Development of microbial resistant -...

Development of

microbial resistant

Ag0-transdermal surgical cellulose scaffolds

via

environmental

friendly green

process Development of

PABs

3.1 Introduction

The fact that non-microbicidal wound scaffolds allow microorganisms to the site of

wound to exhibit undesirable effects on wounds and it is one of the main factors for disease

transmission. This factor has stimulated intensive research, for the development of antimicrobial

wound dressings and bandages, in the medical field. Therefore, it has become crucial to impart

antimicrobial activity to the wound dressing materials, in order to protect the user from

microorganisms against contamination [1].

In order to impart antimicrobial activity to the medical textile materials, a number of

chemicals have been employed. These chemicals include: inorganic salts, organometallics,

iodophors (substances that slowly release iodine), phenols, thiophenols, antibiotics, heterocyclics

with anionic groups, nitro compounds, ureas, formaldehyde derivatives and amines [2].

However, many of these chemicals are toxic and show undesirable effects in humans. The use of

antibiotics in these materials has also given rise to a wide scope for the development of multi-

drug resistance microbes, such as: meticillin-resistant Staphylococcus aureus (MRSA) and

vancomycin-resistant Enterococcus (VRE) [1]. But, all these problems can be avoided very

much by the use of metal or metal nano particles [3], which do not act via cell receptors and so

the chance of immune response in microbes to develop resistance is ruled out.

Many studies have indicated that silver is a useful prophylactic and therapeutic agent for

the prevention of wound colonization by organisms. Silver is also considered as a broad-

spectrum antimicrobial agent that controls yeast, mold and bacteria, including methicillin-

resistant staphylococcus aureus (MRSA) and vancomycin-resistant enterococci (VRE) [4,5]. The

acceleration of delayed wound healing and rapid diabetic wound healing [6,7] are the additional

characteristics of silver nano particles (AgNPs), as reported by Tian et al [6] and Mishra et al [7].

Hence, the use of silver as nano particles is always a better choice to curb the antimicrobial

growth.

Silver nano particles produced by chemical reducing agents are usually associated with

environmental toxicity or biological hazards. Consequently, the development of AgNPs based on

environmentally-friendly and biodegradable reagents is always preferred. From this perspective,

naturally occurring carbohydrate polymers, gum acacia (GA) (Gum Arabic) consisting mainly of

β-(1→3) galactose units in the backbone with branches of arabinose and rhamnose [8] and guar

gum (GG) consisting, mainly of (1→4)-β-D-mannopyranosyl backbone with branch-points from

the six-position linked to single α-D-galactopyranosyl residues [9], have been utilized for the

formation and stabilization of AgNPs [10-12]. These carbohydrate polymers are readily available

not only at low cost, but also in abundance in nature and have excellent emulsifying and surface-

active properties [13-15]. Therefore, these carbohydrate polymers are highly beneficial in

designing antimicrobial wound dressing materials.

Surgical cellulose roller gauge bandage is a specially designed functional clothing of

cellulose (a homopolymer of α-D-glucopyranose units linked together by (1→4)-glycosidic

bonds) [16], usually used as a secondary dressing layer over the primary wound dressing

materials. This bandage, amongst others, holds primarily wound dressing, supports injured

wound, absorbs blood and secretions exudates and provides pain relief. It serves as the outermost

protective dressing for wound, thereby preventing wound contamination from the outside

environment. Hence, imparting anti-microbial activity in surgical bandages is duly essential to

make them effective as adequate barriers against highly communicable bacteria and fungus. It

was also concluded from the review of Miller and co-workers that the patients who are treated

the treated wounds with silver containing dressing had a significantly quicker healing rate than

equivalent non-silver containing dressing [5].

In view of the above discussion the present investigation is an attempt to develop

microbial resistant transdermal wound dressing scaffolds from the cellulose roller gauze bandage

from natural carbohydrate polymers, gum acacia powder (GA), and gaur gum powder (GG). The

scaffolds (Poly-AgNPs bandages) that are developed, exhibited excellent antimicrobial activity

when tested against E. coli (gram negative bacteria), S.aureus (gram positive bacteria) and C.

Albicans (fungus). Hence, the scaffolds developed in this work from natural carbohydrate

polymers via an environmentally-friendly green process can be used as effective microbial

resistant transdermal scaffolds for wound care applications.

