Chapter - M

IDENTIFICATION OF FUNCTIONAL GROUPS IN 100% CPVC AND 1:l CD:CHVC

IDENTIFICATION OF FUNCTIONAL GROUPS IN 100%CPVC AND

1:lCD:CHVC

In this chapter functional groups present in vermicasts of 100% coir pith

(100%CPVC) and in the vermicasts of 1 1CD CH were identified through

FT-IR spectra The water extracts of these two vermicasts had inhibtted the radlal

growth of the two plant pathogens Rhlzoctonra solani and Rh~zopus stolonrfer

(chapter VII) Hence these two vermcasts - 100% CP VC and 1 1 CD CH VC - were

selected specifically t o identify the functional groups for their pathogen

suppressrve property

100% CP VC SPECTRA

The extract of 100%CPVC was allowed to pass through silica gel

column (chromatography) using acetone, chloroform, methanol and hexane as

solvents The elutes obtained are shown In the flow chart (Fig.51) These elutes

were then developed In thin layer ch romatography(TLC) The e lul tes

developing bands In TLC are shown In Table48 Wlth 100% methanol solvent

2"d jrd and 4* elutes developed bands in TLC having similar Rf value With

100% Hexane only 3'* and 8' elutes developed bands in TLC The elultes 1-8"

obtalned in I lHexane Methanol solvent and the elutes 51h and 61h obtained in

1 9 Hexane Methanol solvent developed bands in TLC having same Rf value

The only 3d elute to develop in TLC with 9 1 Hexane Methanol solvent also

has a similar Rf value as that of1 1 Hexane Methanol (Table 48) FTIR spectra

of those elutes wh~ch developed bands in TLC was taken in BOMEM MB 104

FT-IR spectrometer Three spectra were obtalned and a re presented in

F i g u r e 52. The spectrum obtained with 100%CPVC (without running

t h r o u g h column ch romatography) 1s a l so p re sen ted fo r compar i son

Figure 51: Flowchart of elutes obtained with 100% CPVC extract in Silica-Gel column chromatography using different organic solvents

100% CP Vermicast extract

11

1 100% Acetone 100% Chloroform 100% Hexane 100% Methanol

@ 7 8 9 10 6 7 8 9 10 1 I Hexane Methanol 1 9 Hexane Methanol 9 1 Hexane Methanol

Table 48: Rf value o f 100% coif plth-vennicast extract in TLC.

Figure 52: FT-IR Spectra of elutes of 100% CPVC that developed bands in TLC and 100% CPVC extract

SPECTRUM-I

In spectrum I (Figure 52) the slight sharp peak at 3391 29 cm" indicates

H bond in OH group Slightly broad small peak at 2364 13 cm-'depicts the P-in

CH Stretch 1742 05 cm-I, 1695 73 cm.' and 1645 9 cm.'-the three associated

broad peaks indicate the N-H band of amide group in open anion (Silverstein

and Webster, 1998) Two associated peaks at 1540.97 cm.' and 1576 92 cm"

indicate N-H bending v~bration The small peak at 1457 47 cm-' indicates the

CH stretch in ring structure 1360 9 cm-' in rlng structure indicates the CH

bending

SPECTRUM-11

In spectrum I 1 (Figure 52) the broad peak at 3397 4 cm-' Indicates the

H bond In OH group The small signal at 2361 22 cm-' ind~cates the phosphate

In CH Stretch and the well pronounced peak at 1645 81 cm-' ind~cates N-H

band in amide group, The two broad associated peaks at 1415 08 cm-' and

1354 69 cm-' are the N-H and CH stretches The peak at 1045 72 cm-' ind~cates

phosphate contain~ng C=O group (Chen et al. 1995)

SPECTRUM-111

The spectrum-11 and spectrum-I11 are nearly similar (Figure.52)

