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Bioconversion of rice straw into improved fodder for cattle


Preliminary treatments
The present investigation


Usha George and T.K. Ghose

Biochemical Engineering Research Centre, Indian Institute of Technology, Hauz Khas, New Delhi, India

The 38.6 million hectares of Indian agricultural land under rice cultivation in 1977 yielded 42.8 million tons of grain (1) and, as by-products, about 81 million tons of agricultural residues - namely, 66 million tons of rice straw and 15 million tons of husks. The cellulosic and hemicellulosic components of these residues (cf. table 1) are potentially available for saccharification or bioconversion to microbial biomass as an improved feed supplement. Research into the use of these residues is being conducted in our laboratory.

In developed countries these residues are disposed of mostly by burning, but now, there being increased interest in recycling these materials that are available in such large amounts, attention is turning toward making better use of them. The traditional uses of rice husks in India have exploited their calorific and abrasive qualities, and they are now being considered for the prediction of good-quality construction materials, while rice straw finds use as fuel, thatch, packing material, and as cattle feed (3; 4).

TABLE 1. Average Composition of Rice Straw and Rice Husk

 

Composition (%)

Constituent

 

Rice strawa

Rice huskb

Cellulose

43

35

Hemicellulose

25

25

Lignin

12

20

Crude protein (N x 6.25)

3-4

3

Ash

16-17 (silica 83%)

17 (silica 94%)

a. Linko (2).
b. NCST (3).

 

 

TABLE 2. Energy and Protein Requirements of Cattle and Availability in Rice Husk and Straw

 

Requirementa

Availabilityb

 

Adult (maintenance)

Calf

Pregnant cow

Lactating cow

Per kg rice husk

Per kg rice straw

Digestible energy (x 103 kcal/day)

7.5-12.4

7.5-22.9

10.5-20.6

16.5-49.9

1.28

1.97

Digestible crude protein (g/day)

110-150

250-410

30-260

240 - 1,660

239

-

a. Adapted from Cuthbertson (6)
b. Ranjhan (5).

Forty-two per cent of the adult cattle in India, employed primarily for draught power, are fed and maintained almost entirely on rice straw; it is also fed to cattle of other age groups with supplements. Quite a number of studies are being carried out in India on the use of other vegetable wastes as feed supolements (5). It is therefore of interest to compare the energy and protein requirements of cattle with the energy and protein available in rice straw (table 2). It is evident that both straw and husk, if untreated or unsupplemented, are not of adequate quality even as maintenance rations. The poor nutritive value results from the resistance of these materials to enzymatic attack by rumen microorganisms. This resistance can be correlated to ( i ) the degree of lignification (7; 8), (ii) the crystallinity of the cellulosic component (9; 10), and (iii) the high silica content (11). In addition, the abrasive quality of rice husks makes ingestion of untreated materials dangerous. Treatment of these materials before feeding to animals is essential.

 

Preliminary treatments


Non-biological treatments have been used to render straws and other lignocellulosic materials more digestible for cattle (12; 13), or more susceptible to microbial or enzymatic attack (12). Briefly, physical treatments include: (i) size reduction, which increases the surface area of cellulosic and lignocellulosic residues, decreases the crystallinity (14; 15), and therefore increases the susceptibility to chemical action (16) or enzymatic attack (fig.1) (10; 14); ( ii ) moist heat treatment, resulting in thermal hydrolysis and caramelization of sugars (17); (iii) ultraviolet, gamma, or electron irradiation, which lowers the degree of polymerization of cellulose and lignin and partially disrupts the lignocellulosic complex (8). Generally, however, these methods are energyintensive and uneconomical on a large scale.

FIG. 1. Relationship between Lignin Content and In Vitro Digestibility for NaOH-Treated Hardwoods

Chemical treatments, particularly NaOH treatment, have been used successfully (8; 16; 18). These include the use of (i) alkali and (ii) oxidizing agents such as SO2, NaClO2 (or ClO2 ), H2O2, O3, etc. The alkali causes swelling and separation of the cellulose, partial removal of lignin, lowering of the crystallinity of the cellulosic fraction, and partial hydrolysis of the hemicellulose (8; 18). Hot alkali removes a larger portion of the lignin as well as the hemicellulose (Biochemical Engineering Research Centre [BERC], Indian Institute of Technology, Delhi, unpublished data), but such a treatment appears to increase the crystallinity of the cellulose (19). Oxidizing agents act by disruption of bonds in the lignocellulosic complex and within the cellusosic fraction (8).