3.2 Experimental

3.2.1 Materials

Gum acacia powder (GA) and gaur gum powder (GG) and silver nitrate (AgNO3) were

purchased from S.D. Fine-Chem Ltd. (Mumbai, India) and used as received without further

purification. Surgical cellulose roller gauze bandage (dimension 50 mm x 100 mm) was provided

by the Government of Andhra Pradesh to the Primary Health Centre (S.K. University,

Anantapur, India), was used. Double distilled water was used throughout the experiments.

3.2.2 Fabrication of Poly-AgNPs Bandage (PAB)

To prepare PABs (Poly-Silver Nano Particles bandage), 100 mL of GA and GG solutions of

varying concentrations: 0.3%, 0.5% and 0.7% (w/v) were initially prepared by stirring respective

amounts of GA and GG, separately in double distilled water at 300 rpm in an orbital shaking

incubator at 27 oC for 24 h. Then, 5 mL of 0.588 mM of AgNO3 solution was introduced to each

of the stirring solutions and stirring was continued until a constant value in the intensity of

absorption corresponding to AgNPs in the UV-Visible spectrum, was observed. The swelling

method was carried out in order to introduce the AgNPs over the cellulose roller gauge bandage,

for which 10 strips of cellulose roller gauge bandage were immersed in each of the solutions and

stirred for 24 h at 27 oC, followed by sonication for 30 min. Finally, the Ag

0 (silver nano

particles) loaded cellulose bandages (PABs) were obtained. PABs obtained from the various

formulations of GA and GG were taken out dried and kept in desiccator prior to characterization.

3.2.3 Characterizations

UV-Visible spectra of silver nano particles were recorded with UV-Vis

spectrophotometer (Elico-SL 164, Hyderabad, India). Bonding interaction of AgNPs with

cellulose roller gauge bandage was established by performing Fourier transform infrared (FTIR)

measurement on Perkin Elmer (Model Impact 410, Wisconsin, MI, USA) spectrophotometer.

Transmission electron microscopy (TEM) (JEM-1200EX, JEOL, Tokyo, Japan) was carried out

by dispersing two to three drops of (1 mg/1 mL) poly-AgNPs solution on a 3mm copper grid.

Thermal properties were determined using the TGA data, using SDT Q 600 thermal analyzer

(TA Instruments-water LLC, Newcastle, DE, USA), at a heating rate of 20 oC/min by passing

nitrogen gas at a flow rate of 100 mL/min. Scanning electron microscopy-energy dispersive

spectroscopy (SEM-EDS) analysis was performed using a JEOL JEM-7500F (Tokyo, Japan),

operated at an accelerating voltage of 3 kV. All (SEM-EDS) samples were gold coated, prior to

examination on the field emission scanning electron microscope.

3.2.4 Swelling ratio studies

Normally, the swelling ratio plays a significant role in biomedical applications,

particularly for antibacterial applications [17]. In the present investigation, the swelling ratios of

PABs were measured at ambient temperature by using gravimetric method. Initially, known

weights of dried PABs were immersed in 50 mL of double distilled water until their weight

becomes constant. The PABs were then removed and their surfaces were blotted with filter paper

and weighed.

The swelling ratio or swelling capacity, S(g/g) of the PABs developed, was calculated

using the equation:

Swelling ratio (Sg/g) = [Ws - Wd]/ Wd (1)

Where, Ws and Wd denote the weight of the swollen cellulose bandage at equilibrium and

the weight of the dry cellulose bandage, respectively. The data provided is an average value of

three individual sample readings. This data is presented in the form of figure in Fig.7 under

Results and Discussion section.

3.2.5 Antimicrobial activity studies

For testing antimicrobial activity of the developed PABs, E. coli (gram negative bacteria,

MTCC-1668), S.aureus (gram positive bacteria, MTCC-7443) and C. Albicans (fungus, MTCC-

7315) were chosen as model microbes over modified agar diffusion assay maintained at 37oC for

an incubation period of 24 h. The nutrient agar medium required was prepared by mixing

peptone (5.0 g), beef extract (3.0 g) and sodium chloride (5.0 g) in 1000 mL of distilled water

and the pH was adjusted to 7.0. Finally, agar (15.0 g) was added to the solution. Sterilization of

agar medium was done in a conical flask in an autoclave (MAC, MACRO scientific works Pvt.

