THE EFFECT OF LACTIC ACID BACTERIA AND DIFFERENT LEVEL OF CARBOHYDRATE SOURCES ADDITION ON TOFU WASTE INDUSTRY FERMENTATION

The purpose of this research was to figure out the affinity value of soluble carbohydrates from pollard with lactic acid bacteria (LAB) and its effect on tofu waste silage. Lactic acid bacteria (LAB) isolates used as inoculum were commerce LAB isolates (B13-1) and yolk LAB isolates (K6-3). The selected of LAB isolates used in the tofu waste silage were tofu waste (TW) and pollard (P) with different proportion (70:30), (60:40), and (55:45). The result showed that the addition of soluble carbohydrates sources in the tofu waste silage did not affect lactic acid and pH value. However, LAB addition significantly increased lactic acid (P<0.05). The TW:P proportion (55:45) produced the highest lactic acid concentration with the value of 3.54%DM with pH value 3.90. Proportion of TW:P gave significant effect on dry matter (DM) and organic matter (OM) (P<0.01). Meanwhile, LAB addition presented significant difference in declining of dry matter percentage (P<0.01) but it showed non-significant effect on organic matter percentage from the tofu curd silage. It can be concluded that pollard addition with the value of 45% shows that tofu waste silage has the highest lactic acid concentration and ideal pH value. (


Introduction
Semi-intensive and intensive ruminant production in Indonesia is characterized by a high demand and dependencyy of concentrate (mixed cereals) which are expensive.This makes feeds to be high, which represents 60-70% of the economic input in the livestock production system.The optimal utilization of local natural resources is a strategic step in achieving business efficiency ruminant livestock production in Indonesia.The raw materials generally associated with industrial-waste processing industry are the primary raw materials of agricultural origin.Nkosi et al. (2010), feed cost and animal competition with humans for feed item, strongly suggest that alternative energy sources such as food by-product is used partially or totally to replace cereals in livestock diet to reduce and enhance cheaper meat production.
Not only could the by-products be utilized as a source of nutrients for ruminants, but they can also be used to replace imported commercial feedstuffs to save energy in transportation, and, possibly, to reduce the environmental impact of burning them as waste of burying them in landfills.Xu et al. (2001) reported that food by-product such as tofu waste is high in crude protein and fatty acid.Katayama (2001) and Osaka (2001) implied that many food by-products are high in moisture content and often stored by ensiling them to avoid the energy costs of drying.Although some wet by-products can be ensiled without any additives, much cost is wasted on transporting water rather than nutrients.Imai (2001) stated that preparing total mixed ration (TMR) silage is one practice where by food by-products are stored and utilized as animal feeds in Japan.Our previous study (Cao et al., 2009) shows that TMR silage with 30% dried tofu waste has the higher lactic acid content than that with rice bran or green tea waste.Segawa (1991) reported that in tofu curd mixed with rice straw produces good silage and is ready to use by livestock in Japan.
However, even if there is plenty of glucose as a substrate, if it is insufficient in lactic acid bacteria, preparing good quality silage is difficult.Many researchers report that molasses addition can stimulate fermentation by supplying substrates for the growth of lactic acid bacteria (Guo et al., 2014).Cai et al. (1999) reported that the factors involved in fermentation quality include chemical composition, particularly the water-soluble carbohydrates content of the silage material, and physiological properties of epiphytic bacteria.
During ensiling, lactic acid bacteria (LAB) is used as additives to increase the preservation of nutritive value of feedstuff and enzyme activity, and to inhibited deleterious epiphytic microbial populations.LAB isolates (B13-1) and yolk LAB isolates (K6-3), inoculated in this study, are facultative homofermentative LAB, and able to ferment a wide variety of substrates and quickly produce large amounts of lactic acid.The purpose of this experiment was to examine the effects of LAB inoculant on the tofu curd silage, and to figure out the affinity value of soluble carbohydrates from pollard with LAB and its effect on tofu curd silage.

Ensiling
Composition of tofu waste silage before being fermented shows at Tabel 1. Curd waste silage was mixed with tofu waste (TW) and pollard (P) as the main ingredient with different proportion.The tofu waste silage was TW and P with different proportion (70:30), (60:40), and (55:45).Targeted DM tofu waste silage was respectively 30 to 45% of DM.The experiment was ensiled with or without LAB inoculant.The inoculant used in this study were, LAB isolates (B13-1), and LAB isolates (K6-3), and those isolate were homofermentative LAB.The persentage of the inoculants appliedd 10% of fresh tofu waste silage.The tofu waste silage was stored in vacuum-packed plastic pouches.Fresh by-products or tofu curd silage mixtures (300 g) were put into a pouch, and an airtight silo was prepared by using a vacuum sealer.Respectively, Silage was made in triplicate and stored at ambient temperature at 25°C for 21 days.

Chemical analysis
Moisture, ash, crude protein, ether extract, and crude fibre contents were determined by general methods.The pH of the silage was determined using a glass electrode of pH meter (D-21, Horiba, Kyoto, Japan).Lactic acid was analysed using the method of Barker and Summerson (Hawk et al., 1954).

