Fibers and Polymers 2009, Vol.10, No.2, 161-166
DOI 10.1007/s12221-009-0161-2
Study of Antimicrobial Activity of Aloevera, Chitosan, and Curcumin on Cotton, Wool, and Rabbit Hair L. Ammayappan* and J. Jeyakodi Moses1
Division of Textile Manufacture and Textile Chemistry, Central Sheep and Wool Research Institute, Avikanagar-304501, (Via) Jaipur, India 1 Department of Chemistry, PSG College of Technology, Coimbatore-641004, India
(Received May 14, 2008; Revised January 13, 2009; Accepted January 13, 2009)
Abstract: Aloevera, chitosan, and curcumin were applied in alone and in combination with each others on cotton, wool and
rabbit hair by exhaustion method for the assessment of their antimicrobial activity. The antimicrobial activity of these natural ingredients was better in peroxide treated cotton, formic acid treated wool/rabbit hair fibrous substrates than their corresponding intact ones. Aloevera shown better antimicrobial activity than chitosan and curcumin when applied alone and its antimicrobial activity was enhanced by addition of both chitosan and curcumin. The application of aloevera+chitosan+curcumin combination on peroxide treated cotton and formic acid treated wool/rabbit hair fibrous substrate was fast up to twenty five washing cycles. Keywords: Aloevera, Antimicrobial, Chitosan, Cotton, Curcumin, Wool, Rabbit hair
Introduction
Based on this literature review, it was decided to apply these natural antimicrobial ingredients in alone and in combination with each other on cotton, wool and rabbit hair fibrous substrates by exhaustion method for the assessment of their antimicrobial activity.
Microbial growth on textile materials has been considered as a major cause of biodegradation, which has led to development of antimicrobial technologies for preservative purposes. Such studies have further stimulated the research on antimicrobial textiles with focuses on development of durable and powerful antimicrobial finishing technologies [1-3]. Some of the typical treatments include sodium dichloroisocyanurate, pentachlorophenol, pentachlorophenyl laurate, quaternary ammonium compounds, chlorinated phenols, and organic tin compounds in finishing processes [4-8]. The active ingredients used in antimicrobial finish need to be registered after they have been demonstrated effective and they should be safe, non-toxic and bio-degradable. Present day’s textile processors preferred eco-friendly chemicals to impart antimicrobial finishing on textiles. Aloevera is a natural plant gives aloe gel, which has antimicrobial activity against various microbes and inhibited the growth of Mycobacterium, Trichophyton, and Bacillus subtilis [9,10]. Chitosan a natural biopolymer has a combination of many unique properties such as biodegradability, non-toxicity, cationic nature and antimicrobial activity. It is chemically called as beta-(1,4)-2-(amino)-2-deoxy-Dglycopyranose. In aqueous solution at pH < 6.5, its protonated form (NH3+) behaves like a cationic polyelectrolyte and interacting with negatively charged molecules i.e. cell wall of the microbes and inhibits their growth [11]. Curcumin is the dried rhizome of the plant Curcuma domestica and chemically called as 1-7-bis (4-hydroxy-3-methoxy phenyl)1,6-heptadiene-3,5-dione. The existence of methoxyl and hydroxyl groups is believed to be responsible for the antimicrobial activity [12].
Experimental Materials
The materials with the following specification were taken for this study, 100 % semi-bleached commercial cotton fabric (plain weave, 80’s count, 78 EPI and 60 PPI, 75 g/m2); Bharat merino wool fiber (23 micron fineness, 75 mm length) and Angora rabbit hair (16 micron fineness and 100 mm length); wool and rabbit hair were supplied by Central Sheep and Wool Research Institute, Rajasthan, India. The wool and rabbit hair were mild scoured with 0.5 % sodium carbonate and 0.1 % non-ionic wetting agent at 50 oC for 30 minutes, washed with distilled water and dried at ambient condition prior to application.