LTD., Delhi) at a pressure of 6.8 kg (15 lbs) for 30 min. This medium was transferred into

sterilized petri dishes in a laminar air flow chamber (Microfilt Laminar Flow Ultra Clean Air

Unit, Mumbai, India). After solidification, 50µl of microbial culture was spread in the petri

dishes. Over the inoculated petri dishes, small pieces of PABs and polymer coated roller gauge

were distributed and incubated for 24 h in an incubation chamber at 37 oC. The inhibition zones

formed by the microbes were measured and photographed. The data of this investigation is given

under antibacterial properties of Results and discussion section.

3.3 Results and discussions

In the present investigation, microbial resistant scaffolds were developed by

impregnating the AgNPs synthesized from natural carbohydrate polymers (GA and GG) over the

cellulose bandage. The procedure involves two steps, as shown, schematically, in Scheme 1: (i)

Preparation of polymer Ago (silver nano particles) solution (poly-AgNPs solution) via a green

reduction process and (ii) Fabrication of polymer Ago-loaded cellulose bandage (PAB) via

swelling method. The loaded AgNPs were considered to bond over cellulose roller gauge

bandage by physical interactions with the surface hydroxyl groups of cellulose [16].

The materials (gum acacia, guar gum and cellulose fibre) used in the present investigation

to develop CSNCFs are naturally available.

Gum acacia (GA), a natural polysaccharide derived from exudates of Acacia senegal and

Acacia seyal trees. It is commonly used as food hydrocolloid. Chemically, it consists β-(1→3)

galactose units in the backbone with branches of arabinose and rhamnose [18].

Guar gum

(GG) is primarily the ground endosperm of guar beans which consists,

mainly (1→4)-β-D-mannopyranosyl backbone with branch-points from the six-position

Acacia seyal tree Gum Acacia

linked to single α-D-galactopyranosyl residues [19]. Guar gum is an effective

hypocholesterolemic agent and prevents hypercholesteromia and reduces body weight.

Cellulose fibre consists majorly cellulose, a homopolymer of α-D-glucopyranose units

linked together by (1→4)-glycosidic bonds [16]. Cellulose is the most abundant organic polymer

on the Earth and present in the primary cell wall of the plants. Cellulose fibres that are naturally

present in cotton consists 90 % of cellulose. These cellulose fibres are used in textile industries

and also to design wound dressing materials/scaffolds. The surgical cellulose roller gauge

bandage used in the present investigation is a specially designed functional clothing of cellulose,

processed under hygienic environment that satisfies medical standards.

Guar gum Guar beans

Cotton Cellulose

Scheme 1: Schematic diagram of preparation of Poly-AgNPs bandages (PABs): i)

Preparation of Polymer Ago solution (poly-AgNPs solution) via green process reduction; ii)

Fabrication of Polymer Ago-loaded cellulose bandage (PAB) via swelling method.

3.3.1 UV-Visible spectral analysis

The initial formation of silver nano particles in GA and GG solutions (polymer solutions)

was established by colour change (form colourless to ruby red) and was confirmed by observing

a broad intense absorbance band formation in the UV–Vis spectra between 430–450 nm, as

shown in Fig.1. This was due to the surface plasmon resonance excitation vibrations of the silver

nano particles [18, 19]. Absorption peaks for concentrations: 0.3%, 0.5% and 0.7% were

observed at 432, 435 and 436 nm for GA (Fig 1A) and at 434, 438 and 442 nm for GG (Fig 1B),

respectively. The formation of AgNPs in the GA/GG solutions was due to the reduction reaction

of the pendent hydroxyl groups present in GA and GG [20]. The AgNPs formed were stabilized

by the high molecular chains present in GA and GG [10]. From Fig.1, it is concluded that the

intensity of absorption peak increases with increasing concentration of the corresponding GA

and GG polymers. It is also evident that for the same concentration, GG-AgNPs solution showed

absorption maxima at higher wavelength (red shift) than GA-AgNPs solution. The red shift was

either due to an increase in size or aggregation of AgNPs [21]. The same can be confirmed by

TEM analysis, where the GG-AgNPs were identified to be larger in size than GA-AgNPs (Fig 2).

Fig

1:

UV

Absorption spectra of Poly-AgNPs solutions of varying concentrations: A) GA-AgNPs

solutions; B) GG-AgNPs solutions.

