~ ~ f 8 (2&3) (2006): 128-131

Post-harvest Quality Deter iorat ion of Cane Juice: Physio-biochemical Indicators

ISHWAR SINGH I*, S. S O L O M O N ~, A. K. SHRIVASTAVA i, R.K. S INGW and J. SINGW

1Division of Plant Physiology & Biochemistry, 2Division of Crop Improvement lndian Institute of Sugarcane Research, Rae Bareli Road, Lucknow 226 002,

P.O. Dilkusha, lndia

A B S T R A C T

Studies were undertaken to ascertain the relative contribution ofpolysaccharide formation and inversion losses responsible for quality deterioration in cane juice after extraction. These included mixing of effective biocide (sodium azide, 1% w/v) to destroy all microbes and heating of juice (at 100~ for 2 min) just after milling. The juice was heated to inactivate acid invertase(s) present in milled juice. The addition ofbiocide in freshly milled cane juice, sustained the quality with not much change in recoverable sugars and juice pH after 96 h of storage, while in the untreated juice there was a drop of over 2.0 units in recoverable sugars and a sharp decrease in juice pH. The dextran, reducing sugars and specific activity of acid invertase were significantly low in biocide treated juice as compared to the untreated control. Although the contents of reducing sugars and acid invertase activity were low in initially heated juice after 96h of storage, the formation ofdextran was quite high with more decrease in recoverable sugars, increase in juice viscosity and drop in juice pH as compared to the biocide treated juice. Microbial infections which lead to formation ofdextran and consequently increased viscosity, are largely responsible for post-harvest quality deterioration of cane juice.

Key words: Sugarcane, recoverable sugars, juice pH, dextran, reducing sugars, acid invertase, juice viscosity.

I N T R O D U C T I O N

Quality losses after the harvest of sugarcane in the field, during transportation, in factory storage pile, or during subsequent milling operations, have become a major concern in recent years, particularly in sub-tropical India. The time lag between harvesting to milling of cane ranges between 3 to 7 days, which entails huge losses in recoverable sugar due to deterioration of harvested cane (Solomon et al., 2001). The quality loss in cane is primarily due to chemical (acid) and enzymatic inversion, and those from microbial invasion through the cut ends or damaged sites of stalk (Eggleston, 2002).

A number of cane deterioration products including high invert sugars, polysaccharides (e.g. dextran) and microbial contamination (e.g., ethanol and lactic acid formation) have been reported to predict and control processing problems at the factory (Solomon et al., 2006, Eggleston et al., 200 l, Lionnet, 1996, Morel du Boil, 1995), but not all deterioration products effect factory processing. Polysaccharide producing soil born

*Author for Correspondence: Ishwar Singh e-mail: singhishwar@vsnl.net

bacteria such as Leuconostoc spp. from cane field enters inside the cane through cut ends or damaged sites and thrives at the expense of stored sucrose, further reduces quality of milled juice. The Leuconostoc bacteria have the ability to synthesize alpha-glucan polysaccharide (Dextran) from sucrose through an extracellular enzyme called dextansucrase as shown below.

The extent of dextran synthesis vgries with the agro-

Dextransucrasc Sucrose + H20 ID (Glucose). + Fructose

Dextran Leuconostoc spp

climatic conditions, cane variety, method of harvesting, cut-to-crush delay and sanitary conditions inside the processing unit. Formation of microbial polysaccharides mainly dextran in cane juice has a negative impact on factory processing (Revelo et aL, 1991). The viscosity of juice increases by the presence of dextran, which ultimately interfere in the process of crystallization during sugar manufacturing (Clarke, 1997, Purchase, 2001, Solomon etal., 2001). Therefore, dextran has often been reported as cane deterioration indicator. Besides, some oligosaccharides have been reported as cane deterioration products (Eggleston et al., 2001, Morel du Boil, 1995) and are responsible for crystal deformation (Morel du Boil, 1991). The present investigation was undertaken to study

Post-harvest quality deterioration of cane juice: physio-biochemical indicators 129

the relative contribution of various physio-biochemical factors responsible for deterioration of stored cane juice.

M A T E R I A L S A N D M E T H O D S

Twenty canes were randomly pulled out from the piles of harvested canes kept at the research farm of the Indian Institute of Sugarcane research, Lucknow, India (26o56 ' N, 80052 ' E, and 11 lm above sea level) for transportation to a sugar factory. The fleshly milled mixed juice with initial juice pH 5.12 was brought to laboratory and 750 ml juice was placed in three separate beakers. In the first beaker, 0.1% (w/v) biocide (sodium azide) was added; in the second beaker the juice was heated at 100~ for 2 min and then cooled immediately on ice; in the third beaker, the juice was left untreated. The pH of the samples was adjusted to the initial pH of 5.12 after treatments. Adding sodium azide increased the juice pH to 5.39, which was lowered to 5.12 by addition of dilute HC1. Heating of cane juice decreased juice pH to 5.02, which was raised to 5.12 by dilute NaOH. Sub-samples (50 ml) from each treatment were then transferred to small beakers and were covered with Para film and aluminium foil to prevent evaporation and external contamination and to simulate juice in closed pipes and tanks in sugar factory. All beakers were stored in a BOD incubator at 284-2 ~ (approximate ambient temperature in North Indian sugar factories). The beakers in triplicate from each treatment were removed after 0, 24, 48, 72 and 96h and analyzed for quality parameters and observations on change in color and odor of the stored juice.