Antimicrobial Agents
Chitosan, with 81 % deacetylation and molecular weight of 65 kilo Dalton was procured from Central Marine Fisheries Research Institute, Cochin-682018 (India). Aloevera with molecular weight of 200 g/mol in gel form and curcumin with molecular weight of 368.4 g/mole in powder form were procured from Institute of Forest Genetics and Tree Breeding, Coimbatore, India. All other chemicals mentioned elsewhere were analytical grade reagents.
Methods
Initially the antimicrobial ingredients were applied to semi-bleached cotton and mild-scoured wool/rabbit hair (intact form) fibrous substrates. To improve their exhaustion,
*Corresponding author:
[email protected] 161
162 Fibers and Polymers 2009, Vol.10, No.2
L. Ammayappan and J. Jeyakodi Moses
hydrogen peroxide bleaching treatment on cotton and formic acid treatment (pretreatment) on wool/rabbit hair was given.
Peroxide Treatment for Cotton Substrate
The semi-bleached cotton substrate was subjected to hydrogen peroxide bleaching treatment as per standard recipe [13].
Formic Acid Treatment for Wool and Rabbit Hair Substrate
The formic acid treatment was given to both wool fiber and rabbit hair. The bath was set with 10 % (owf) formic acid (98 %) and the materials were entered at 25 C and treated for 30 minutes with material to liquor ratio as 1:10. Then they were washed with distilled water and dried. o
Antimicrobial Treatment
Aloevera and curcumin were dissolved with hot distilled water; chitosan was dissolved with 2 % acetic acid solution; and they were filtered before application. The required quantity of antimicrobial ingredients was taken in a bath with material to liquor ratio of 1:100. The sample was entered into the bath and the pH is maintained to 5.0±0.2 with acetic acid solution. The bath temperature was raised to 95 C and kept at this temperature for 30 minutes. After treatment the bath was cooled to 30 C; the sample was taken out and washed with warm water, rinsed with cold water and dried. The concentration of antimicrobial ingredients taken in alone and in combination form was given below. 1. Aloevera 5 g/ 2. Curcumin 5 g/ 3. Chitosan 3 g/ 4. Curcumin 5 g/ + Chitosan 3 g/ 5. Aloevera 5 g/ + Curcumin 5 g/ 6. Aloevera 5 g/ + Chitosan 3 g/ 7. Aloevera 5 g/ + Curcumin 5 g/ + Chitosan 3 g o
o
l
l
l
l
l
l l l
l
l
l
/l
Determination of Add-on (%)
The fibrous substrates were dried in a vacuum oven at 105 C until a constant weight was reached and oven-dried weight was determined before and after antimicrobial treatment and add-on value was calculated using the o
Table 1.
following formula. Add-on (%)=[( − )/ ]×100 Where : weight of fiber substrate before treatment (g) : weight of fiber substrate after treatment (g) W2
W1
W1
W2
Assessment of Antimicrobial Activity of Fibrous Substrates
The antimicrobial activity of intact, pretreated and subsequent antimicrobial ingredient treated fibrous substrates was determined as per standard method described by Quinn [14]. The amount of bacterial and fungal growth in inoculated and incubated samples was determined through serial dilution (10 , 10 and 10 ) and subsequent inoculation of sterile agar. The numbers of bacterial and fungal colonies were counted visually using a microscope. The wash fast test was carried out according to ISO63301984E [15], with a precision washer for 5, 10, 15, 20 and 25 washings followed by the assessment of their antimicrobial activity. -1
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Results and Discussion
Add-on of Antimicrobial Ingredient on Fiber Substrates
The amount of added antimicrobial ingredients on intact and pretreated cotton, wool and rabbit hair fibrous substrates is given in Table 1. It depicts that exhaustion of antimicrobial ingredients is higher in pretreated substrates than their corresponding intact ones. The pretreatment increases wettability, critical surface tension as well as accessible regions and decreases the diffusion barrier of the fibrous substrates. These factors lead to improvement in adhesion, spreading and exhaustion of antimicrobial ingredient into the fibrous substrates [16,17]. It also infers that, the higher add-on value in combined antimicrobial ingredient treatment is responsible for better antimicrobial activity than their corresponding individual treatment.