3.3.2 Transmission electron microscopic (TEM) analysis

The formation of AgNPs in polymer solutions (GA, GG) was confirmed from TEM

analysis. Further, it is also established by TEM analysis, as shown in Fig 2, that the GG-AgNPs

(Fig 2B) with larger size (5±3 nm) than GA-AgNPs (4±2 nm) (Fig 2A), were formed. The

absence of aggregation of the AgNPs in both solutions may be attributed due to the effective

passivation of the surfaces and the suppression of the growth of the nano particles through strong

interactions via the functional molecular groups of GG (-OH) and GA (-OH, -COOH). From

TEM, it is also clear that the overall AgNPs formed were spherical in shape with an average

particle size of in between 2 to 8nm. This is quite expected since GA and GG stabilize the

formed AgNPs by a well-established chemical interaction between the functional groups [22,10]

and silver nano particles.

Fig 2: TEM images of AgNPs formed in: A) 0.7% GA solution; B) 0.7% GG solution.

3.3.3 Fourier transform infrared (FTIR) analysis

In order to examine the formation of AgNPs over poly-AgNPs-coated cellulose bandages

(PABs), the bonding characteristics through Fourier transform infrared (FTIR) spectra was

studied. FTIR spectra of the pure cellulose bandage, GA and GG coated bandage and PABs are

shown in Fig. 3. The shifts in the functional groups positions in PABs confirm the interaction of

AgNPs and GA/GG and the cellulose ban

of pure cellulose bandage, the polymer

respectively.

In the FTIR spectrum of pure cellulose bandage, the following characteristic peaks are

observed:

A large band at 3390 cm

bonded -OH groups; a peak at 2921 cm

corresponding to the C–H bending mode,

assigned to the C–O–C stretching of the glucosidic units or from

[23,24]. In the FTIR spectra of polymer

cm-1

; GG: 3380, 2932, 1308, 1118, 1034 cm

1129,

peaks

bands of

bandages are

result, it can be inferred that poly


AgNPs and GA/GG and the cellulose bandage. Fig. 3 (A) and Fig. 3(B) shows the FTIR spectra

of pure cellulose bandage, the polymer-coated bandage and poly-AgNPs-coated bandage (


A large band at 3390 cm-1

corresponding to the stretching vibration of the hydrogen

OH groups; a peak at 2921 cm-1

due to >CH2 groups; a peak around 1

H bending mode, and the characteristic peaks at 1150 and 1015 cm

C stretching of the glucosidic units or from β-(1 →4)-glucosidic bonds

the FTIR spectra of polymer-coated bandages (GA: 3359, 2920, 1306, 1108, 1035

; GG: 3380, 2932, 1308, 1118, 1034 cm-1

) and PABs (GA: 3358, 2932, 1318, 1129, 1027

cm-1

; GG: 3358, 2931, 1317,

shifted to lower values. A

result, it can be inferred that poly-AgNPs are present in the cellulose bandages.


B) shows the FTIR spectra

coated bandage (PAB),


corresponding to the stretching vibration of the hydrogen

a peak around 1315.59 cm−1

,

and the characteristic peaks at 1150 and 1015 cm-1

are

glucosidic bonds

A: 3359, 2920, 1306, 1108, 1035

) and PABs (GA: 3358, 2932, 1318, 1129, 1027

; GG: 3358, 2931, 1317,

1037 cm-1

), the

of the

characteristic

cellulose

shifted to lower values. As a

Fig

cellulose

coated and

Ago-coated

bandages;

cellulose bandage, 0.7 % GG

coated bandages

3.3.4 Scanning electron microscopy

The presence of AgNPs was confirmed from SEM analysis, as shown in Fig 4.

surface morphology of pure cellulose bandage (Fig 4A(a) and Fig 4B(a)), GA

(Fig 4A(b)), GG-coated bandage (Fig 4B(b)), GA

are presented in Fig 4. The surface morphology of the polymer

bandages (Fig 4A(b) and Fig 4B(b)) reveals no AgNPs on the s

and GG PABs) (Fig 4A(c) and Fig 4B(c)) shows the existence of AgNPs

aggregations (indicated with arrows).