In spectrum-111 the 2928 16 cm-' and 2858 59 cm ' (wavelength) are the two

associated small peaks which are not well developed in spectrum-I1 They

Indicate the CH stretch The small signals at 1542 4 cm ', and 1512 47cm ' indicate the N-H bending of the amide group (Inbar et al , 1989) The small

peak at 821 537 cm.' ~ndicates the CH stretch in spectrum-111

Figure SJ: Flowchart of elutes obtained with 1 1 CDCHVC extract in Silica-Gel column chromatography using d~fferent organic solvents

1:1 CD : CH Vermicnst

1 2 3 4 5 6 7 8 @ @

1 1 Hexane Methanol 1 9 Hexane Methanol 9 1 Hexane Methanol

Table 49: Rf value of 1: 1 CD. CH vermicast extract in TLC

Figure 54: FT-IR Spectra Elutes of 1 1 CDCH VC that developed bands in TLC and 1 1 CD CH VC extract

Wavelength (4000-400 cm") 171

1: 1 CD:CH VERMICAST SPECTRA

T h e ex t r ac t o f 1 1 C D C H vermicast w a s run in co lumn

chromatography The elutes obtained with different solvents are shown in

Figure 53 These elutes were tested in thin layer chromatography (TLC)

The different bands obtained were noted (Table 49) for which FTIR spectrum

was obtained and is presented in Figure 54. Out of the 10 elutes obtained using

100%hexane as solvent only 3Id and 8" elutes have developed bands with TLC

W ~ t h 9 1 Hexane Methanol solvent 3r6 elute alone developed band in TLC

Their Rf-value is similar and the spectrum developed was also similar

(spectrum-I) The 5Ih elute obtained using 1 1 Hexane methanol solvent

mixture developed the spectrum-ll The 9" and 10" elutes obtained by using

1 9 Hexane Methanol solvent mixture developed bands in TLC w ~ t h the same

Rf value The elutes 4 ,5 ,6 ,7 ,and 8Ih -in 100% Methanol solvent a lso

developed bands in TLC and with same Rf value The spectra obtained

from the 9" and 10" elutes of 1 9 Hexane Methanol and 4Lh, Sth. 6", 7*, and

elutes of 100% Methanol were similar (spectrumI11)

SPECTRUM-I

3064 9 6 cm" wave length In spectrum-I (F igu re 54 ) depic ts t he

asymmetric CH stretch (Baes and Bloom, 1989) 2959 1 c m ' , 2930 72 cm"

and 2 8 7 0 2 cm- ' a r e t h e th ree na r row peaks i n d ~ c a t i n g CH-s t r e t ch

The 1729 74 c m ' indicates the amide NH-band (Silverstein and Webster,

1998) Sharp peak at 1461 57 cm-' is another asymmetric band of CH stretch

Both 1381 89 cm" and 1275 93 cm" bands are of CH stretch The sharp peak at

1125 23 cm-' and 1072 31 cm-' indicate the P - 0 bands 956 87 cm-' IS C - 0

g roup , and 743 82 cm.' and 701 78 cm.' both indicate the CH s t re tch

(Niemeyer et al . 1992)

In spetrum 11 (Figure 54) the broad peak at 3410 41 c m ' indicate

H bonds and OH group 2923 02 cm" with the associated peak at

2853 53 cm.' indicate CH bonds of CH stretch The well pronounced peak at

1602.12 cm" indicates the open chain of the amide band 1395 7 cm-' indicates

another C-H band of the CH stretch On 1043 63 cm-' the short peak indicates

the C=O group of the aromatic-compound (Hsu and Lo, 1999)