Antimicrobial Activity of Finished Samples without Pretreatment The numbers of bacterial and fungal colonies observed in intact and antimicrobial ingredient treated fibrous substrates
Add-on (%) of antimicrobial (s) on fibrous substrates Fibrous substrate Cotton
Antimicrobial treatment 1. Aloevera 2. Chitosan 3. Curcumin 4. Chitosan+curcumin 5. Aloevera+curcumin 6. Aloevera+chitosan 7. Aloevera+curcumin+chitosan
W1
Wool
Rabbit hair
Intact
Peroxide treated
Intact
Formic acid treated
Intact
Formic acid treated
0.38 0.75 0.65 -
2.25 2.00 1.80 3.80 4.05 4.25 6.05
0.29 0.53 0.57 -
1.69 1.45 1.31 2.85 3.04 3.19 4.57
0.23 0.41 0.45 -
1.35 1.23 1.11 2.27 2.34 2.45 3.65
Fibers and Polymers 2009, Vol.10, No.2
Antimicrobial Activity on Cotton, Wool, and Rabbit Hair
Number of bacterial colonies observed in antimicrobial treated fibrous substrates Fibrous substrate Intact cotton Intact wool Dilution 10 10 10 10 10 Antimicrobial treatment 1. Blank > 300 > 300 > 300 > 300 164 2. Aloevera > 300 64 11 > 300 50 3. Chitosan > 300 52 2 > 300 128 4. Curcumin > 300 33 2 > 300 60
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Table 2.
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Number of fungal colonies observed in antimicrobial treated fibrous substrates Fibrous substrate Intact cotton Intact wool Dilution 10 10 10 10 10 Antimicrobial treatment 1. Blank > 300 90 20 > 300 24 2. Aloevera > 300 76 3 60 18 3. Chitosan > 300 84 14 > 300 14 4. Curcumin > 300 54 7 > 300 12
Intact rabbit hair 10
10
10
10
21 4 14 4
> 300 > 300 > 300 > 300
208 44 248 20
152 12 130 1
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Table 3.
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Intact rabbit hair 10
10
10
10
3 2 4 1
> 300 47 > 300 > 300
74 21 15 64
12 11 2 4
-3
Number of bacterial colonies observed in antimicrobial(s) treated fibrous substrates Fibrous substrate Peroxide treated cotton Formic acid treated wool Dilution 10 10 10 10 10 10 Antimicrobial treatment 1. Blank 98 22 4 64 21 5 2. Aloevera 4 1 Nil 14 2 Nil 3. Chitosan 12 2 Nil 18 4 Nil 4. Curcumin 12 2 Nil 20 4 Nil 5. Chitosan+curcumin 2 1 Nil 3 1 Nil 6. Aloevera+curcumin Nil Nil Nil Nil Nil Nil 7. Aloevera+chitosan Nil Nil Nil Nil Nil Nil 8. Aloevera+curcumin+chitosan Nil Nil Nil Nil Nil Nil
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Table 4.
