Application of Calophyllum inophyllum oil as antifungal fat-liquor for leather industry

Application of Calophyllum inophyllum oil as antifungal fat-liquor for leather industry

Industrial Crops & Products 105 (2017) 104–112 Contents lists available at ScienceDirect Industrial Crops & Products journal homepage: www.elsevier...

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Industrial Crops & Products 105 (2017) 104–112

Contents lists available at ScienceDirect

Industrial Crops & Products journal homepage: www.elsevier.com/locate/indcrop

Application of Calophyllum inophyllum oil as antifungal fat-liquor for leather industry

MARK



Bindia Sahua, , Aravindhan R.b, Mohammed Abu Javidb a b

CHORD Division, Central Leather Research Institute, Council of Scientific and Research, Adyar, Chennai 600020, India Tannery Division, Central Leather Research Institute, Council of Scientific and Research, Adyar, Chennai 600020, India

A R T I C L E I N F O

A B S T R A C T

Keywords: Calophyllum inophyllum oil. Bishop’s weed Antifungal fat liquor Gas chromatograph with a mass detector (GC/ MS) analysis Thymol Carvacrol

Calophyllum inophyllum oil which contains mixed oleic, linoleic acid, stearic acid and palmitic acid is well known for its antimicrobial properties. Presence of unsaturation leads to provide better opportunity for its modification for fat liquoring. Trans esterified emulsion of Calophyllum inophyllum oil shows good antifungal property. To enhance its antifungal activity against more fungal species the blending of trans esterified emulsion with water extract of Bishop’s weed leads to the formation of an antifungal fat liquor. This blended product mitigates most of the disadvantages associated with conventional tanning. Fresh mature cultures of fungal species such as, Aspergillus Niger, Penicillium notatum, Aspergillus flavus and Paecilomyces variotii have been collected and used as sources of inoculum for antifungal activity of the fat liquor. Gas chromatograph with a mass detector (GC/MS) analysis of both Calophyllum inophyllum oil and Bishop’s weed were carried out separately. The presence of unsaturated acids like oleic acid and linoleic acid provide the platform for its modification by transesterification process and the presence of two compounds namely thymol and carvacrol in the Bishop’s weed enhances the antifungal activity of fat liquor. The antifungal activity of fat liquor is confirmed by Agar well diffusion method. The aim of this study is to investigate the possibility of using Calophyllum inophyllum oil as antifungal fat liquor for leather industry.

1. Introduction In leather processing hide undergo several chemical and mechanical treatments. Chemical treatments of hide release fat content which in turn causes collagen fibers to stick together. In order to prevent this fiber sticking fat liquoring of leather is essential for soft and flexible leather (Liu et al., 2007). Fat liquoring is a beneficial process of wet blue finishing which usually involves introduction of fat into the wet blue and improves both physio–chemical properties of leather (Zarlok et al., 2014). Most of the fat liquors are special softening oils, synthesized by modifying oils by different chemical reaction like sulphonation, sulphation, and trans esterification, in order to insert the oil droplets with the water molecules in the form of fine emulsion into the fibers of skin. In order to avoid sickness of the fibers fat liquoring is essential to separate them apart (Bajza and Vinkovic Vrcek, 2001). The size of the emulsion particles plays a crucial role in fat liquoring. The size of the emulsion particles and pores of skin are closely associated, in order to achieve deep penetration of emulsion compatibility between emulsion and fibers requires (Bajza and Vinkovic Vrcek, 2001). The resulting finished



leather exhibited the enhanced properties like soft and high tensile strength of leather (Sivakumar and Rao, 2001). Calophyllum inophyllum oil is non edible oil. The botanical name is Calophyllum inophyllum. The plant is known by different local names English – Alexandra Laurel, Hindi – Sultana Champa (Shankar Mishra et al., 2010). It is dark green in color and contains both free fatty acids and saturated as well as unsaturated acids such as oleic, palmitic, linoleic, and stearic acids. Calophyllum inophyllum oil is commonly used as component of drugs, biologically active additives, cosmetics, and food industry. In addition, many latest studies report on its biological activity such as antibacterial, anti-inflammatory, antiseptic, fungicidal, antioxidant, and antiradical activities (Sirvaityte et al., 2012a). It has been reported that Calophyllum inophyllum oil contains most important group of compounds such as glycerides, terpenoids, sterols, steroids such as canophyllal, canophyllol, and canophyllic acid and coumarinic derivatives such as calophyllolids, inophyllolids and calophyllic acid which are responsible for its antimicrobial activity (Dweck and Meadowsy, 2002; Kedzia et al., 2011). Bishop’s weed usually known as trachyspermum ammi, or ajwoin, or carom seed is a straight yearly herb with antibacterial activity (Hassan