SPECTRUM-111

In spectrum-I11 (Figure 54) the slightly broad peak with the wave length

3441 43 cm-I is ~ndicative of the H band of OH group (Silverstein and Webster,

1998) The small peak at 3065 12 cm-' indtcates the CH stretch of the

aromatlc ring 2958 98 cm' . 2930 48 cm ', 2869 81 cm-' and 2866 43 cm-' are

well pronounced narrow assoc~ated peaks and they stand for CH stretch of

the aromatic rtng (Hsu and Lo.1999) 2336 83 c m l is the phosphate bond in

CH stretch The sharp peak at 1729 18 cm-' is the N-H band ofthe amide group

The two assoctated sharp peaks at 1599 71 cm-' and 1580 78 cm-' indicate

N-H bond in amide group The sharp peak at 1462 45 cm'l stands for the ring

structure of CH stretch 1276 14 cm" band wavelength indtcates phosphate

containing ring structure 1125 4 cm' , 1072 26 cm.' and 1040 908 cm-' are the

three short sharp peaks indicating C - 0 bands in al tphat tc chain

956 90 cm.' indicates the CH stretch, 762 222 c m ' , 743 888 cm-', and

701 67 cm ' tndicate the CH stretch in the ring structure, 651,455 cm-', and

578 122 cm-' indicate the C-H out plane b e n d ~ n g (St lverstein and

Webster, 1998)

CONCLUSIONS

Different solvent used in the column chromatography have

aided in t he distinct separat ion of t he act ive compound present in t he

vermicasts and these have been expressed in the FTIR spectra obtained

However from the spectra o f both 100%CPVC and that o f 1 . ICD.CHVC

(without passing through column chromatography), it is discernible that the

separation o f the compounds was limited

From the spectra I, 11 and 111 developed with the water extract o f

100% C P V C , it i s evident t ha t t h e phospha te g r o u p in CH-s t r e t ch

(in ring s t ructure) and N-H band in amide group occur in spectrum I

Phosphate containg C=O appeared in spectrum 11 CH stretch and amide

group, N-H bend~ng are conspicuous in spectrum I11

In the spectra 1, I1 and I11 which developed w ~ t h the extract o f

1 ICD CHVC, it is observed that the NH band In amide group, asymmetric

band o f CH stretch, C=O and P - 0 band occur in spectrum I C=O group

of the aromatic compound occurs in spectrum I1 The H band of OH group,

CH stretch of aromatic ring structure, phosphate containing ring structure,

C=O band in ring sturecture and CH out plane bending is noticeable in

The water extracts of 100% CPVC and I ICD CHVC demonstrated

(in our in ivtrp study, chapter VII) significant effect in the suppression of

plant pathogens Rhrroclonra solanr and Rhrzopus stolonrfer P-0 . C=O and

N-H (amide group) occuring in aromatic ring structure is the vermicasts of 100%

CPVC and I ICD CHVC are identified as contributing in the suppression

of these pathogens

DISCUSSION

E. eugeniae is very effective in the recycling of coir pith followed by the

anecic earthworm L.mauritii to be followed by P. excavatus. The contribution of

0.serrata in both vermicasts production and biomass production is significantly very

less. In all the four species in the production of vermicasts the common trend was

100%CD>3: lCD:CP> 1 : 1 CD: CP>1:3CD: CP>100%CP. In the recycling of coir

pith, it is preferable to use cow dung to aid in the process. Perionyx excavatus is

reported as the first species to invade the coffee pulp piles in the field. Since it can

survive field conditions and high temperatures characterized by the initial

decomposition of the coffee pulp, it has been used satisfactorily in open field conditions

(Aranda and Barois, 2000). Perionyx excavatus is often used for vermicomposting

because it is a tolerant species for the temperature and pH variations and has a

higher capacity to invade the organic matter at an early decomposition stage itself

(Aranda et al, 1999). In the present study, the rate potential of P excavatus in the

recycling of coffee husk with five different feed ratios is tested. The trend observed

with all the four earthworms in the recycling of coffee husk was 100%

CD>3: 1 CD:CH>l :I CD:CH> 1 :3CD:CH>1 OOCH, thus indicating that the feed

combinations have an impact on recycling in different species. Though Penier (1972)

reported Perionyx excavatus to be well suited for coffee husk recycling, from the

results of the present study E eugeniae followed by L.mauritii is found to have

recycled coffee husk with better (output) quantities of vermicasts as well as biomass

production. Vermicasts are effective in vegetable cultivation and act as good carrier

media for bio-fertilizers (Ismail, 1997).

