a comparative study on the influence of temperature …

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50

A COMPARATIVE STUDY ON THE INFLUENCE OF TEMPERATURE

AND ACID CONCENTRATION ON WASTE AGRICULTURAL

CELLULOSE BIOMASS DEGRADATION

Nnam Raphael Eze1*, Oko Nnam Ahamefula

2, Ugah Chukwuemeka S.

3, Okoro Otuu

Inya2

and Chihurumnanya Ola Oji2

1Department of Food Technology Akanu Ibiam Federal Polytechnic, Unwana Ebonyi State

Nigeria.

2Department of Science Laboratory Technology Akanu Ibiam Federal Polytechnic, Unwana

Ebonyi State Nigeria.

3Department of Sciences, National Institute of Construction Technology, Uromi

Edo State Nigeria.

ABSTRACT

Bioconversion of agricultural waste products to produce value-added

food products like simple sugar; fuels and chemicals offers potential

economic, environmental and strategic advantages over traditional

fossil-based products. A comparative study on the effect of

temperature and concentration on waste cellulose biomass degradation

to glucose was studied at temperature range of 80 – 1300C and

concentration range of 2, 4, 4.5, 5.0, 5.5 and 6Mol/dm3 for a period of

48hours. The percentage (%) glucose yield was calculated at 8h

interval for the various temperatures and concentration ranges. This

study was done using two different acids namely sulphuric acid and

hydrochloric acid. The research showed that glucose is present in a

reasonable amount in sawdust and its %yield is greatly influenced by the acid concentration

and temperature. It is also influenced by the type of acid used and the reaction time. For a

temperature of 1300C the %glucose yield when sulphuric acid was used for the cellulose

degradation at 2.0, 4.0, 4.5, 50, 5.5 and 6.0mol/dm3 after 48h is 28.4%, 37.2%, 35.6%,

47.8%, 56.7% and 61.2% while the %glucose yield for hydrochloric acid at the same

temperature and concentrations stood at 25.1%, 33.2%, 30.0%, 45.4%, 51.2% and 58.1%.

The results showed that the rate of hydrolysis by virtue of glucose yield, generally increased

WORLD JOURNAL OF PHARMACY AND PHARMACEUTICAL SCIENCES

SJIF Impact Factor 7.632

Volume 10, Issue 1, 50-66 Research Article ISSN 2278 – 4357

*Corresponding Author

Nnam Raphael Eze

Department of Food

Technology Akanu Ibiam

Federal Polytechnic,

Unwana Ebonyi State

Nigeria.

Article Received on

29 October 2020,

Revised on 19 Nov. 2020,

Accepted on 09 Dec. 2020

DOI: 10.20959/wjpps20211-18021


51

Nnam et al. World Journal of Pharmacy and Pharmaceutical Sciences

with increase in temperature and acid concentration except for acid concentration from 4,0 -

4.5mol/dm3

were we noticed a decline in %glucose yield for both acids and all the

temperatures.

KEYWORDS: Acid Hydrolysis, Temperature, Acid Concentration, Sawdust, Agricultural

Waste.

1. INTRODUCTION

The agricultural activities of man have resulted in the production of large quantities of

agricultural waste biomass that tends to dominate and pollute the environment. Many of these

agro-wastes are allowed to rot away unutilized or underutilized (Latinwo, and Agarry 2015;

Obot et al., 2008). According to Adeoye et al (2019), the environmental impact of

indiscriminate agricultural wastes disposal in Nigeria has brought untold hardship on the

citizenry. Notably, consequence of this indiscriminate waste disposal can be classified into

five, vis-a-vis, global warming, photochemical oxidant creation, abiotic resource depletion,

acidification and eutrophication (Lamond et al. 2015). Bioconversion of agricultural wastes

biomass to produce value-added fuels and chemicals offers potential economic,

environmental and strategic advantages over traditional fossil-based products (Anex et al.,

2007).

Among all the constituents of agricultural wastes biomass, cellulose constitutes the highest

percentage because it is a strong elastic material that forms the cell wall of nearly all plants

(Latinwo, and Agarry 2015). Nnam et al. (2020) noted that cellulose is one of the major

constituent of all plant materials, which formed about half to one-third of all plant tissues.