The present investigation


TABLE 3. Yields of Microbial Protein Derived from Lignocellulosic Materials

Substrate

Treatment

Organisms

Maximum Protein

Yield (%)

Rice straw

alkali

Cellulomonas sp. and Alcaligenes faecalis

20a

Paper mill waste fibre

-

S. pulverulentum

13.8b

Barley straw

alkali

T. reesei

25.8c

Sawdust

acid

Ch. cellulolyticum

21d

Sawdust

-

Ch. cellulolyticum

22.2e

Bleached kraft pulp

-

Ch. cellulolyticum

21.2e

Wheat straw

alkali

Cochliobolus specifier and other fungi

13.9f

Rice straw and potato peelings

-

S. pulverulentum

12.5g

a. Han (20)
b. Eriksson and Larsson (21).
c. Peitersen (22).
d. Moo-Young et al. (23).
e. Pamment et al, (24).
f. Chalal et al. (25).
g. Ghose, George, and Selvam, unpublished data (BERC, 1979)

Chemical treatment has also been used to render lignocelluiosic materials susceptible to microbial attack (table 3). Work is being done in our laboratory to develop a low-technology process for the conversion of rice straw to improved fodder. It is particularly useful and highly desirable to use such a fodder in Indian villages. A lignocellulosic fungus strain of Sporotrichum pulverulentum, with a well-defined enzymatic capacity (26; 27) and having a favourable amino acid composition of its potential biomass (26; 28), was chosen for this preliminary work.

The effect of mild chemical treatments on protein production of S. pulverulentum grown on rice straw was studied. The treatment and growth conditions are shown in table 4 and the results in table 5. Very poor growth (mostly in the form of spores) and production of protein were observed. Another series of experiments was conducted, therefore, to study the effect of a supplementary carbon source (cane molasses) and slightly different preliminary treatments on protein production by S. pulverulentum. The treatments were similar to those shown in table 4 with some modifications. Instead of the "dry" caustic treatment, as reported by Wilson and Pigden (18), a partially de-lignified straw was prepared by autoclaving 10 9 of straw with 3 9 NaOH in 100 ml of water for one hour. Following the treatments, the caustic or acid was removed by washing until the wash water was neutral. The medium used contained 0.1 g cane molasses, 0.1 9 (NH4)2SO4 per 10 ml distilled water, 2 9 straw, and 10 ml medium in a 250 ml Erlenmeyer flask inoculated for five days at 37 C. Three flasks and one control were used for each treatment. In addition, protein production by an Aspergillus sp. isolated from rice straw was also studied in the same manner. The results are shown in table 6.

TABLE 4. Treatment of Rice Straw and Growth Conditions for Protein Production ( Kjeldahl ) by Sporotrichum pulverulentum

 

Treatment
 

1a

2b

3

Chemical

NaOH

NaOH

H2SO4

g chemical /100 g straw

6

6

1.6

Ratio of water to straw

1:4

6:1

3:1

Incubation

30 C, 24 h

100C, 15 min

100 C, 30 min

pH adjustment

washing

washing

5 % NH3

Final pH

pH 7

pH 7

pH 4.5-5.0

Nutrients

     

(g/40 ml water/20 g straw)

     

(NH4)2SO4

1.2

1.2

-

KH2PO4

0.75

0.75

0.75

MgSO4,7H2O

0.12

0.12

0.12

Inoculum

mycelial fragments of S. pulverulentum

Incubation

37C, 7 days

a. Wilson and Pigder (18).
b. Han and Callihan (29).

TABLE 5. Effect of Various Treatments of Rice Straw on Protein Production by S. pulverulentum

 

Mg Protein (N x 6.25)/g Straw

Treatment
 

Uninoculated

Inoculated

 

control

 

Untreated

26

28

NaOH (spray),0.06 g/g strew

25

27

NaOH (100 C), 0.06 g/g straw

32

33

H2SO4 (100C), 0.016 g/g straw

22

29

 

TABLE 6. Effect of Treatment of Rice Straw on Protein Production by Aspergillus sp. and S. pulverulentum with Molasses as Supplementary Carbon Source

Mg Protein (N x 6.25)/g Straw

 

Aspergillus sp.

S. pulverulentum

Treatment
 

Uninoculated control

Inoculated straw

Uninoculated control

Inoculated straw

Untreated

35

47

33

58

NaOH (100C), 0.06 g/g straw

45

49

36

67

NaOH (120C), 0.3 g/g straw

25

39

32

66

H2SO4 (100C), 0.016 g/g straw

36

44

32

64

Aspergillus sp. produced little protein and further studies with this organism were suspended. S. pulverulentum did show a little increase in protein, even in untreated straw. There was little difference in protein production in the straw following different types of preliminary treatments, and not enough increase over untreasted straw was observed to justify the higher cost

Further studies were conducted with untreated straw. The effect of starch and potato peelings or an infusion from potato peelings (10 g peelings, 50 ml distilled water, boiled for 30 minutes) was explored. These experiments were performed in a manner similar to those using molasses, but with the substitution of 0.1 g starch or potato peelings for molasses. In the case of the infusion, 0.1 g (NH4)2SO4 was added to 10 ml infusion and no other carbon source was used. The results are shown in table 7. The results obtained with potato peelings are probably due to the increased nitrogen content and perhaps to the presence of some growth factors.