The EDS spectrum of the pure bandages

spectra of A)

bandage, 0.7 % GA

cellulose bandage, 0.7 % GG coated and 0.7 % GG

3.3.4 Scanning electron microscopy-energy dispersive spectroscopic (SEM-EDS) analysis

The presence of AgNPs was confirmed from SEM analysis, as shown in Fig 4.

surface morphology of pure cellulose bandage (Fig 4A(a) and Fig 4B(a)), GA

coated bandage (Fig 4B(b)), GA-PABs (Fig 4A(c)) and GG-PABs (Fig 4B(c))

are presented in Fig 4. The surface morphology of the polymer-coated (both GA and GG

bandages (Fig 4A(b) and Fig 4B(b)) reveals no AgNPs on the surfaces, whereas PABs (both GA

and GG PABs) (Fig 4A(c) and Fig 4B(c)) shows the existence of AgNPs

aggregations (indicated with arrows). The EDS spectra shown in Fig 5, support the SEM data

pure bandages (Fig 5(a)) shows no characteristic peak of silver, while

3: FTIR

spectra of A)

bandage, 0.7 % GA-

0.7 % GA-

B)

coated and 0.7 % GG-Ago-

EDS) analysis

The presence of AgNPs was confirmed from SEM analysis, as shown in Fig 4. The

surface morphology of pure cellulose bandage (Fig 4A(a) and Fig 4B(a)), GA-coated bandage

PABs (Fig 4B(c))

coated (both GA and GG-coated)

urfaces, whereas PABs (both GA

and GG PABs) (Fig 4A(c) and Fig 4B(c)) shows the existence of AgNPs without any

support the SEM data.

s no characteristic peak of silver, while

the GA-PABs (Fig 5(b)) and GG

silver, confirming the existence of elemental silver nano particles, Ag

and GG-PABs (Fig 5(c)) clearly shows the characteristic peaks of

confirming the existence of elemental silver nano particles, Ago in PABs.

clearly shows the characteristic peaks of

Fig

5:EDS

spectrum

of (a)

cellulose

bandage(b) 0.7 GA-PABs (c) 0.7 GG

3.3.5 Thermal (TGA) analysis

Thermal properties remained as a supporting evidence for the presence of AgNPs in the

PABs. The thermal properties of PABs were examined with reference to pure cellulose ba

by thermogravimetric analysis (TGA). The primary thermograms are GA

GG-PABs (Fig 6B) are shown in Fig 6. The results indicated that, in all the samples, there is an

initial weight loss at a temperature below 100

present on the surface. The maximum decomposition of all the PABs was occurred at a slightly

higher temperature, around 399.72

studies lead to the conclusion that with i

PABs the stability of PABs also increased. This might be due to the rise in the number of nano

particles on the surface of the PABs, which is due to the development of strong bonding

interactions between AgNPs and the cellulose surface of PABs [13]. This reason might be quite

expected as the TGA residue at 600

GG concentrations (due to the increased deposition of AgNPs on PABs). Overall, the therma

analysis indicates that the developed and designed PABs were thermally stable due to more

percentage of AgNPs.

Fig

PABs (c) 0.7 GG-PABs


PABs. The thermal properties of PABs were examined with reference to pure cellulose ba

by thermogravimetric analysis (TGA). The primary thermograms are GA-PABs (Fig 6A) and

PABs (Fig 6B) are shown in Fig 6. The results indicated that, in all the samples, there is an

initial weight loss at a temperature below 100 oC, due to the loss of moisture (water molecules)


higher temperature, around 399.72 oC, when compared to that of pure bandage. The thermal

studies lead to the conclusion that with increase of GA/GG percentage (%) in the corresponding



AgNPs and the cellulose surface of PABs [13]. This reason might be quite

expected as the TGA residue at 600 o

C also increased proportionately with increase in GA and

GG concentrations (due to the increased deposition of AgNPs on PABs). Overall, the therma



PABs. The thermal properties of PABs were examined with reference to pure cellulose bandage

PABs (Fig 6A) and

PABs (Fig 6B) are shown in Fig 6. The results indicated that, in all the samples, there is an

of moisture (water molecules)


C, when compared to that of pure bandage. The thermal

ncrease of GA/GG percentage (%) in the corresponding



AgNPs and the cellulose surface of PABs [13]. This reason might be quite

C also increased proportionately with increase in GA and

GG concentrations (due to the increased deposition of AgNPs on PABs). Overall, the thermal