These wastes biomass consist of cellulose, hemicelluloses, lignin and other materials called

extractive (Ghose1956). Generally, agricultural wastes from different sources have different

physical properties such as surface area, lignifications, crystallinity and other different

chemical compositions that could hinder the accessibility and susceptibility of cellulose for

hydrolysis (Aberuagba, 1997; Caritas and Humphrey, 2006). However, they may be modified

to enhance their susceptibility to hydrolysis through pretreatment processes. Chemical

modification of cellulose is performed to improve process ability and to produce cellulose

derivatives (cellulosics‟) which can be tailored for specific industrial applications (Akira

2001). Cellulosic waste materials are in general strong, reproducible, recyclable and

biocompatible (Conner A. H 1995).


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Wide varieties of biomass sources are available for further conversion and utilization. Proper

selection of the biomass feedstock is of paramount importance from both a techno- and socio-

economical point of view. The biomass feedstock should not compete with the food chain

and waste streams with a low or even negative value, such as agricultural waste, are

preferred. Furthermore, it is also advantageous to select sources that are not prone to diseases

and are preferably available throughout the year (Girisuta et al., 2008). Based on these

criteria, the wood sawdust is an excellent biomass feedstock for further conversion and

utilization.

Wood sawdust is a lignocellulosic byproduct of sawmill that is available at low cost

throughout the year (Islam and Mimi Sakinah, 2011). It is produced in enormous quantities

by sawmills and the economical disposal of them is a serious problem to the wood based

industries. Sawdust is commonly used as fuel in producing plants and local utilities. Other

uses of sawdust are: as litter and bedding material in poultry and livestock structures, for the

production of fiberboards and paper pulp (Harkins, 1969; Arends et al., 1985; Islam and

Mimi Sakinah, 2011; Nnam et al 2020; Kim et al 2017).

Over the last decades, the hydrolysis of cellulose and lignocellulosic materials (Klemm et al

1998; Mian et al 2017) has been a subject of intensive research for the development of large

scale conversion processes that would be of benefit to mankind (Patel and Bhatt, 1992;

Nicolettal et al., 2002; Hahn-Hagerdal et al., 2006; Qu et al., 2006). These processes would

among other things help to solve modern disposal problems, reduce pollution of the

environment and reduce man‟s dependence on fossil fuels by providing a convenient and

renewable source of energy in the form of bioethanol (Cowling et al., 1976).

Conversion of cellulose and lignocellulosic biomass to glucose and other monomeric sugars

can be achieved by acid and enzyme hydrolysis (Badger, 2002; Benkun, et al, 2008,

Megawati et al., 2010; Wu et al., 2010). The relative advantages of enzyme and acid

hydrolysis of cellulose is a subject of continuing research study. Olaru et al., (1997); Hienze

and Pfeiffer, (1999); Togrul and Arslan, (2004); Adinugraha et al., (2005); Pushpamalar et

al., (2006); Suzana, (2009) noted that cellulose potentially can be modified as carboxymethyl

cellulose, hydroxy propyl cellulose, methyl cellulose and hydroxyl propyl methyl cellulose.

Hutomo et al., (2015) argued that the sulfuric acid and hydrochloric acid are strong acid but

difference in effectively for cellulose degradation. The hydrochloric acid consist of one

proton per mole so the sulfuric acid content two protons per mole. Cellulose degradation by


53


acid is cheaper than by enzyme (Lu and Hsieh, 2010; Habibi et al., 2010; Devabaktuni

Lavanya et al 2011).

Enzymatic hydrolysis of cellulose to glucose is carried out by cellulase enzymes which are

highly specific catalysts (Molina et al 2014; Saha et al., 2005). The enzymatic process is

believed to be the most promising technology because enzymatic hydrolysis is milder and

more specific and does not produce by products (Wen et al., 2004; Benkun et al, 2008).