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are given in Table 2 and 3 respectively. It depicts that the residual impurities present in intact fibrous substrates influence the growth of microbial [18] and also reduces the exhaustion of antimicrobial ingredient. This trend is also observed in combined antimicrobial treatments; hence peroxide treatment for cotton and formic acid treatment for wool/rabbit hair was given to study their effect
Antibacterial Activity of Finished Samples with Pretreatment Table 4 shows the number of bacterial colonies observed in pretreated and antimicrobial ingredient treated fibrous substrates. It reveals that the number of bacterial colonies is reduced after application of antimicrobial ingredients either alone or in combination form. The antimicrobial activity of aloevera treated cotton substrate is better than chitosan and curcumin treated ones, while in wool/rabbit hair substrate it
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Formic acid treated rabbit hair 10
10
10
104 12 24 20 3 Nil Nil Nil
15 1 2 1 1 Nil Nil Nil
4 Nil Nil Nil Nil Nil Nil Nil
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is in the order of aloevera>curcumin>chitosan. It is noted that acemannon, anthroquinone, and salicylic acid component present in aloevera helps to promote antimicrobial activity and is further enhanced by amino acids, zinc and saponins present in it [19]. The antimicrobial activity of chitosan differs with respect to fibrous substrates. The sorption of chitosan on cellulose/protein fibrous substrates is due to ionic interaction between negative charges [hydroxyl anions in cellulose polymer (-O−) and carboxylate anions (-COO−) in protein polymer] and protonated amino groups of chitosan (NH3+), hydrogen bonding and van der Waals’ forces. However its’ affinity can be generally regarded as weak and so its antimicrobial activity is weak when applied alone [20-22]. Curcumin has only phenolic groups, which form hydrogen bonding with hydroxyl groups of cellulose polymer and amino/hydroxyl groups of protein polymer. Since the molecular weight of curcumin is smaller
164 Fibers and Polymers 2009, Vol.10, No.2
L. Ammayappan and J. Jeyakodi Moses
and so improve their activity on fibrous substrates. The charged state of new functional groups present in antimicrobial treated fibrous substrate attacks the cell wall of the bacteria and inhibits the microbial growth in finished fibrous substrates [27].
than chitosan, its exhaustion is higher than chitosan in all fibrous substrates and so its’ antibacterial activity is slightly better than chitosan [23]. When the antimicrobial ingredients are applied in combination form, the antibacterial activity is better in aloevera combinations than non-aloevera combination. It depicts that the antibacterial activity of aloevera is further enhanced by the addition of both chitosan and curcumin. The interaction between chitosan/curcumin and the functional groups such as hydroxyl, carboxyl and amino groups present in fibrous substrates may be weak and so the antibacterial activity of chitosan+curcumin combination is lesser than aloevera combination. In aloevera combination, the adsorbed chitosan, which is rich in protonated amino groups increases the attractive forces to nucleophiles present in aloevera and enhanced their exhaustion. It is also observed that antimicrobial activity of aloevera, chitosan and curcumin treated fibrous substrates with pretreatment is better than their intact ones. Formic acid treatment modifies disulphide linkages and wettability of wool/rabbit hair and peroxide treatment improves the accessible regions of cotton [24,25]. These modifications improve swelling of fibrous substrates in aqueous state and increase the adhesion, spreading, and exhaustion of antimicrobial ingredients [26]. The exhausted ingredients further form chemical bonding with functional groups present in fibrous substrate and also in other ingredients. These chemical bonding further increases the exhaustion/fixation of antimicrobials
Antifungal Activity of Finished Samples with Pretreatment
Table 5 shows the number of fungal colonies observed in pretreated and antimicrobial treated cotton, wool and rabbit hair fibrous substrates. The antifungal property of finished fibrous substrate is similar to their antibacterial property. Among three antimicrobial ingredients, aloevera shows better antifungal activity in wool/rabbit hair than cotton. The phenolic groups (in aloevera and curcumin) and anthroquinone groups (in aloevera) are binding microbial proteins, inhibiting their growth and so exhibit both antibacterial and antifungal activity [28,29]. The antifungal activity of aloevera further enhances by the addition of chitosan or curcumin, while chitosan and curcumin combination shows lesser activity than aloevera combination.