6: Thermo-

gravimetric analysis of: A) GA

3.3.6 Swelling studies

The swelling ratio is a significant parameter that i

developed bandage for blood and secretion exudates from injured wounds. The swelling

behaviours of pure cellulose bandage (C), polymer

PABs of GA (Fig 7A) and GG (Fig 7B) are show

influenced by the concentration of GA/GG in the polymer. With an increase in the polymer

concentration, increased swelling ratio values were observed. This is due to the hydrophilic

nature of GA and GG. The overall swelling studies shows that the swelling ratios of polymer

coated bandages were greater than the PABs and pure bandage. This lowering in the swelling

ratio of PABs can be attributed due to the interaction of poly

and other groups present in polymer

cross-links with in the bandage networks. Consequently, the increase in swelling phenomenon of

polymer coated bandages over pure cellulose bandage can play significa

applications, particularly in antimicrobial applications. Based on this characteristic, many

researchers have prepared different types of material for antibacterial applications [25

Fig 7: Swelling ratio of A) cellulose bandage a

bandages (brown); B) cellulose bandage and GG

bandages (brown).

3.3.7 Antimicrobial properties

gravimetric analysis of: A) GA-PABs; B) GG-PABs

The swelling ratio is a significant parameter that indicates the absorption capacity of the


behaviours of pure cellulose bandage (C), polymer-coated bandages (GA/GG bandages) and

PABs of GA (Fig 7A) and GG (Fig 7B) are shown in Fig 7. The values of the swelling ratio were

fluenced by the concentration of GA/GG in the polymer. With an increase in the polymer


overall swelling studies shows that the swelling ratios of polymer


ratio of PABs can be attributed due to the interaction of poly-AgNPs with electrons of hydroxyl

d other groups present in polymer-cellulose bandage matrices, thereby producing additional

links with in the bandage networks. Consequently, the increase in swelling phenomenon of

polymer coated bandages over pure cellulose bandage can play significant role in biomedical


researchers have prepared different types of material for antibacterial applications [25

Swelling ratio of A) cellulose bandage and GA-coated bandage (black), GA

bandages (brown); B) cellulose bandage and GG-coated bandage (black), GA

ndicates the absorption capacity of the


coated bandages (GA/GG bandages) and

n in Fig 7. The values of the swelling ratio were

fluenced by the concentration of GA/GG in the polymer. With an increase in the polymer


overall swelling studies shows that the swelling ratios of polymer-


AgNPs with electrons of hydroxyl

cellulose bandage matrices, thereby producing additional

links with in the bandage networks. Consequently, the increase in swelling phenomenon of

nt role in biomedical


researchers have prepared different types of material for antibacterial applications [25-28].

coated bandage (black), GA-PABs

coated bandage (black), GA-PABs

The efficiency of the developed PABs as microbial resistant transdermal materials was

determined by conducting the antimicrobial activity against E. coli (Fig 8(A1, B1)), S.aureus

(Fig 8(A2, B2)) and C. Albicans (Fig 8(A3, B3)). The results revealed a strong reduction in the

number of microbial colonies around the samples. The inhibition zone for all the PABs shown as

in Fig.8 was found to be higher than 2.0 mm. According to the Standard Antibacterial test “SNV

195920–1992”, specimens showing more than 1 mm microbial zone inhibition can be considered

as good antibacterial agents [29]. Hence, the PABs that are developed in the present

investigation by a different approach can be considered as good antibacterial agents that are (and

should) effective in killing the microbes. It is noticed that with increasing concentration of

GA/GG in the PABs, the antimicrobial activity also increased. This is due to the formation of

more number of AgNPs on the PABs with increase in concentration of GA/GG. It is also noticed

that for the same concentration, GG-PABs showed higher inhibition zone than the GA-PABs.

This is quite expected due to more number of AgNPs are accumulated on the surface of GG-

PABs than on the GA-PABs (this is evident from TGA and SEM-EDS data). This conclusion is

supported by the exceptional high inhibition zone attained, as shown in Table 1.

