<p><italic>Sclerotium rolfsii</italic> Sacc. is known to secrete cell degrading enzymes leading to collar rot of chickpea. Bateman (<xref ref-type="bibr" rid="bib001">1969</xref>) emphasized the importance of cellulase (Cx) in pathogenesis of <italic>S. rolfsii</italic>. The pectic enzymes were the primary factor involved in the rots caused by <italic>S. rolfsii</italic>. Chandrasekaran <italic>et al</italic>. (<xref ref-type="bibr" rid="bib002">2000</xref>) found that both total and ortho-dihydroxy phenolics compound were more in resistant cultivars of soybean. The activity of peroxidase was higher. It also produced polygalacuronase in carboxy methyl cellulose and pectin media. Hence, they suggested that both oxalic acid and pectic enzymes are involved in the destruction of host tissues by <italic>S. rolfsii</italic>. The present study deals with inducer source relationship of macerating activity, PG and Cx production under optimum time interval and pH conditions.</p><p>Cellulose activity was determined by measuring the reduction in viscosity of 0.5% Carboxy Methyl Cellulose (CMC) solution. Viscometric measurements were made with Ostwald's viscometer at time intervals of 0, 30, 60, 90, 120, 150, 180 and 210 min. The reaction mixture consisted of 5 ml of 0.5% carboxy methyl cellulose solution, 2 ml of sodium citrate buffer (at pH levels of 4, 7 and 9.2) and 2 ml of enzyme preparation. The enzyme preparation heated to 100°C for 10 min served as control. Polygalacturonase activity was determined by measuring loss in viscosity of 1.2 pectin solution. Viscometric measurements were made with Ostwald's viscometer at different time intervals i.e. 0, 30, 60, 90, 120, 150 and 180 min. The reaction mixture consisted of 5 ml of 1.2% pectin solution, 2 ml of sodium citrate buffer (at pH levels of 4, 7 and 9.2) and 2 ml of enzyme preparation. The reduction viscosity for CMC as well as PG was calculated by the following formula: % loss in Viscosity = (T<sub>0</sub> – T<sub>1</sub>)/(T<sub>0</sub> – T<sub>W</sub>) x 100. Where, T<sub>0</sub> = Flow time of reaction mixture at ‘0’ min, T<sub>1</sub> = Flow time of reaction mixture at a particular time interval, T<sub>W</sub> = Flow time of distilled water.</p><p>The results (<xref ref-type="table" rid="tab001">Table 1</xref>) clearly showed that the cellulolytic activity differed with change in pH levels 4 to 9.2 and time intervals 0 to 210 minutes. The per cent loss in viscosity was more (68.1%) at pH 9.2 as compared to 66.6% and 40.0% at pH 4.0 and 7.0, respectively. The per cent loss in viscosity was maximum at 180 min in all range of pH. <table-wrap id="tab001"><label>Table 1</label><caption><p>Influence of culture filtrate of <italic>Sclerotium rolfsii</italic> - activity of carboxy methyl cellulose and pectin</p></caption><table><thead><tr><th rowspan="7" align="center">Time interval (min)</th><th colspan="6" align="center">% loss in viscosity</th></tr><tr><th colspan="6"><hr /></th></tr><tr><th colspan="6" align="center">pH of enzyme mixture</th></tr><tr><th colspan="6" align="center"><hr /></th></tr><tr><th colspan="3" align="center">Carboxy methyl cellulose</th><th colspan="3" align="center">Pectin</th></tr><tr><th colspan="3"><hr /></th><th colspan="3"><hr /></th></tr><tr><th align="center">4.0</th><th align="center">7.0</th><th align="center">9.2</th><th align="center">4.0</th><th align="center">7.0</th><th align="center">9.2</th></tr></thead><tbody><tr><td colspan="7"><hr /></td></tr><tr><td align="center">0</td><td align="center">0.0</td><td align="center">0</td><td align="center">0.0</td><td align="center">0</td><td align="center">0.0</td><td align="center">0</td></tr><tr><td align="center">30</td><td align="center">66.6</td><td align="center">20</td><td align="center">13.6</td><td align="center">0</td><td align="center">0.0</td><td align="center">20</td></tr><tr><td align="center">60</td><td align="center">66.6</td><td align="center">20</td><td align="center">36.3</td><td align="center">0</td><td align="center">11.1</td><td align="center">20</td></tr><tr><td align="center">90</td><td align="center">66.6</td><td align="center">20</td><td align="center">45.4</td><td align="center">0</td><td align="center">11.1</td><td align="center">20</td></tr><tr><td align="center">120</td><td align="center">66.6</td><td align="center">40</td><td align="center">50.0</td><td align="center">20</td><td align="center">22.2</td><td align="center">30</td></tr><tr><td align="center">150</td><td align="center">66.6</td><td align="center">40</td><td align="center">54.5</td><td align="center">30</td><td align="center">22.2</td><td align="center">30</td></tr><tr><td align="center">180</td><td align="center">66.6</td><td align="center">40</td><td align="center">68.1</td><td align="center">30</td><td align="center">22.2</td><td align="center">30</td></tr><tr><td align="center">210</td><td align="center">66.6</td><td align="center">40</td><td align="center">68.1</td><td align="center">30</td><td align="center">22.2</td><td align="center">30</td></tr></tbody></table></table-wrap></p><p>The enzymatic activity of culture filtrate started at 30 min and became maximum (66.6%) at pH 4.0. However, maximum activity of culture filtrate in other range of pH was recorded at 180 min. The pH 9.2 was most suitable for degradation of CMC by culture filtrate of <italic>S. rolfsii</italic>.</p><p>Influence of culture filtrate of <italic>S. rolfsii</italic> on the activity of PG at different pH levels (<xref ref-type="table" rid="tab001">Table 1</xref>) indicated that the pectin activity differed with change in pH and time intervals. The % loss in viscosity was started at 30 min of pH 9.2 followed by 60 min of pH 7.0 and 120 min of pH 4.0. Thereafter, it became stagnant at all the pH range after 120 min. The % loss in viscosity was maximum at 120 min. However, Dutta and Jindal (<xref ref-type="bibr" rid="bib003">2002</xref>) reported that pectin had minor role in disease incidence. The % loss in viscosity was maximum at 9.2 pH. Thus pH of 9.2 was better in degradation of pectin by culture filtrate of <italic>S. rolfsii</italic>.</p><p>Singh (<xref ref-type="bibr" rid="bib005">2004</xref>) observed that nitrate reductase activity, chlorophyll content, total sugar, protein, phenol contents were decreased on the infected plant parts of <italic>Brassica</italic> spp. Jain and Yadav (<xref ref-type="bibr" rid="bib004">2003</xref>) reported significant positive correlation of total and reducing sugars contents in susceptible finger millet.</p></sec></body><back><ref-list><title>References