TABLE 7. Effect of Starch, Potato Peelings, and Potato Peeling Infusion on Protein Production by S. pulverulentum in Rice Straw

Mg Protein/g Straw (Dry)

Supplement
 

Uninoculated control

Inoculated

Starch, 0.1 g/g straw

30

84

Potato peelings, 0.1 g/g strew

62

125

Infusion,a 5ml/g straw

54

88

BERC, unpublished data.
a. Prepared by boiling 10 g peelings with 50 ml distilled water for 30 minutes.

The protein yields obtained by certain investigators on lignocellulosic material are shown in table 3. Clearly, our protein yields are lower than those reported by others, but the fact that we have been using simple or no preliminary treatment and much simpler media (25) and yet obtaining enrichment of rice straw appears most encouraging. We have plans to screen several organisms with a view to ascertaining their ability to grow on untreated rice straw either singly or in mixed culture. We are hoping (on the basis of more recent data not reported here) to be able to develop a simple, inexpensive process for the bioconversion of rice straw into improved cattle fodder.

References

1. India, Ministry of Information and Broadcasting, India: A Reference Manual, 1977-1978 (New Delhi).

2. M. Linko, in Adv. Biochem. Eng., 5: 2511977).

3. NCST, Utilization and Recycling of Agricultural Wastes/By-products: A Country Report (New Delhi, 1974).

4. D.H. Grist, Rice, 5th ed. (Longmans, London, 1975), pp. 432-448.

5. S.K. Ranjhan, Animal Nutrition and Feeding Practices in India (Vikas Publishing House, New Delhi, 1977).

6. D. Cuthhertson, "Nutrient Requirements for Farm Livestock: 2. Ruminants," in D. Cuthbertson, ea., Nutrition of Animals of Agricultural Importance, part 2 (Pergamon Press, Oxford, UK, 1969), Pp. 883920.

7. W.C. Feist, A.J. Baker, and H. Tarkow, in J. Animal Sci., 30: 832 (1970).

8. M.A. Millet, A.J. Baker, and L.D, Slatter, in Biotech. Bioeng., 20: 107 (1978).

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12. Y.W. Han, in Adv. Appl. Microbial., 23: 119 (1978).

13. M.G. Jackson, Treating Rice Straw for Animal Feed ( FAO, Rome, 19781.

14. T.K. Ghose and J.A. Kostick, in Adv. Chem. Ser., 95: 415 (1969).

15. J. Nystrom, in Biotech. Bioeng. Symp., 5: 221 (1975).

16. Y.W. Han, W.P. Chen, and T.R. Miles, in Biotech. Bioeng., 20: 567 ( 1978).

17. N. Nesse, J. Wallick, and J,M, Harper, in Biotech, Bioeng., 19: 323 (1977).

18. R.K. Wilson and W.J. Pigden, in Canad. J. Animal Sci., 44: 122 (1966).

19. M. Tanaka, M. Taniguchi, T. Morita, R. Matsuno, and T. Kamibuko, in J Ferment, Technol., 57: 186 (1979).

20. Y.W. Han, in Appl. Microbiol., 29: 510 (1974).

21. K.-E. Eriksson and K. Larsson, in Biotech. Bioeng., 17: 327 (1975).

22. N. Peitersen, in Biotech. Bioeng., 17: 129 (1975).

23. M. Moo-Young et al., in Biotech. Bioeng., 20: 107 (1978).

24. N. Pamment et ah, in Biotech. Bioeng., 20: 1735 (1978).

25. I.D.S, Chalal, M. Moo-Young, and G.S. Dhillon, in Canad. J. Microbiol., 25: 793 (19791.

26. B. von Hofsten, "Cultivation of a Thermotolerant Basidiomycete," in G.G. Birch, K.J. Parker, and J.T. Worgan, eds, Food from Waste (Applied Science Publishers, London, 1976).

27. K-E. Eriksson, in Biotech. Bioeng., 20: 317 (1978).

28. B. von Hofsten and A.L. Ryden, in Biotech. Bioeng., 17: 1183 (1975),

29. Y.W. Han and C.D. Callihan, in Appl. Microbiol., 27: 159 (1974).