However, enzymatic hydrolysis of cellulose have been observed not to be economically

viable because of high cost of enzymes, slow rate of depolymerization and high enzyme

loading to realize reasonable rates and yields (Layokun, 1981; Aberuagba, 1997; Grohmann

and Baldwin, 1995; Wyman, 1999). Vaccarino et al; (1989); Grohmann et al., (1995);

Talebnia et al., (2008) reported that the advantages of acid hydrolysis for peel liquefaction

and releasing carbohydrates prior to enzymatic treatment have been studied. Acid hydrolysis

of cellulosic biomass is relatively fast and of low cost (Palmqvist and Hagerdal, 2000;

Megawati et al., 2010). The key variables that might have impacts on the rate and extent of

cellulose and lignocellulosic biomass by acid hydrolysis are temperature, acid concentration

(or pH), total solid fraction (TS) and time duration (Grohmann, et al., 1995; Talebnia et al.,

2008; Brudecki et al., 2012; Jeya et al., 2012; Liu et al., 2010).

Several studies have been made on cellulose hydrolysis using purified raw materials

especially commercial cellulose as well as cellulose derived from agricultural wastes such as

Eucalyptus wood, Meranti wood sawdust, corncobs, sugarcane bagasse, sorghum straw,

brewer‟s spent grain, and OPEFB using dilute sulphuric acid and very high temperature (100

– 2500C (Parajo et al., 1994; Dominquez et al., 1997; Lavarack et al., 2002; Mussato and

Roberto, 2005; Rahman et al., 2007; Xiang et al 2003). However, very few studies using

concentrated sulphuric acid and moderate temperature have been carried out (Aberuagba,

1997; Ajani et al., 2012; Adeoye et al 2019). Therefore the objective of this study is to

examine the effect of concentration and low to moderate temperatures on the saccharification

of cellulosic waste material (wood sawdust) when sulphuri and hydrochloric acids are used.

2. MATERIALS AND METHOD

Wood sawdust was obtained from Afikpo Timber Shade, Afikpo North Local Government

Area, Ebonyi State, Nigeria. Sulfuric acid and hydrochloric acid used for hydrolysis and

Diethyl ether used for the removal of lignin and extractives which are products of E. Merck


54


(Darmstadt Germany) were purchased from a chemical store in Abakaliki Ebonyi State,

Nigeria. Other chemicals used were of analytical or biochemical grade.

2.1. Pretreatment of wood Sawdust and Cellulose isolation

Samples of wood were oven dried at 70 until a constant weight was obtained and further

dried in a furnace for 10 min. The dried wood samples were reduced to very small sized

particles of about 300μm by grinding using a grinding machine (Flammer Mill, Model 200,

UK) incorporated with 2mm sieve.

2.2. Dissolution of wood

Cellulose was isolated from the small particle sized Scaphium spp. sawdust samples using

the modified procedure described by Layokun (1981). Dimethyl Sulfoxide (DMSO) in liquid

form was prepared for stirred with the sawdust. In dissolution process, 5 different mass of

sawdust has been prepared which were in 10g, 20g, 30g, 40g and 50g. Then, all the samples

were dispersed into Diethyl ether (100 ml) added to the sample (300g) in a 250ml Erlenmeyer

conical flask so as to remove the extractives. The resultant residue (free of extractives) was

filtered and washed thoroughly with sterile distilled water. To the washed residue was added

500ml of 14M sulfuric acid which then dissolved the cellulose and hemi cellulose leaving

lignin as a hard precipitate. Lignin was filtered off and 8M sodium hydroxide solution was

added to the filtrate to obtain a residue that was predominantly cellulose, while hemi

cellulose remained in solution. The solution was filtered and the resultant cellulose residue

was then washed thoroughly with sterile distilled water until a neutral pH was obtained. The

cellulose residue was oven dried at 800C until a constant weight was obtained for subsequent

hydrolysis.