APPS

Annals of Plant Protection Sciences

0971-3573

Society of Plant Protection Sciences

apps-13-2-062

SHORT COMMUNICATION

Enzymatic activities of Sclerotium rolfsii on pectin and cellulose

Toorray

N.K.

Verma

K.P.

Sinha

A.K.

Tripathi

B.P.

Department of Plant Pathology, Indira Gandhi Agricultural University, Raipur – 492 006, India

92005

132495496

<p><italic>Sclerotium rolfsii</italic> Sacc. is known to secrete cell degrading enzymes leading to collar rot of chickpea. Bateman (<xref ref-type="bibr" rid="bib001">1969</xref>) emphasized the importance of cellulase (Cx) in pathogenesis of <italic>S. rolfsii</italic>. The pectic enzymes were the primary factor involved in the rots caused by <italic>S. rolfsii</italic>. Chandrasekaran <italic>et al</italic>. (<xref ref-type="bibr" rid="bib002">2000</xref>) found that both total and ortho-dihydroxy phenolics compound were more in resistant cultivars of soybean. The activity of peroxidase was higher. It also produced polygalacuronase in carboxy methyl cellulose and pectin media. Hence, they suggested that both oxalic acid and pectic enzymes are involved in the destruction of host tissues by <italic>S. rolfsii</italic>. The present study deals with inducer source relationship of macerating activity, PG and Cx production under optimum time interval and pH conditions.</p><p>Cellulose activity was determined by measuring the reduction in viscosity of 0.5% Carboxy Methyl Cellulose (CMC) solution. Viscometric measurements were made with Ostwald's viscometer at time intervals of 0, 30, 60, 90, 120, 150, 180 and 210 min. The reaction mixture consisted of 5 ml of 0.5% carboxy methyl cellulose solution, 2 ml of sodium citrate buffer (at pH levels of 4, 7 and 9.2) and 2 ml of enzyme preparation. The enzyme preparation heated to 100°C for 10 min served as control. Polygalacturonase activity was determined by measuring loss in viscosity of 1.2 pectin solution. Viscometric measurements were made with Ostwald's viscometer at different time intervals i.e. 0, 30, 60, 90, 120, 150 and 180 min. The reaction mixture consisted of 5 ml of 1.2% pectin solution, 2 ml of sodium citrate buffer (at pH levels of 4, 7 and 9.2) and 2 ml of enzyme preparation. The reduction viscosity for CMC as well as PG was calculated by the following formula: % loss in Viscosity = (T<sub>0</sub> – T<sub>1</sub>)/(T<sub>0</sub> – T<sub>W</sub>) x 100. Where, T<sub>0</sub> = Flow time of reaction mixture at ‘0’ min, T<sub>1</sub> = Flow time of reaction mixture at a particular time interval, T<sub>W</sub> = Flow time of distilled water.</p><p>The results (<xref ref-type="table" rid="tab001">Table 1</xref>) clearly showed that the cellulolytic activity differed with change in pH levels 4 to 9.2 and time intervals 0 to 210 minutes. The per cent loss in viscosity was more (68.1%) at pH 9.2 as compared to 66.6% and 40.0% at pH 4.0 and 7.0, respectively. The per cent loss in viscosity was maximum at 180 min in all range of pH. <table-wrap id="tab001"><label>Table 1</label><caption><p>Influence of culture filtrate of <italic>Sclerotium rolfsii</italic> - activity of carboxy methyl cellulose and pectin</p></caption><table><thead><tr><th rowspan="7" align="center">Time interval (min)</th><th colspan="6" align="center">% loss in viscosity</th></tr><tr><th colspan="6"><hr /></th></tr><tr><th colspan="6" align="center">pH of enzyme mixture</th></tr><tr><th colspan="6" align="center"><hr /></th></tr><tr><th colspan="3" align="center">Carboxy