The electrical conductivity and pH were near neutral in the vermicasts obtained

with 1:lCD:CP combination than those obtained from the other four feed ratios

i.e.l00%CD,3: lCb:CP, 1:3CD:CP and 100%CP. The carbon and nitrogen (C:N)

ratio projects the rate of digestion of organic waste by the earthworms (Kale et al.,

1994). It was observed that the C:N ratio obtained with the vermicasts of 1:l CD:

CP is more favorable for gemination as well as for plant growth in our experiments.

C:N ratio was less balanced in 100% CP vermicasts of Eudrilus sp.and Ochchaetona

sp., than the other two earthworm species. However C:N ratio in the vermicast of

coffee husk was relatively lesser in 0. serrata than in the other three earthworm

species. It was in L.mauritii (52.91:1)1:3CD:CH, E.eugeniae (38.42: 1) 3: ICD:CH,

0. serrata (30.86: 1) 3: 1 CD:CH, P.excavatus (49.29: 1) 100%CH vermicasts. The

carbon and nitrogen (C:N) ratio projects the rate of digestion of organic waste by

the earthworms (Kale et al., 1994). It was observed that the C:N ratio obtained

with the vermicasts of 1:l CD: CP is more favorable for germination as well as for

plant growth in our experiments.

Available nitrogen, phosphate and potassium content was higher in 1: ICD:CH,

100%CH and3:lCD:CH. Buck et.al., (1999) report that the plant nutrients are

generally more concentrated in vermicasts along with a higher microbial population.

That vermicasts were enriched with organic phosphate was reported by Scheu (1987).

But the earthworms prefer feed with low CM ratio (Hameed et al., 1993; Fledge1 et

a1.,1998). However the amount of organic matter in the vermicasts depends on the

earthworm's feeding ecology which is reported higher in detritivorous and lower in

geophagous (Corima Buck, 1989). Seasonally the total nitrogen content in vermicasts

is higher in summer than in other seasons but ammonical nitrogen showed no seasonal

differences (Matsumoto and Kasai, 1989). Earthworm activity has the potential to

contribute substantially to nutrient cycling in agro-ecosystems, more so when the

return of organic residues is increased thus reducing the intensity of cultivation.

Earthworm activity is of importance in arable lands by altering the soil structure,

aeration, providing channels for root growth, and drainage. These activities help to

reduce the tillage and the use of heavy machinery in conventionally filled soils

(Atlavinyte and Zirnkuviene, 1985; Lal, 1988; Bostrom, 1988).

However in coffee husk recycling available Phosphate (260.62 mg~lrg) and

potassium (155.5 mgkg) contents were relatively higher in 1 : 1CD:CH vermicasts

but less in nitrogen content. Available nitrogen (260.7 mgkg) content was higher in

100% CH vermicasts. However the amount of organic matter in the vermicasts

depends on the earthworm's feeding ecology which is reported to be higher in

detritivorous and lower in geophagous (Buck et.a1.,1989).

Though Wilson et.al., (1980) reported that the optimum pH levels for

composting is in the range of 5.5 to 8.0, in the present study pH in all the compost

cells by 90th day was above neutral except in control and CP-A,with good compost

maturity reached by 90th day. Similarly EC increased slightly from the beginning of

compost and it is observed to be so for every fifteen days interval. In CP-H reactor

with Trichoderma sp, and Actinomyceres-G compost maturity was highly significant.

This result indicates the release of easily decomposable compounds in the compost

solution (Saviozzi et aL.1987). EC increased sharply from 75th day to 90th day.

Similar observation was reported by Inbar et. al., (1993). Reduction in total organic

carbon was gradual by 90th day especially in CP-I reactor with all the 4 microbes in

combination.