2.2.Experimental design for acid hydrolysis of cellulose

To determine the effect of different acid concentrations on the hydrolysis, 10g sample of the

cellulose in a 250ml conical flask which served as a batch reactor was added 100ml of

concentrated sulfuric acid (2mol/dm3). The flask was placed in a gyratory shaker set at a

temperature of 800C with an agitation speed of 150 rpm and was allowed to operate for 48h;

and at intervals of 8h, samples were withdrawn to determine the glucose concentration. The

experiment was repeated at other temperatures of 90, 100, 110, 120 and 1300C and sulfuric

acid concentrations of 4.0, 4.5, 5.5 and 6.0 moles/ dm3, respectively. The same method,

temperature and concentrations were used in the hydrolysis involving hydrochloric acid.


55


2.3. Determination of glucose concentration

The reducing sugar content (glucose) was determined by the DNS method with glucose as

standard (Miller, 1959; Marsden et al., 1982). Absorbance was measured at 540nm. However,

the DNS reagent was modified according to Mwesigye (1988). Two hundred grams of

potassium sodium tartarate (Rochelle salt) were dissolved in 200ml of sterile distilled water.

Ten grams of sodium hydroxide was measured at 540nm. Ten grams of sodium hydroxide

was also dissolved separately in 200ml of sterile distilled water in a 500 ml beaker. To the

sodium hydroxide solution was added 10 g of DNS (3, 5dinitrosalicyclic acid) and 2.52ml

(2g) of 80% (w/v) phenol simultaneously. After stirring to complete dissolution, the mixture

was added to the Rochelle salt solution. The resultant solution was then made up to one litre

with sterile distilled water. This mixture gave the stock of the modified DNS reagent

containing 1% (w/v) DNS acid, 0.2% (w/v) phenol, 1% (w/v) sodium hydroxide and 20%

(w/v) Rochelle salt (Mwesigye, 1988). The DNS reagent was then stored under refrigeration

in an amber coloured bottle.

2.4.Yield Calculation and Kinetics of acid hydrolysis

For molar cellulose concentration (Cc), cellulose is treated as glucan (anhydrousglucose), i.e.

(1)

In this way, cellulose is converted to glucose equivalents, which represent the maximum

theoretical glucose from cellulose. The cellulose concentration as glucose equivalents is

calculated as in Eq. (1) (Latinwo and Agarry (2015):

(2)

Where is the mass of cellulose or the mass of weighed cake from experiment, V is the

liquid volume, and MM is the molar mass. The glucose yield ( ) is defined in this

study per initial cellulose concentration as glucose equivalents.

3. RESULTS AND DISCUSSION

3.1. EFFECT TEMPERATURE

The percentage cellulose yield from the agricultural waste of saw-dust are 28.4%, 37.2%,

35.6%, 47.8%, 56.7% and 61.2% for Sulphuric acid; 25.1%, 33.2%, 30.0%, 45.4%, 51.2%

and 58.1% for hydrochloric acid at 1300C for 2M, 4M, 4.5M, 5.5M and 6.0M respectively

with Sulphuric yielding more glucose than hydrochloric acid for all temperature and


56


concentration under review. Benkun et al. (2009) obtained 60.12% cellulose from the acid

hydrolysis of wheat straw. The effect of temperature on glucose yield from acid hydrolysis of

wastes cellulose from saw-dust using sulphuric and hydrochloric acid is shown in Fig.1and 2

respectively. From Fig 1 and 2, it is seen that the glucose yield from the agricultural waste

cellulose (saw-dust) increased with increase in temperature for both Sulphuric and

Hydrochloric acids. A similar observation has been reported about agricultural waste

(Aberuagba, 1997; Adoeye et al. 2019; Hutomo et al 2015; Ajani et al. 2011; Layokun 1981;

Ajani et al., 2012; Kupiainen et al., 2014). However, Layokun (1981) was more specific

about saw-dust. He observed an increase in glucose yield as temperature increases for acid

hydrolysis of saw dust. Kim et al. (2012) reported that the pretreatment with 1% NaOH and