methyl cellulose</th><th colspan="3" align="center">Pectin</th></tr><tr><th colspan="3"><hr /></th><th colspan="3"><hr /></th></tr><tr><th align="center">4.0</th><th align="center">7.0</th><th align="center">9.2</th><th align="center">4.0</th><th align="center">7.0</th><th align="center">9.2</th></tr></thead><tbody><tr><td colspan="7"><hr /></td></tr><tr><td align="center">0</td><td align="center">0.0</td><td align="center">0</td><td align="center">0.0</td><td align="center">0</td><td align="center">0.0</td><td align="center">0</td></tr><tr><td align="center">30</td><td align="center">66.6</td><td align="center">20</td><td align="center">13.6</td><td align="center">0</td><td align="center">0.0</td><td align="center">20</td></tr><tr><td align="center">60</td><td align="center">66.6</td><td align="center">20</td><td align="center">36.3</td><td align="center">0</td><td align="center">11.1</td><td align="center">20</td></tr><tr><td align="center">90</td><td align="center">66.6</td><td align="center">20</td><td align="center">45.4</td><td align="center">0</td><td align="center">11.1</td><td align="center">20</td></tr><tr><td align="center">120</td><td align="center">66.6</td><td align="center">40</td><td align="center">50.0</td><td align="center">20</td><td align="center">22.2</td><td align="center">30</td></tr><tr><td align="center">150</td><td align="center">66.6</td><td align="center">40</td><td align="center">54.5</td><td align="center">30</td><td align="center">22.2</td><td align="center">30</td></tr><tr><td align="center">180</td><td align="center">66.6</td><td align="center">40</td><td align="center">68.1</td><td align="center">30</td><td align="center">22.2</td><td align="center">30</td></tr><tr><td align="center">210</td><td align="center">66.6</td><td align="center">40</td><td align="center">68.1</td><td align="center">30</td><td align="center">22.2</td><td align="center">30</td></tr></tbody></table></table-wrap></p><p>The enzymatic activity of culture filtrate started at 30 min and became maximum (66.6%) at pH 4.0. However, maximum activity of culture filtrate in other range of pH was recorded at 180 min. The pH 9.2 was most suitable for degradation of CMC by culture filtrate of <italic>S. rolfsii</italic>.</p><p>Influence of culture filtrate of <italic>S. rolfsii</italic> on the activity of PG at different pH levels (<xref ref-type="table" rid="tab001">Table 1</xref>) indicated that the pectin activity differed with change in pH and time intervals. The % loss in viscosity was started at 30 min of pH 9.2 followed by 60 min of pH 7.0 and 120 min of pH 4.0. Thereafter, it became stagnant at all the pH range after 120 min. The % loss in viscosity was maximum at 120 min. However, Dutta and Jindal (<xref ref-type="bibr" rid="bib003">2002</xref>) reported that pectin had minor role in disease incidence. The % loss in viscosity was maximum at 9.2 pH. Thus pH of 9.2 was better in degradation of pectin by culture filtrate of <italic>S. rolfsii</italic>.</p><p>Singh (<xref ref-type="bibr" rid="bib005">2004</xref>) observed that nitrate reductase activity, chlorophyll content, total sugar, protein, phenol contents were decreased on the infected plant parts of <italic>Brassica</italic> spp. Jain and Yadav (<xref ref-type="bibr" rid="bib004">2003</xref>) reported significant positive correlation of total and reducing sugars contents in susceptible finger millet.</p></sec></body><back><ref-list><title>ReferencesBateman