Fig

8:

Antibacterial activity of: A) (o) Pure cellulose bandage, (a) 0.3 GA-PAB, (b) 0.5 GA-PAB,

(c) 0.7 GA-PAB and B) (o) Pure cellulose bandage, (a) 0.3 GG-PAB, (b) 0.5 GG-PAB, (c)

0.7 GG-PAB against E. coli (A1, B1), S.aureus (A2, B2) and C. Albicans (A3, B3).

Microbial culture Inhibition zone of GA-PABs (mm) Inhibition zone of GG-PABs (mm)

0.3 GA-

PAB

0.5 GA-

PAB

0.7 GA-

PAB

0.3 GG-

PAB

0.5 GG-

PAB

0.7 GG-

PAB

E. coli 4.5 4.8 5 4.5 5.2 5.8

S. aureus 3.2 3.5 4 4.2 4.4 5

C. Albicans 3 3.2 4 3.2 4.1 4.8

Table 1: Inhibition zone exhibited by various PABs

Mechanism proposed for the antibacterial activity

The mechanism of growth inhibitory effect of AgNPs against the microorganisms is not

clear [30]. However, among the various possible mechanisms proposed by many authors [30],

‘Inhibition by formation of pits,’ was the mechanism assumed here. The initial contact of PABs

with the microbes allows AgNPs to interact with the cell wall of the microbe membrane to form

pits. These pits cause the leakage of lipopolysaccharide molecules and membrane proteins of

bacteria/fungus, leading to microbial death [31,32]. The same can be concluded from Table 1,

where the inhibition zone observed for E.coli (containing thin peptidoglycan cell wall layer) is

more than that of S.aureus (thick peptidoglycan layer) [33]. This mechanism was further

supported by the studies of Singh et al, which postulated that nano particles formed in the 1-10

nm range, attach to the surface of the cell membrane and drastically disturb its proper function

by forming pits [34]. The AgNPs formed in the current approach is also in between 2-8 nm

range, which supports strongly the proposed mechanism.

3.4 Conclusion

Silver nano particles were developed from natural carbohydrate polymers: Gum acacia

(GA) and Gaur gum (GG). The silver nano particles developed, when coated as poly-AgNPs on

cellulose roller gauge bandage, exhibited excellent antimicrobial activity against E. coli (gram

negative), S.aureus (gram positive) and C. Albicans (fungus). Unlike the use of conventional

reducing agents for the synthesis of AgNPs, which causes environmental toxicity, the use of

naturally occurring polymers as reducing and stabilizing agents is highly advantageous, based on

the environmental concern. Hence, the scaffolds developed by this environmentally-friendly

green process can be used, effectively as microbial resistant transdermal scaffolds in biomedical

applications.

The work presented in this investigation is an important contribution in the field of

development of antibacterial surgical cellulose roller gauge bandage using green process

for biomedical applications.

The work developed in the present studies has been applied for a Patent and it has

crossed all the preliminary stages and is in the final review process and hoping for final

approval.

3.5 References

1. T. Ristic, L. Z. Zemljic, M. Novak, M. K. Kuncic, S. Sonjak, N. G. Cimerman and S.

Strnad, Science against microbial pathogens: communicating current research and

technological advances, 0 (2011), 36-51.

2. Y. Gao and R. Cranston, Text Res J, 78(2008), 60-72.

3. N. S. Z. Haider, U. Kiran, I. M. Ali, A. Hameed and S. A. N. Ali, Brno, Czech Republic,

EU, 10(2012), 23 - 25

4. S. Bishara, A. Atiyeh, Michel Costagliola, N. Shady, Saad A. Hayek and Dibo, Burns,

33(2007), 139-48.

5. C. Miller, Communicating current research and technological advances, A. Méndez-

Vilas (Ed.), 1(2011),14-22.

6. J. Tian, K. K. Wong, C. M. Ho, C. N. Lok, W. Y. Yu, C. M. Che, J. F. Chiu and P. K.

Tam, Chem Med Chem, 2(2007), 129 – 136.

7. M. Mishra, H. Kumar and K. Tripathi, Dig J Nanomater Bios, 2008(2008), 49-54.

8. D. Yael, C. Yachin and Rachel Yerushalmi-rozen, J. Appl. Polym. Sci. B, 44(2006),

3265–3271.

9. S. Saeed, H. Mosa-Al-Reza, S. D. Atiyeh and A. A. Marziyeh, J. Phyt. Med., 1(2011),

36-42.

10. E. S. Abdel-Halim, M. H. El-Rafie, and Salem. S. Al-Deyab, Carbohydr. Polym.,

85(2011), 692-697.