2% MCA for a total of 4 hours at 75and a hydrolysis period of 48 hours was determined to be

the optimum conditions for the carboxymethylation technique applied to sawdust. Talebnia et

al. (2008) reported that in the acid hydrolysis of orange peels at low temperature range, sugar

yield increased with increase in temperature and at very high temperature range, sugar yield

declines. However, Megawati et al. (2010) reported that in the acid hydrolysis of rice husk, at

high temperature range (160 – 2200C total sugar concentrations increased with increase in

temperature. Akpan et al., (2005) reported that the yields of glucose from wastes cellulose of

banana skin (0.20 - 0.37%) and maize stalks (0.20 - 0.41%) are relatively comparable at the

temperature of 70 – 1000C However, they are both lower than the glucose yield from wastes

cellulose of cowpea shells and rice husk, respectively. Also, the glucose yield obtained from

wastes cellulose of cowpea shells (0.64%) and rice husk (0.57%) are relatively comparable at

lower temperature range of 700C however, the yields from wastes cellulose of cowpea shells

(0.83 - 1.17%) are higher than the yields from rice husk (0.61 - 0.73%) at higher temperature

range of 80-1000C Aberuagba (1997) reported a glucose yield of 0.74 - 1.27% from wastes

cellulose of maize cobs and 0.67 - 1.17% from groundnut shells in an acid hydrolysis at a

temperature range of 65 – 80oC and 2.5M sulfuric acid concentration. Megawati et al (2010)

reported a sugar yield of 12.70% at a temperature of 2200C.

3.2. Effect of concentration

The results revealed that there was a general increase in glucose yield as the acid

concentration increased in the range 2 - 6mole/dm3. However, there was a decline in glucose

yield from 4.0 to 4.5moles/dm3. A similar observation has been reported for the acid

hydrolysis of cellulose from maize cobs and groundnut shells (Aberuagba, 1997; Hutomo et

al 2015). This observation could be attributed to the fact that at high acid concentration and


57


relatively high temperature; glucose can be converted to organic acid which led to a decrease

in glucose concentration (Aberuagba, 1997 Hutomo et al 2015). This suggests that maximum

glucose yield could be obtained at low to moderate acid concentration. Talebnia et al. (2008)

reported that at low acid concentration and low temperature, sugar yield increases with

increase in dilute acid concentration. The glucose yield obtained from wastes cellulose of

sawdust for sulphuric (0.30%) and hydrochloric acids (0.24%) at acid concentration of

2moles/dm3 each are relatively comparable. Rahman et al. (2006) also reported that the acid

hydrolysis of oil palm empty fruit bunch with acid concentration of 2 -6% produced a sugar

yield of 31.74%.

4. CONCLUSION

The acid hydrolysis of wood sawdust with sulphuric acid and hydrochloric acid was

successfully carried out in this research. The results obtained from the experiment reveal that

glucose is present in a reasonable amount in sawdust and its %yield is greatly influenced by

the acid concentration, nature of acid and temperature. Acid hydrolysis of wood sawdust

cellulose was successfully accomplished at different acid concentrations (2.0 – 6.0mol/dm3)

for both sulphuric and hydrochloric acid; temperature (80 – 1300C and reaction time (0 –

48h). The acid hydrolysis of sawdust cellulose increases with increase in acid concentration,

reaction time and temperature. The nature of the acid used also affected the yield as sulphuric

acid yielded the highest glucose per cent at all temperature and concentration more than the

hydrochloric acid.

1(a)


58


1(b)

1(c)

1(d)


59


1(e)

1(f)

Fig. 1(a), 1(b), 1(c), 1(d), 1(e) and 1(f) represents the % Glucose Yield for temperature

range of 80 – 1300C at 2.0, 4.0, 4.5, 5, 5.5 and 6mol/dm

3H2SO4 respectively.

Fig. 1g: A comparative analysis of % Glucose yield at 130 for all the different

concentrations.


60


2(a)

2(b)

2(c)


61


2(d)

2(e)

2(f)

Fig. 2(a), 2(b), 2(c), 2(d), 2(e) and 2(f) represents the % Glucose Yield for temperature

range of 80 - 130 at 2.0, 4.0, 4.5, 5.0, 5.5 and 6mol/dm3H2SO4 respectively.


62


2(g): A comparative analysis of %Glucose yield at 1300Cfor all the different

concentrations.

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