Antimicrobial Activity of Finished Samples with Respect to Washing Table 4 and 5 reveals that, the antimicrobial activity of aloevera+chitosan+curcumin combination is better than their individual and dual combinations. Hence this triple combination was selected and applied on pretreated fibrous
Number of fungal colonies observed antimicrobial(s) treated fibrous substrates Fibrous substrate Peroxide treated cotton Formic acid treated wool Dilution 10 10 10 10 10 10 Antimicrobial treatment 1. Blank 90 20 4 24 3 1 2. Aloevera 16 2 Nil 8 2 Nil 3. Chitosan 24 5 Nil 4 1 Nil 4. Curcumin 20 2 Nil 8 1 Nil 5. Chitosan+curcumin 2 1 Nil 1 Nil 6. Aloevera+curcumin Nil Nil Nil Nil Nil Nil 7. Aloevera+chitosan Nil Nil Nil Nil Nil Nil 8. Aloevera+curcumin+chitosan Nil Nil Nil Nil Nil Nil
Table 5.
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Formic acid treated rabbit hair 10
10
10
14 4 5 8 1 Nil Nil Nil
4 1 2 2 Nil Nil Nil
1 Nil Nil Nil Nil Nil Nil Nil
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Number of bacterial colonies observed in aloevera+curcumin+chitosan treated fibrous substrates with respect to washing cycles Fibrous substrate Peroxide treated cotton Formic acid treated wool Formic acid treated rabbit hair Dilution 10 10 10 10 10 10 10 10 10 0 washing Nil Nil Nil Nil Nil Nil Nil Nil Nil 5 washing Nil Nil Nil Nil Nil Nil Nil Nil Nil 10 washing Nil Nil Nil Nil Nil Nil Nil Nil Nil 15 washing Nil Nil Nil Nil Nil Nil Nil Nil Nil 20 washing Nil Nil Nil 1 Nil Nil 1 Nil Nil 25 washing Nil Nil Nil 2 Nil Nil 2 Nil Nil
Table 6.
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Antimicrobial Activity on Cotton, Wool, and Rabbit Hair
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Number of fungal colonies observed in aloevera+curcumin+chitosan treated fibrous substrates with respect to washing cycles Fibrous substrate Peroxide treated cotton Formic acid treated wool Formic acid treated rabbit hair Dilution 10 10 10 10 10 10 10 10 10 0 washing Nil Nil Nil Nil Nil Nil Nil Nil Nil 5 washing Nil Nil Nil Nil Nil Nil Nil Nil Nil 10 washing Nil Nil Nil Nil Nil Nil Nil Nil Nil 15 washing Nil Nil Nil Nil Nil Nil Nil Nil Nil 20 washing Nil Nil Nil Nil Nil Nil Nil Nil Nil 25 washing 2 Nil Nil Nil Nil Nil Nil Nil Nil
Table 7.
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substrates in order to assess their durability with respect to washing. Table 6 and 7 show the effect of washings on antimicrobial activity of finished fibrous substrates in terms of number of bacterial as well as fungal colonies observed. It depicts that, there are no bacteria and fungal growth in the finished fibrous substrates up to 20 washings and after 25 washings two bacterial colonies at 10 dilution is observed in wool/ rabbit hair and two fungal colonies at 10 dilution in cotton substrate. It reveals that this combined antimicrobial finishing is more suitable for pretreated cotton/wool/rabbit hair based textile products which can provide odour-free or hygienic functionality garments/fabrics. -1
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Conclusion
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The application of aloevera, chitosan and curcumin on peroxide treated cotton and formic acid treated wool/rabbit hair fibrous substrates show better antimicrobial activity than their corresponding intact ones. The pretreatment improves wettability, critical surface tension as well as accessible regions of fibrous substrate which enhances the exhaustion of antimicrobial ingredients. aloevera shown better antimicrobial activity than chitosan and curcumin when applied alone and its antimicrobial activity was enhanced by addition of both chitosan and curcumin. In aloevera+chitosan+curcumin combination treatment , the functional groups present in the exhausted antimicrobial ingredient in fibrous substrates further improve the fixation of other antimicrobial ingredient. This synergistic action enhances the overall antimicrobial activity of fibrous substrate. The aloevera+chitosan+curcumin combination finishing on pretreated cotton/wool/rabbit hair fibrous substrates is durable up to twenty five washing cycles. These antimicrobial ingredients are eco friendly and so give good effect to human skin in addition to values on garments.
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