). The cake might be considered rich in calcium (1.5%), potassium (0.8%),
phosphorous (0.6%) and magnesium (0.5%) compared to other minerals (Na, S, B, Zn, Mn, Cu and Fe). The
processes of protein extraction showed yields between 16 and 38%, based on the initial content of protein in
the cake. In general, the increase in pH contributed to the increase in the yield of protein extraction from the
castor bean cake. Furthermore, higher yields were obtained in the extraction with low concentration of cake
in the extraction solution. An analysis electrophoresis of the extracted protein showed the presence of
albumin and globulin, proteins widely used in biodegradable materials technology. It can be concluded that
castor bean cake is a good source of protein for the industry of biodegradable materials, and that the
extraction of these proteins continues to generate considerable residue with high protein content, which may
be considered for use as fertilizer or, if detoxified, as a component of animal feeds.
Keywords: Protein; extraction yield; film biodegradable; renewable resource.
INTRODUCTION
Castor bean has recently been highly rated as a source of raw material (oil) for biodiesel production, because
beyond its high oil content (25 – 55%), it is a culture of great social appeal in Brazil by intensive use of
workmanship in the field and allows for intercropping with other crops as beans, groundnuts or maize [1,2].
In addition, castor bean cultivation is encouraged in areas of low water availability and is genetically
improved to produce biofuel.
Castor bean cake is the main by-product of castor oil production. Pressing one ton of castor beans for oil
extraction produces around 550 kg of cake, but this value can vary with seed oil content and oil extraction
process [4]. Castor bean cake can be used as fertilizer, fungicide, in plant-parasitic nematode control, in
animal feed (if detoxified), in recovery of depleted soils and as raw material for ethanol production [4, 5, 6].
Also, the cake may be used as filler to obtain composite material prepared from widespread polymers, such
as high impact polystyrene, low density polyethylene, polypropylene and polyhydroxyalkanoate [7].
Castor bean cake is a product with high protein content (28 – 43%), composed of globulins (60%), albumins
(16%), proteoses (4%), glutelins (20%), conjugated proteins and non-protein nitrogen compounds [4]. This
composition makes the castor bean proteins a good raw material for biodegradable material production,
because many of these materials have been developed with similar protein fractions [8].
The objectives of this work were to determine the proximate composition and mineral profile of castor bean
cake produced by a Brazilian industry, and extract proteins for the production of biodegradable materials
(films) to be used in agriculture. Therefore, the valorization of this by-product could contribute to the success
of the biodiesel production from castor beans. In addition, the residues generated during protein extraction
would still be used, either as fertilizer, or in animal feed, depending on its toxicity.
MATERIALS & METHODS
The castor bean cake was donated by the company A. Azevedo Indústria e Com. de Óleos Ltda. (Itupeva/SP
– Brazil). For the characterization of the raw material, the cake was submitted to classical chemical analyses
[9, 10, 11, 12].
For protein extractions tests, first, the cake was ground and separated into three fractions (coarse,
intermediate and fine) using sieves of 12-48 mesh. These fractions were analyzed for protein, lipids, crude
fiber, ash and moisture [9].
The extraction of the proteins was studied by solubilization in an alkaline medium under different conditions
of pH (8, 9, 10 and 12), temperature (30 and 50ºC), concentration of the cake in the extraction solution (10
and 20%), and agitation speed (400 and 600 rpm). The protein extracts (supernant) were separated from the
solid residues (decanting) by centrifugation (4000 rpm/20 minutes) and analyzed for protein, dry matter and
ash [9]. The molecular mass profile of lyophilized protein was evaluated using SDS-polyacrylamide gel
electrophoresis (SDS-PAGE) according to Laemmli [13].
The experimental results were evaluated by ANOVA and Tukey’s multiple test using ‘Statistical Analysis
System’ computer programme version 8.0.
RESULTS & DISCUSSION
The castor bean cake presented a high protein (40%) and fiber (29%) content (Table 1), and may in principle
have a good indication for use as animal feed, if there was no problem of toxicity. There are several studies in
literature concerning the detoxification of castor bean cake [14] and its use in animal feed. Diniz et al. [15]
concluded that castor bean cake treated with calcium oxide can replace all or part of the soy cake used in
diets for young bulls. Moreover, the cake might be considered rich in calcium (1.5%), potassium (0.8%),
phosphorous (0.6%) and magnesium (0.5%) compared to other minerals (Na, S, B, Zn, Mn, Cu and Fe)
(Table 1). According to Bose and Wanderley [16], castor bean cake is one of the best fertilizers when
compared to other organic fertilizers such as bovine manure, mixed manure and cottonseed meal.
The castor bean cake also presented high values of NDF and ADF (Table 1), and these values should be
considered in formulations of animal diets. As NDF and ADF increase, dry matter intake and digestibility of
forage generally decrease, respectively. The cake is a product slightly acid (pH 5.9) and required a low
amount of liming to neutralize (acidity index of 0.95).
Table 1. Proximate composition and mineral content of whole castor bean cake.