Results of the present study also indicate gradual increase in conductivity

with progress in the composting of coir pith, pH in the CP-I compost reactor was

above neutral and in control reactor CP-F, it was acidic. pH attaining above neutral

condition processing with maturing of compost was reported by Jacobewitz and

Teenhuis (1984). However, the difference in pH in different reactors of the present

experiments noted is in correlation with the report ofAtchley and Clark (1979). who

nported that it is based on moisture, temperature and aeration condition in the compost

cells. However in pure coir pith vermicasts and composted coir pith it is the high

C:N ratio which reflects the digestion rate of organic waste by earthworms (Kale

et. al., 1994). This has been reflected in a better balancing of the C:N ratio in

177

CP-I compost. In comparison to control (CP-F) reactor C:N ratio in all other

reactors is well balanced. Decrease in C:N ratio indicates the degradation of organic

waste by the microbial action (Bonazzi et. al., 1990). Decrease in C:N ratio

indicates the positive influence of compost turning nitrogen-mobilization, nitrogen

toxification. Compost is applied to soil to prevent nitrogen immobilization and to

prevent and control soil nitrogen efficiency (Illmer and Schinner, 1997). The low

C/N ratio is the cause for ammonia toxicity and both extremes interfere with plant

growth (Inbar et.aL.1990).

In other compost reactors also N, P and K content was significantly higher

when compared to compost in control (CP-F). The N, P and K concentrations

indicate the immobilization of nutrient content (Illmer and Schinner, 1996). Providing

aeration to the compost cells helps to increase available nutrient content in the uunpost

(Bonazzi et a1.,1990). That N, P and K content in composted coir pith is higher than

the non-composted is reported by Jothimani(l993) and Joseph (1995). NPK help

in the absorption of plant nutrients, resulting in healthy growth and higher yield. It

also aids for faster germination of seeds (Salisbury et. a1.,1986). Compost when

added to soil increases nitrogen, phosphorus and potassium contents in the soil

(Zucconi and de Bertoldi, 1987; McCalla, 1975; Genevini et.al, 1983; Bauduin,

1987; Genevini et.al, 1987). Earthworm population in a compost added soil enhances

water availability to the plants (Jacobowitz and Steenhuis, 1984). Similarly when

compost is added to the soil, it not only increases water retention capacity but also

increases the temperature of the soil. All this add to improve soil quality because

when soil temperature is low, it delays seed germination and there is slow seedling

development (Van Wijk,et.a1,1959). Compost was reported to suppress the Dollar

spot (Sacatrotinia homoeocarpa) of Turfgrass (Boulter et.al, 2002) and suppress

other plant diseases through a combination of physico-chemical and biological

mechanisms (Boulter et.al., 2002).

The nutrient levels in the organic matter besides moisture and pH reduce

178

disease setting directly or indirectly affecting the pathogen or by increasing the host

capacity for resistance (Henis and Chet, 1975; Lockwood, 1986; Whipps, 1997).

Compost when used as manure provides nitrogen supplies which in turn increases

the yield. The pH gets maintained due to the buffering capacity of compost (Buchanan

et d.,1991). Compost is an environmentally and economically valuable material. The

enormous quantities of wastes are eliminated and become a product of value, and

improve the soils (Kuhlman,1990). Composted agricultural waste used as a container

media for plants, from l5thday to 90th day indicates the need for salt leaching in

composting (Inbar et.al., 1988). pH is higher than the optimum level of pH for

compost (5.5 - 8.0) reported by Wilson et. al., (1980). pH is also much higher in

coffee husk compost than that in coir pith compost on 15th day. The C:N ratio in

CH-I compost was 20.93:l on 15th and on 90th day C:N ratio in CH-I compost

was significantly reduced to 7.55:l. This reduction balancing the ratio of C to N has

been noted in all CH compost reactors when compared to control reactor. The

reduction in the ratio between C andN is indicative of compost maturity, which is

the result of induced decomposition and nitrogen immobilization.