D.F.

(1969). Some characteristic of the cellulase system produced by Sclerotium rolfsii Sacc. Phytopathology 59: 37-42.Chandrasekaran

Narasimhan

Rajappan

(2000). Changes in the activity of phenolics and oxidising enzymes in soybean cultivars resistant and susceptible to Colletotrichum truncatum. Ann. Pl. Protec. Sci. 8: 176-178.Dutta

Jindal

J.K.

(2002). Hydrolytic enzymes and exopolysaccharide production of Xanthomonas axnopodis pv. vignaeradiatae inciting bacterial leaf spot of mungbean. Ann. Pl. Protec. Sci. 10: 295-298.Jain

A.K.

Yadav

H.S.

(2003). Biochemical constituents of finger millet genotypes associated with resistance to blast caused by Pyricularia grisea. Ann. Pl. Protec. Sci. 11: 70-74.Singh

H.V.

(2004). Biochemical transformation in Brassica spp due to Peronospora parasitica infection. Ann. Pl. Protec. Sci. 12: 301-304.Bateman

D.F.

(1969) Some characteristic of the cellulase system produced by Sclerotium rolfsii Sacc. Phytopathology 59: 37–42.Chandrasekaran

Narasimhan

Rajappan

(2000) Changes in the activity of phenolics and oxidising enzymes in soybean cultivars resistant and susceptible to Colletotrichum truncatum. Ann. Pl. Protec. Sci. 8: 176–178.Dutta

Jindal

J.K.

(2002) Hydrolytic enzymes and exopolysaccharide production of Xanthomonas axnopodis pv. vignaeradiatae inciting bacterial leaf spot of mungbean. Ann. Pl. Protec. Sci. 10: 295–298.Jain

A.K.

Yadav

H.S.

(2003) Biochemical constituents of finger millet genotypes associated with resistance to blast caused by Pyricularia grisea. Ann. Pl. Protec. Sci. 11: 70–74.Singh

H.V.

(2004) Biochemical transformation in Brassica spp. due to Peronospora parasitica infection. Ann. Pl. Protec. Sci. 12: 301–304.