Components
Content (%)1
Minerals
Content (%)1
Moisture
8.76 ± 0.13
Iron
0.1909
0.0004
Crude protein
39.78 ± 1.06
Zinc
0.012
0.0003
Lipids
3.05 ± 0.1
Manganese
0.0073
0.0006
Ash
10.33 ± 0.01
Copper
0.0037
0.0005
Crude fiber
28.84 ± 0.83
Sodium
0.029
0.004
NDF
49.44 ± 1.09
Magnesium
0.46
0.01
ADF
40.58 ± 0.43
Potassium
0.75
0.04
Lignin (H2SO4)
23.54 ± 0.38
Calcium
1.54
0.06
Phosphorus
0.55
0.02
Cellulose
10.45 ± 0.25
Hemicellulose
9.51 ± 1.06
NDF-N
0.86 ± 0.13
ADF-N
1.40 ± 0.10
NPN
1.80 ± 0.19
pH
5.91
0.00
Acidity Index2
0.95 0.00
1
Values expressed on dry basis, except moisture. 2 Value expressed in mg de NaOH/g of cake. NDF =
Neutral Detergent Fiber, ADF = Acid Detergent Fiber, NDF – F = Nitrogen in NDF, ADF – F = Nitrogen
in ADF, NPN = Non-Protein Nitrogen.
The castor bean cake showed a low residual oil (3%), and thus can be used as part of animal feed, if
detoxified. However, removal of residual oil may be necessary, in formulations with a high proportion of
cake, because the oil is not edible by animals. Absorption of the cake by soil is also facilitated when the oil is
present in low concentrations (1 – 1.5%) [4]. The moisture (8.8%) of the cake is considered satisfactory,
being favorable to storage.
According to analysis of the three fractions (coarse, intermediate and fine) of the cake, obtained after
grinding, only 3% of the proteins were in coarse fraction. So, this fraction was discarded. Thus, the protein
extraction tests were performed using the fine and intermediate fractions that make up 95% of whole castor
bean cake. The yields of extracted proteins ranged from 16 to 38%, based on initial protein content in the
cake. At pH 9, the highest extraction yield was obtained using low concentration of cake in the extraction
solution (Figure 1) compared to other extraction conditions studied. The agitation speed did not significantly
affect the extraction yield of protein.
Figure 1. Yield of extracted proteins from castor bean cake (agitation speed = 400rpm).
Extraction yield of castor bean cake proteins increased linearly with pH (Figure 2), however, increasing pH
up to 12 demanded a considerable consumption of sodium hydroxide (Figure 2), that should be considered in
the characterization of films obtained from protein extracted under these conditions. Studies on the extraction
of protein from red pepper seed, watermelon seed and pigeon pea [17, 18, 19] showed that increasing
solvent/cake ratio (decreasing of cake concentration in the extraction solution) and increasing pH resulted in
higher yields in the extraction of proteins, as observed in this study.
Figure 2. Yield of extracted proteins in relation to soda volume used at different extraction pH.
Protein and ash contents of liquid extracts obtained with different treatments ranged from 57 to 69% (dry
basis) and 8 to 24% (dry basis), respectively. The residues generated had protein and ash contents between 25
and 37% (dry basis) and 12 and 14% (dry basis), respectively, which suggests that these residues may still be
used as organic fertilizer or animal feed, as they present reasonable levels of protein.
An analysis by electrophoresis of extracted proteins showed three groups of molecular weight (MW) bands:
37 – 40 kDa, 20 – 23 kDa, 6 – 7 kDa, and traces of high molecular weight fractions (between 50 -90 kDa)
(Figure 3). The first two groups (20 – 40 kDa) were identified as 11S globulin with their dissociated subunits,
and 4 – 6S albumins (lectins) [20]. The bands of low MW (6 – 7 kDa) represent the subunits of 2S albumin
that typically have MW around 11 – 12 kDa [21, 22]. Thus, it can be assumed that the proteins extracted
from castor bean cake were composed mainly of albumin and globulin, which are proteins widely used in the
technology of biodegradable materials [8]. The two subunits of ricin (A and B of approximately 35 and 29
kDa, respectively) were not visualized and may be masked by large fractions, as observed in the Figure 3.
Figure 3. SDS-Page electrophoresis of the castor bean lyophilized protein extract. Conditions of the system: stacking gel
= 4% and running gel = 12%. Legend. MWM = Molecular weight marker (BioRad).
Samples of these proteins were tested in essays on films production. These films were prepared by casting a
film-forming solution with 7.5 g lyophilized protein/100 g of film-forming solution, 25 g of glycerol/100 g of
proteins, used as plasticizer, and 0.8 g of glutaraldehyde/g of proteins, used as crosslinker agent. The filmforming solutions where poured on plexi-glass support and dried at 30ºC/16h, in an oven with air circulation.
Thus, under these conditions, it was observed that the lyophilized proteins produced in this study were able to
produce films, as can be observed in the Figure 4.
Figure 4. Examples of films produced with lyophilized castor bean proteins extracted in pH = 11 (left) and 12 (right).
The produced films were homogeneous, relatively opaque and with a brown color; characteristics
appropriated for use in agriculture. The brown color was due possibly to Maillard reaction that can occur
during the extraction process at 50 ºC and under alkaline conditions.
CONCLUSION
Castor bean cake is a good source of protein for the industry of biodegradable materials, and the extraction of
these proteins continues to generate considerable residue with high protein content, which may be considered
for use as fertilizer or, if detoxified, as a component of animal feeds.
ACKNOWLEDGEMENTS
The authors acknowledge FAPESP for their financial support (08/11341-5) and for the Post Doctoral
fellowship provided to the third author (09/10172-8).
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