In the vermicomposting of cornposted coir pith and coffee husk experiment

biomass production was higher in CP-E and CP-A of coir pith and in CH-A and

CH-H of coffee husk compost reactors. Significant increase in both EC and pH of

the vermicasts of composted coir pith (CP-I) indicate a higher availability of

nutrients (Lambert et, d.,1994). A higher total organic nitrogen balanced by a lesser

total organic carbon in CP-I, has made the C/N ratio relatively better balanced than

in all other composted cells making it more favorable for germination as well as

healthy plant growth. However in pure coir pith vermicasts and composted coir pith

it is the high C:N ratio which reflects the digestion rate of organic waste by earthworms

(Kale et. al., 1994). Nitrogen and potassium content in vermitreated composted

coir pith cell (CP-I) is higher than those of composted coir pith and vermicasts of

coir pith. NPK help in the absorption of plant nutrients, resulting in healthy growth

and higher yield. It also aids for faster germination of seeds (Salisbury et. al., 1986).

Accelerated germination (100%) was observed with the vermicasts,

especially of 1:l CD: CP and 1:3CD : CH as is reflected in the root length, shoot

length, fresh and dry weight of roots and shoots besides the number

of lateral roots. That vermicompost manure induces good rooting and yield responses

in turmeric was reported by Kale (1998). Kale et al., (1994) and Kale (1998) have

reported good yield responses when vermicast is used as manure in cash crops,

cereals and other plantations besides that in ornamental plants. Similar observations

were also presented in the yield responses of cardamon and vanilla cuttings. The

present study results are in correlation with the yield parameter like number of fruits,

number of seeds, fresh and dry weight of fruits and the weight of 100 seeds (Table

45). Though vermicompost is effective for yield responses (Kale,1998), in the present

experiment it is established that in the coir pith recycling 1: 1 ratio of CD:CP vermicast

has been more effective than the other combinations of feed in providing better yield

responses as is observed from the black gram pot study.

Vermicasts when used as container media or mixed with soil, improved its

physical, chemical and microbial properties and stimulated plant growth (Kale et al.,

1992, Riggle, 1998). Similarly the vermicasts rich in essential plant nutrients, when

used as organic manure improves physicochemical and biological properties of soil

(Purakayastha and Bhatnagar, 1997). Tomati et al. (1988) have demonstrated that

vermicasts promote plant growth, plant propagation; promote root initiation,

carpophore formation and nitrogen assimilation in Agaricus bisporus. Since

vermicompost does not contain toxic levels of any trace elements (Handreck, l986),

it serves as an ideal potting media.

After 12hours with 100% CPVC, the inhibition observed was 84.02% and

with 1 :I CD:CHVC it was 86.3%. Similarly the percentage of inhibition was 63.88%

with 100SCCPVC (Plate.15,B) and 61.66% with I : 1CD:CH VC after 24th hours.

The suppression of growth of Rhizopus srolonfler is greater than that of Rhizocronia

solani by 12 hours with both 1OO%CPVC and I: 1CD:CHVC. Szaechetd. , (1993)

180

reported on the inhibition of club rot caused by Plasmodiophora brassicae nor. by

vermicasts. Kostaka et al., (1996) reported that when potato plants were treated

with vermicompost, they became less susceptible to Phyfophthora infestans (mont)

de Bary than those plants treated with inorganic fertilizers (Arnaravadi et a1.,1990).

However Szezech and Smolinska (2001) reported that vermicompost of

all organic waste do not suppress plant pathogens. Suppression against plant

pathogens depends on the kind of organic materials used to produce the vermicasts.

It is the property of the vermicast that suppresses the pathogen. It also varies from

organism to organism i.e. different pathogens. Szezech, observation is based on the

experiments where vermicompost did not inhibit all the microorganisms present in

the potting media. Vermicompost also did not inhibit the total number of microorganism

in the potting media when compared with control (Szezech and Smolinska, 2001).