Recoverable sugars: The ~ in juice was recorded with Erma hand refractrometer. The clarified juice was analyzed for sucrose content with High Sensitive Polarimeter (Model SEPA 300, Horiba, Japan). The recoverable sugars in juice were calculated by the following equation.

Recoverable sugars (%) = 1.022 S -O . 292 B; where S = Sucrose%juice and B = ~ meter.

Reducing sugars: The reducing sugars in juice were estimated as per the procedure described by Nelson (1944) and were expresses as per 100 ~

pH: juice pH was measured at room temperature (254-2 ~ by Cyber Scan 500 pH. (Eutech Instruments) calibrated at room temperature using three different pH buffers (pH 4, 7 and 10).

Dextrau: The dextran content in juice was estimated by Rapid Haze method (Clarke et al., 1987)

Acid lnvertase: The activity of acid invertase in juice was determined by the method described by Rosario and Santisopasri (2003)

Viscosity: The viscosity of cane juice was measured by the procedure given in the Hands-On- Activi t ies Manual

(Anon., 1996).

R E S U L T S AND D I C U S S I O N

To study the relative contribution of various physio- biochemical factors (polysaccharide formation or inversion losses or both) responsible for post-harvest quality deterioration in cane juice especially in sub-tropical India, laboratory studies were undertaken. These included mixing of effective biocide to destroy all microbes and heating of juice just after milling. The juice was heated to inactivate acid invertase(s) present in milled juice, pH of treated (biocide or heat treated) was adjusted to the initial value of untreated one, in order to eliminate the changes caused by acid sucrose inversion.

The freshly milled juice was dark brown in color and had a characteristic fresh odor of cane juice. The untreated juice began to change color within 24 h and became yellowish brown with characteristic 'wine' odor after 96 h of storage. The heated juice became light brown with stale odor after 96 h of storage. On the other hand the addition of biocide had a remarkable effect on juice quality. There was no apparent change in color and odor of biocide treated juice even after 96 h, indicating a major role of microbes in deterioration of quality of cane juice. Therefore, color and odor of cane juice could serve as visual indicator of deterioration of juice quality (Eggleston, 2002).

The addition of biocide sustained the juice quality with not much change in recoverable sugars even after 96 h of storage, while there was about 2.27 and 1.43 units drop recoverable sugars in untreated and heated juice, respectively after 96 h (Fig. 1). Solomon et al. (2006) demonstrated the retention of juice quality by application of antibacterial chemicals-based formulation during post-harvest staling of cane juice.

~11.5 o

I1~ 10.5

10 0 24 48 72 96

Storage duration (h) [ --4,-Untreated -IF-Heated --jr-Biocide I

Fig. l. Effect of biocide and heating of cane juice on recoverable sugars.

The pH changes with time in stored milled juice are shown in Fig. 2. The cane quality deterioration as indicated by decrease in pI-I, strated immediately after milling in the untreated cane juice. The pH also decreased in the heated juice, but only after

5.2"l

4.64 4.54

4.44 4.3 I

0 24 48 72 9E

Storage duratuion (h)

I ~p- Untreated -~1~ Heated .-t]~ Bioclde I

Fig. 2. Effect ofbiocide and heating of cane juice on juice pH

24 h. In biocide treated juice there was no significant change in juice pH even after 96h, further supporting that microbial growth is mostly responsible for cane juice deterioration.

The microbial infestation in cane juice resulted in formation of polysaccharide (dextran) and increase in juice, viscosity as shown in Fig.3 and Fig. 4, respectively. In the

45 40

35 ~ 3o o. 25

2o

10

0 24 48 72

Storage duration (h)

i -t-Untreated -E-Heated -k-BiocideJ

9 6

Fig. 3. Effect ofbiocide and heating of cane juice on dextran formation

2,4'

2.2-

E 1.8.

~1.6; i 1.4:'

1,2-

1 0 24 48 72

Storage duration (h)

I -4k-Untreated -I~Heated -&-Biocide "1

96

Fig. 4. Effect ofbiocide and heating of cane juice onjmce viscosity

161

1 4 1 -

.o I 0 24 48 72 ;6

Storage duration (h)

[ -$- Untreated --liP- Heated -k- Biocide I

Fig. 5. Effect ofhiocide and heating of cane juice on formation of reducing sugars.