Statistical analysis
Data of pH value, lactic acid production of tofu curd silage, and dry matter (DM) and organic matter (OM) of tofu curd silage opened after day 21 was analysed using a completely randomized design with 3 × 3 (variety of proportion × additive treatment) factorial treatment structure.

pH value
Fermentation process on tofu curd silage causes pH value to be low.pH value measurement resulted in tofu curd silage is presented in Table 2.
Based on the result presented in Table 2, the different proportion of dissolved carbohydrate source showed a very significant effect (P>0.01) on pH value changes, and the addition of LAB showed also significant effect (P<0.05) whereas the interaction between those two treatments didn't show a significant effect.All treatments reached the critical pH value with a mean 3.80 to 3.90 simultaneously on day 21.It shows that all treatments produced successful tofu curd silage because of the formation of the LAB.Meeske et al. (2002) showed that the fermentation of corn (maize silage) with addition of lactic acid bacterial inoculants gives a decrease of pH value more than what is expected -which is about 3.90 to 3.85.Gao et al. (2008) reported that, after 30 days of fermentation, the acidic smell formed by straw fermentation using lactic acid bacteria with pH 3.8 is lower than the control (4.1).Danner et al. (2003) implied that corn silage with addition of homo-fermentative bacteria has about 3.8 of pH value and lactic concentration between 31.9 to 35.4 g/kg, while the corn silage with addition of hetero-fermentative bacteria has the highest pH value which is 4.11.

Lactid acid production
Based on the result presented in Table 3, it can be concluded that the proportion of dissolved carbohydrate in tofu curd silage did not give effect on the level of lactic acid (P<0.05),but the three treatments gave different result of lactic acid production.The proportion of tofu curd and pollard (55:45) produced lactic acid with the highest average which is 3.54% BK.However, other results present that the addition of LAB significantly affected the levels of lactic acid (P<0.05).High content of BETN was more susceptible to degradee resulting in a higher lactic acid production.The result showed that the proportion BETN tofu curd and pollard (55:45) by 60.17% higher lactate production compared to (70:30) by 53% and (60:40) by 58.10%.Hartadi et al. (1999) reported that the extract without nitrogen-containing mono-, di-, tri-and tetra-saccharides and starch added by some material substances includes hemicellulose.Molin (2008) reported that L. plantarum is a homo-fermentative bacteria which is able to utilize 100% of carbohydrates in the form of glucose or other simple sugar.The more complex sugar bonding that occurs in carbohydrate, the lower capability of L. Plantarum to utilize these carbohydrate.
The addition of soluble carbohydrate can improve the quality of fermented silage using LAB.Hhigher concentration of soluble carbohydrate will increase the production of lactic acid, and lower the propionic acid and butyric acid.At an excessive concentration of dissolved carbohydrates, production of lactic acid will be saturated.High concentration of lactic acid can reduce the loss of crude protein which occurs during the ensilage process (Yang et al., 2006).Lactic acid levels in total mixed ration silage (TMR) which uses tofu curd for 14 days of incubation by 1.9%, while BK for 56 days of incubation produce lactic acid levels by 2.53% BK.Total mixed    ration (TMR) silage which uses beer dregs produces lactic acid levels by 1.15% during the 14 day incubation, while BK for 56 day incubation produces lactic acid level by 1.56% BK (Wang and Nishino, 2008).

Dry matter (DM) and organic matter (OM) on tofu curd silage
The result of observation of dry matter and organic matter in tofu curd silage at 21d was presented in Table 4 and 5.
The result presented in Table 4 shows that the difference of soluble carbohydrate source proportion and addition of LAB significantly affects DM of tofu curd silage (P<0.01) with significant interaction (P<0.05).Addition of bacteria reduces percentage of DM of tofu curd silage significantly.The lowest reduction of percentage of DM occurs on k 6-3 treatment (34.77%), B13-1 (35.46%), and control (37.21%).The highest percentage of DM reduction occurred on K-63 treatment proves bacteria ability in optimal utilization of soluble carbohydrate.Based on the result presented in table 5, it shows that proportion of soluble carbohydrate gives significant effect on OM of tofu curd silage (P<0.01).However, addition of LAB does not affect the percentage of OM of tofu curd silage (P<0.05).
Some hydrolysis process which break hemicellulose into pentose occurs during fermentation process (McDonald et al., 2002).Liu et al. (2009) reported that quality of fermentation product which has moderate KA (60.24%) is better than fermentation product which has high KA (73.68%) and low KA (28.88%).

Conclusion
It can be concluded that pollard addition with the value of 45% shows that tofu curd silage has the highest lactic acid concentration with ideal pH value, and has the highest dry matter.Whereas, addition of LAB increases lactic acid significantly.The tofu curd silage having some beneficial could be included as component of ration in livestock industry.

Table 1 .
Composition of tofu curd silage before being fermented : Nutritional Biochemistry Laboratory Faculty of Animal Science Gadjah Mada University. Author

Table 2 .
pH value on tofu curd silage

Table 4 .
Percentage of dry matter (%DM) on tofu curd silage Means in the same column with different superscripts (P<0.01).x,y,zMeans in the same row with different superscripts (P<0.01).

Table 5 .
Percentage organic matter (OM) of %DM on tofu curd silage x,y Means in the same row with different superscripts (P<0.01).