In the present investigation growth inhibition of Rhizocotonia solani and Rhizopus

srolont$envas quite significant. Amaravadi et al., (1990) reported that the earthworm

enzymes possessing virucidal activity are responsible for the control of Tobacco

mosaic virus (TMV) and cowpea mosaic virus (CPMV). Szezech et al. (1995) have

reported that when vermicast was added to the potting media and inoculated by the

pathogenic fungi Phyfophthora nicotianae var. nicotianae or Fusarium oxysporum

f. sp. Lycopersici, there is an increase in the yield and quality of tomatoes and hence

concluded that vermicasts as potting media have favourable pathogen controlling

effect.

In the present investigation growth inhibition of Rhizocofonia solani and

Rhizopus stolonifcr was quite significant. Amaravadi et al., (1990) reported that

the earthworm enzymes possessing virucidal activity are responsible for the control

of Tobacco mosaic virus (TMV) and cowpea mosaic virus (CPMV). Szezech et al.

(1995) have reported that when vermicast was added to the potting media and

inoculated by the pathogenic fungi Phyfophfhora nicotianae var. nicotfmae or

Furmlum oxysporum f. sp. Lycopersici, there is an increase in the yield and quality

181

of tomatoes and hence concluded that vermicasts as potting media have favourable

pathogen controlling effect. From the present invitro study, it is evidenced that among

the different extracts of vermicasts (based on feed ratio) 100%CPVC and

I :I CD:CHVC had significant suppressive effect against both the plant pathogens,

R. solani and R. slolonifer 1:l CD: CHVC than that by 100% CPVC suggesting

that the suppressive effect of vermicast varies from pathogen to pathogen and that it

also varies depending on the organic wastes from which the vermicasts are derived.

Those vermicastings which indicated better suppression of the plant pathogens - Rhizoctonio solani and Rhizopus srolonifer were tested for the characterization of

functional groups. C-H stretch of C=C bond of the aromatic structure was observed,

in CH-B, C,D, E, F, G and I compost cells and 100%CH vermicast. Aliphatic C-H,

OH, C - 0 stretches of the various compounds indicated the formation of humic acid

(Chen et.al.1995) during both composting and vermicomposting. Humic acid is one

of the soil conditioner and increase the population of the beneficial microorganisms,

humic substances carrying the micro nutrients in the soil and making them available

later to plant root hair (Hayes and Swift,1978; Stevenson,l982). Humic substances

participate are dissolution of minerals (Hoch et.al., 2000), and binding of small organic

molecules (Chiou et, al., 1986), reduction of metal ions(Szilagyi,l971). The humic

substances are important role in many agronomic, environmental and biochemical

processes (Hayes and Swift,1978; Stevenson,l982). The OH group and H band,

C=O groups CH-CH stretch; aromatic C=H stretch; P-0 group and N-H stretch

are the results of digestion of carbohydrates, amino acids, proteins and formation of

humic acid, in earthworm gut (Niemeyer,et,al.,l992). In pure coffee husk the

polymeric structure of OH group is an indication of digestive activity by the earthworm

(Chen.et.al, 1995). The two associated peaks in coffee husk vermicast spectrum

indicate the C-H stretch of the CH group as a result of digestion by the earthworm.

CH groups and amide groups observed in 3: ICD:CPVC, and 1:3CD:CPVC,

the P-H stretch in 100%CPVC, 3: ICD:CPVC, I: I CD:CPVC, and 1 :3CD:CPVC,

182

indicate the products of the earthworm gut activity. The second amide groups

developed in the spectrum of 1:ICD:CPVC and I:3CD:CPVC indicate that the

organic nitrogen was digested in the earthworm gut that contains the soil processing

and digesting enzymes.

Humic acid is one of the soil conditioner and increase the population of the

beneficial microorganisms, humic substances carrying the micro nutrients in the soil

and making them available later to plant root hair(Hayes and Swift,l978;Stevenson,

1982). Humic substances participate are dissolution of minerals (Hoch et.al., 2000),

and binding of small organic molecules

(Chiou et. al., 1986), reduction of metal ions (Szilagyi, 1971). The humic

substances play an important role in many agronomic, environmental and biochemical

processes (Hayes and Swift, 1978; Stevenson, 1982).