Result and discussion

The Total Organic carbon content of the medium (KB)

Organic compounds are oxidized by potassium dichromate (K2Cr2O7) and
concentrated sulfuric acid (H2SO4). The reaction is as follows:

Based on the results noted that the total organic carbon content in water washing rice greater
than broth chicken intestine pieces. This is in accordance with the views that the water
contains dissolved carbohydrates in rice [19], vitamin B1, and B12 [20], while the source of
carbon from the broth chicken intestines can come from dissolved protein [21].
Table 1. The Total Organic Carbon Content Of The Medium
Growing Bacillus sp.

Kadar C organik total

Kaldu Usus Ayam Potong Air Cucian Beras

Medium Luria Bertani

0,156 (%) (0,032)

0,502 (%) (0,067)

0,784 (%) (0,072)

The Total Nitrogen content of the medium (KB)

The process of destruction on nitrogen element change sample be ion ammonium
(NH4 +) and bind to ion sulfate (SO42-) to form ammonium sulphate. As for the organic
element is transformed into CO2 and H2O. Distillation process revamp of sulfuric gas into
ammonia ammonium by the addition of NaOH.

The results of the research conducted is known that nitrogen broth chicken intestine
three times larger than the water washing the rice. This is due to the animal's organs are
generally made up of the protein as it does on the intestine [21]. The nitrogen content is also
found in water washing the rice. Although the water washing rice contains a lot of dissolved
carbohydrates, there are still content of nitrogen derived from vitamin B1, B12, as well as
inorganic nitrogen [20].
Table 2. The Total Nitrogen Content Of The Medium Grow
Bacillus sp.

Kadar Nitrogen Total

Kaldu Usus Ayam Potong Air Cucian Beras

Medium Luria Bertani

0,185 (%)

0,080 (%)

0,223 (%)

( 0,016)

( 0,016)

( 0,014)

Gram Staining Bacterial Isolates

Bacillus sp. including Gram-positive bacteria and this unconfirmed on the staining
test was conducted. Staining Bacillus SP. isolates from results of staining was observed under
the light microscope with a magnification of 40 and 400 times. (Picture 1. A) saw the lines
that caused the evaporation of the solvent in the process of fixation of the glass above the
preparations taking place in diskontinyu. Magnification 400 times (picture 1. B) shows the
number of bacteria is relatively dense and purple, this indicates bacterial isolates tested is a
species of Gram-positive bacteria and supporting data that the Bacillus sp including Grampositive bacteria.

Figure 1. a) results of staining microscope 40 x magnification

light. b) magnification 400 x.
KOH test 4% Bacterial Isolates
The test results with the staining was persuaded by KOH test 4% for the
determination of the type of gram bacteria. Lipid bilayer gram-negative would rupture by
KOH so that the contents of the cell out mixed with a solution of test. This led to the
viskosiatas test solution is increasing and appears to be slimy. Based on the research results
obtained that the test solution dilute isolates of bacteria origin, meaning abdomen termite soil
rejuvenated including Gram-positive group. Positive controls are not carried out due to the
absence of gram-negative bacteria of the collection.
The growth of Isolate Bacillus sp
Variation of mixed medium liquid broth chicken intestine cut and water washing of
rice include: 12:5; 1:4; 2:3; 3:2; 4:1; and 5:0. The results of the measurements show that the
stationary phase begins to look at the 16th after planting, which was not found to significantly
increase in absorbance. The best known growth on the mix volume medium KB (1:4). The
highest absorbance values at the 16th until the 20th became the basis of the determination.
Medium liquid water washing rice without adding the broth has a similar growth medium KB
(2:3), in terms of both its chemical liquid medium is different in terms of content of carbon
and nitrogen. This means that not all of the carbon and nitrogen utilized by bacteria to grow.
This is allegedly due to several factors such as carbon and nitrogen content of functionalist
functionalist, these allegations need to be strengthened by further research and not addressed
in this research because including the limitation problem.

Figure 2. Profile the growth of Bacillus SP. on variations

volume was medium at a temperature of 37 oC KB
Temperature of bacteria growth on these penilitian vary from 27 oC, oC, 37 and 47 c.
Research results indicate that optimum growth of Bacillus sp. liquid medium KB (1:4) at a
temperature of 37 c (Figure 3). These data correspond to the views that the optimum growth
temperature condition of Bacillus SP. i.e. 37 oC [22] [23].

Figure 3. Profile the growth of Bacillus SP. in medium

KB (1:4) at a temperature of 27 c, 37 c, and 47 oC

Figure 4. Isolates of bacteria growth profile of Bacillus sp.

in medium KB (1:4) compared to the
medium LB at a temperature of 37 oC
Comparison of the growth medium KB (1:4) and the medium LB turns out to have a
semblance of a graph as shown in (Figure 4). Although there is a difference between the
absorbance of the medium but the difference is not very significant with the difference in
absorbance ( 0.02). This data shows that artificial medium still has the prospect to replace
the medium LB as a medium growing Bacillus sp.
Levels Of Protein
Protein levels (per ml) medium supernatan KB (1:4) is greater than on medium LB.
This means that an enzyme produced medium KB more. In terms of the growth rate of
bacteria Bacillus sp on medium LB better than liquid medium (1:4). This is allegedly because
most of the carbon and nitrogen on medium LB used to form the bacterial cells, while in
medium KB (1:4) only a handful of carbon and nitrogen were used to form bacterial cells.
Table 3. The levels of the protein enzyme endo--1.4-xilanase

Kadar protein (g/ml )

Supernatan medium KB (1:4)

Supernatan medium LB

2,600

1,421

( 0,412)

( 0,225)

Enzyme Activity
Based on the data obtained, the specific activity of medium KB (1:4) smaller than in
medium (LB), although larger protein levels. The existence of other enzyme contaminants
into the cause of the small value of the enzyme activity in medium KB (1:4). However, in
terms of a medium protein KB (1:4), the activity can be increased when done the process of
purification of the enzyme.

Table 4. The enzyme activity of endo--1.4-xilanase

Aktivitas enzim (U/ml)

Aktivitas spesifik (U/g )

Supernatan medium KB (1:4)

Supernatan medium LB

0,210

0,152

( 0,016)

( 0,020)

0,080

0,11

Molecular Weight Of The Enzyme

Based on the results of sodium dodecyl sulfate (SDS-PAGE) samples of the enzyme
that is done, it can be explained that the medium broth chicken intestine apparently has a
protein content with the emergence of a peak between 25 to 45 kDa (Figure 5). For water
washing rice not found the existence of a peak because it is largely made up of melekul
dissolved carbohydrates. A similar case is also available on medium LB liquid without the
bacteria, even though the content of nitrogen liquid LB greater than medium broth gut, it
didn't found the existence of a peak. This case can be explained because the medium LB
contains material of nitrogen in the form of amino acids and does not bind to the stationary
phase, when the process of electrophoresis in progress.
The results of the produsi enzymes from medium KB (1:4) and LB peak appears
between turns 25 to 66 kDa. This is due to the enzyme still contain other protein
contaminants. However, based on the results of the SDS-PAGE is found the existence of a
peak among 45 to 66.2 kDa correspond to the results of the SDS-PAGE (Figure 5). This is in
accordance with the study mentioned that the enzyme endo--1.4-xilanase been 45000 Dalton
to 66200 Dalton [24].

Figure 5. The results of the SDS-PAGE A) bowel broth chicken pieces;

B water washing rice); C) medium KB (1:4); D)

liquid LB medium; M) marker proteins; X) crude

enzyme from the medium LB; Y) crude enzyme from
medium KB