9O

~ . 80 S~. 70 ~ 6 o

~=~ 4o 3o

o o 24 48 72 96

Storage duration (h)

I -(k-Untreated -~Hea ted -tk-Biocide ]

Fig. 6. Effect ofbiocide and heating of cane juice on acid invertase activity.

untreated juice the dextran formation started immediately after milling and increased at a steady rate in stored juice. The initial microbial high load in cane juice could be that transferred from harvested cane in milled juice. In contrast, in heated juice less dextran was formed during first 24 h, because intial heating might have reduced the number of viable Leuconostoe

bacteria to a level where lag phase growth occurred. The bacteria later multiplied and produced dextran. On the other hand, in biocide treated juice there was no formation ofdextran UP to 24h, and further increase in its content was very slow, as compared to other treatments. Juice viscosity also increased in untreated and heated juice, but there was no significant change in the viscosity ofbiocide treated juice even after 96h, further, providing evidence for larger contribution of microbial activities in cane deterioration. Revelo et al. (1995) applied the disinfectant IFOPOL TM to stored billeted cane and observed significant reduction in the formation ofdextran as well as total polysaccharides.

The data related to inversion losses based on formation of reducing sugars and acid invertase activity is presented in Fig. 5 and Fig. 6, respectively. The reducing sugars content

Post-barvcst quality deterioration of cane juice: physio-biochemical indicators 131

increased sharply in untreated juice with concomitant increase in the activity of acid invertase. In heated juice there was no significant change in the reducing sugars and acid invertase activity up to 72h. The reducing sugars content in biocide treated juice also remained low as compared to the untreated one.

Deterioration of cane juice in milling tandem can occur in several ways. The stored sucrose could be hydrolysed in reducing sugars (glucose and fructose) by acid invertase enzyme (acid inversion of sucrose). Some species of yeasts (e.g. Saccharomyces) found at sugar factories converts sucrose into ethanol(Chen and Chou, 1993), while few yeast species secrete acid invertase enzyme (Hanko and Rohrer, 2000). Sucrose losses might occur due to its utilization by the microbes. The milling area inside a sugar factory is often referred as 'high infection zone' . The Leuconostoc bacteria utilizes glucose molecule fi'om sucrose to form dextran.

However, it appears that microbial infections are largly responsible for deterioration o f juice quality during factory processing, especially presence o f Leuconostol bacteria in milling tandem leads to formation of dextran which is often associated with increased juice viscosity and subsequent loss in recoverable sugars, Therefore, sucrose losses in milling tandem could be minimized by the application o f an effective biocide.

R E F E R E N C E S

Anonymous (1996). Laboratory manual for 'Hands-On-Activities '. On-line [http://www/spacegrant.hawaii.edu/class_acts/ Viscosity.html].

Chen, J.C.P. and Chou, C.C. (1993). Cane Sugar Handbook, 12th ed. John Willey and Sons Inc., New York.

Clarke, M.A. (1997). Dextran in sugar factories: Causes and control (Part I &II) Sugar YAzucar, October/November 1997.

Clarke, M.A., Bergeron, J. and Cole, F. (1987).A rapid dextran screening test. Sugar YAzucar, 82(3): 23-24.

Eggleston, G., Legendre, B.L., and Richard, C. (2001). Effect of

harvest method and storage time on cane deterioration. I: Cane quality changes. InternationalSugarJournal, 103, 331-338.

Eggleston, G. (2002). Deterioration of cane juice-sources and indicators. Food Chemistry, 78, 99-107.

t tanko, V.P. and Rohrer, J.S. (2000). Determination of carbohydrates, sugar alcohol and glycols in cell cultures and fermentation broths using high performance ion exchange chromatographs with pulsed amperometric detection. Analytical Bioehem., 283:192-199.

Purchase, B.S. (2001).Losses caused by micro-organisms. Proe. ISSCT, 24(1): 379.

Lionnet, G.R.E. (1996). A review of cane quality in South Africa and its impact on factory performance. Proc. ISSCT, 22: 103-113.

Morel du Boil, P.G. (1991). The role ofoligosaccharides in crystal elongation. In: Proceedings of South African Sugar Technologists Meeting. Pp. 171-178.

Morel du Boil, P.G. (1995). Cane deterioration- oligosaccharides formation and some processing implications. In : Proceedings of South African Sugar Technologists Meeting, Pp. 146-154.

Nelson, N. (1944). A photometric adoptation of the somogy's method of determination of glucose. J. Biol. Chem. 243:6281 - 6283.

Revelo, S.B.; Ramos, E.L. and Mejiras, R. (1991). Sugar cane deterioration and its implications in the factory, lnt. SugarJ., 93: 82-86.

Revelo, S.B.; Ramos, E.L. and Torres, B.M.P. (1995). Origin of oligo and polysaccharides in cane juice and their effect on sugar mill efficiency. Proc. ISSCT, 21(3): 894-901.

Rosario, E.J. and Santisopasri, S. (2003). Characterization and inhibition of invertase in sugarcane juice. Phytochem., 16: 443 -445.

Solomon, S., Shahi, H.N., Suman, A., Gaul', A., Deb, S. and Singh, I. (2001). A survey of post-harvest biological losses in Indian sugar factories: An emerging challenge. Proc. ISSCT, 24(1): 380-381.

Solomon, S., Banerji, R., Shrivastava, A.K., Singh, P., Singh, I., Verma, M., Praj apati, C.P. and Sawnani, A. (2006). Post-harvest deterioration of sugarcane and chemical methods to minimize sucrose losses. Sugar Tech, 8(1): 74-78.

Received February 18, 2006; Accepted August 23, 2006