11. K. Vimala, K. Samba Sivudu, Y. Murali Mohan, B. Sreedhar and K. Mohana Raju,

Carbohydr. Polym., 75(2009), 463-471.

12. Y. Park, Y. N. Hong, A. Weyers, Y. S. Kim and R. J. Linhardt, IET Nanobiotechnol,

5(2011), 69–78.

13. G. M. Raghavendra, T. Jayaramudu, K. Varaprasad, R. Sadiku, S. Sinha Ray and K.

Mohana Raju, Carbohydr. Polym., 93(2013), 553– 560.

14. M. Akhtar, E. Dickinson, J. Mazoyer and V. Langendorff, Food Hydrocolloids,

16(2002), 249-256.

15. K. S. Mikkonen, M. Tenkanen. P. Cooke, C. Xu, H. Rita, S. Willfo, B. Holmbom, K. B.

Hicks and M. P. Yadav, LWT - Food Sci. Technol. Int., 42(2009), 849–855.

16. D. J. Gardner, G. S. Oporto, R. Mills and M. A. S. Azizisamir, J. Adhes. Sci. Technol.,

22(2008), 545-567.

17. T. Jayaramudu, G. M. Raghavendra, K. Varaprasad, R. Sadiku and K. Mohana Raju,

Carbohydr. Polym., 92(2013), 2193–2200.

18. N. Duran, P. Marcato, G. I. H. De Souza, O. L. Alves and E. Esposito, J. Biomed.

Nanotechnol., 3(2007), 203–208.

19. S. S. Shankar, A. Rai, A. Ahmad and M. Sastry, J. Colloid Interface Sci, 275 (2004),

496–502.

20. D. Dorjnamjin, M. Ariuna and Y. K. Shim, Int. J. Mol. Sci., 9(2008), 807-820.

21. M. A Hettiarachchi and P. A. S. R. Wickramarachchi, Journal of Science of Kelaniya Sri

Lanka, 6(2011), 65-75.

22. B. Sreedhar, D. Keerthi Devi and Deepthi Yada, Catal. Commun., 12(2011), 1009–1014.

23. D. M. Suflet, G. C. Chitanu and Pop VI, React Funct Polym, 66 (2006), 1240–1249.

24. Shu-Ming Li, NingJia, Jie-Fang Zhu, Ming-Guo Ma, FengXu, Bo Wang and Run-Cang

Sun, Carbohydr. Polym., 83(2011), 422–429.

25. P. S. K. Murthy, Y. Murali Mohan, K. Varaprasad, B. Sreedhar and K. Mohana Raju, J.

Colloid Interface Sci., 318(2008), 217–224.

26. K. Varaprasad, K. Vimala, S. Ravindra, N. Narayana Reddy, G. Venkata Subba Reddy

and K. Mohana Raju, J. Mater. Sci. Mater. Med., 22(2011), 1863-1872.

27. S. Ravindra, Y. Murali Mohan, N. Narayana Reddy and K. Mohana Raju, Colloids Surf.

A, 367(2010), 31–40.

28. P. Ranga Reddy, K. Varaprasad, N. Narayana Reddy, K. Mohana Raju and N. Subbarami

Reddy, J. Appl. Polym. Sci., 125(2012), 829-1656.

29. M. Pollini, M. Russo, A. Licciulli, A. Sannino and A. Maffezzoli, J. Mater. Sci. Mater.

Med., 20(2009), 2361–2366.

30. H. H. Lara, E. N. Garza-Treviño, L. Ixtepan-Turrent and K. S. Dinesh Silver, J

Nanobiotechnology, (2011), 9-30.

31. N. A. Amro, L. P. Kotra, K. Wadu-Mesthrige, A. Bulychev, S. Mobashery and G. Liu,

Langmuir, 16(2000), 2789-2796.

32. A. Nasrollahi, Kh. Pourshamsian and P. Mansourkiaee, Int.J.Nano.Dim, 1(2011), 233-

239.

33. S. H. Kim, H. S. Lee, D. S. Ryu, S. J. Choi and D. S. Lee, Korean J. Microbiol.

Biotechnol, 39(2011), 77–85.

34. M. Singh, S. Singh, S. Prasada and I. S. Gambhir, Dig J Nanomater Bios. 3(2008), 115-

122.

Development of microbial resistant -...

Documents

Transcript of Development of microbial resistant -...