Corresponding author at: Central Leather Research Institute, Council of Scientific and Research, Adyar, Chennai 600020, India. E-mail addresses: [email protected], [email protected] (B. Sahu).

http://dx.doi.org/10.1016/j.indcrop.2017.04.064 Received 28 May 2016; Received in revised form 28 April 2017; Accepted 30 April 2017 0926-6690/ © 2017 Elsevier B.V. All rights reserved.

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Shahian et al., 2014). Bishop’s weed seeds are small in size, their taste is spicy, and it is stomachic, carminative, laxative and diuretic in nature (Morshed et al., 2012). It has been found that the antibacterial activity of Bishop’s weedis mainly due to presence of two chemical compounds which are, thymol and carvacrol. These compounds have phenolic group in their structure and show high antimicrobial activity against the test microbes (Dwivedi et al., 2012). The thymol and carvacrol chemicals show effective antifungal activity against various fungal species (Mathela et al., 2010). Transesterification is process of reacting of vegetable oils with alcohol, produces a mixture of fatty acids alkyl esters and glycerol in the presence of a strong acid or base as catalyst (Schuchardt et al., 1998). Diglycerides and monoglycerides are the intermediates in this process showing reversibility in the reaction (Meher et al., 2006). It was also observed that when alkali such as sodium hydroxide, sodium acetate and potassium hydroxide are used as catalysts they enhance the rate of reaction much faster than other catalysts. In the formation of fat liquor, the proportion molar ratio of triglyceride and alcohol has found to be plays major role (Schuchardt et al., 1998). An excess of the alcohol favors the formation of the products.

Table 1 Specification of Calophyllum inophyllum oil. 1. Iodine Value:

78 g Iodine/100 g oil

2. Saponification value

200 mg of KOH/gram of oil

3. Fatty acid composition:

Palmitic acid C16: 0 Stearic acid C18: 0 Oleic acid C18: 1 Linoleic acid C18: 2 Docosanoic acid C22: 0 Tetraecosonoicacid C24: 0

9.90% 8.22% 40.25% 30.85% 3.94% 2.6%

4. Sterols:

Campesterol Stigmasterol Beta-Sitosterol

0.70% 2.9% 3.1%

Table 2 Pore size of fat liquor emulsion. No.

Symbol of emulsion

Average particle size (nm)

1 2

U-PEG Castor PEG

25.6 13.5

1.1. Antifungal activity of oil spectrometric detector (7890A series, GC system, with Mass spectrometry MSD with triple axis detector: 2009) in EXCEL lab at CSIR-CLRI, Chennai. Volatile compounds and unsaponifiable matter were separated. Analysis of iodine value and saponification value of oil was also carried out by titration method. Further the Bishop’s weed was collected from local areas of Chennai and extracted in a soxhlet extractor using 150 ml distilled water (100 °C) for 8 h. The extracts were concentrated under vacuum to obtain crude extracts and were stored in desiccators for further studies. At last for antifungal analysis, Potato Dextrose Agar and Agar–Agar was supplied by High Media, Chennai.

Leather is made up of animal skin that has been treated to preserve and make them suitable for use. Leather is a natural product, which can be bio deteriorated by bacteria or fungal species (Rao et al., 1998). Microbes, specially bacteria and fungus are everywhere in the environment. These microbes quickly colonize on food materials, surface, clothing, and on footwear (Sirvaityte et al., 2012b). Most of these organisms are not harmful to human health. However, they may cause infection to a diabetic and asthmatic patients or to infants. Leather can act as carriers for microbial growth. In contact with human body, garments offer a favorable condition for microbes to grow (Sirvaityte et al., 2012b). Vegetable tanned leathers are easily attacked by microbes than chrome tanned leather, but chrome tanned leathers have also been bio deteriorated by microbes. It is required that all the fungicides are effective against a wide range of organisms that bring about deterioration of the leather (Akpomie et al., 2012). Currently 2-(thiocyanomethylthio) benzothiazole (TCMTB), N-OITZ (N-Octylisothiazolinone), OPP (Ortho phenyl phenol), PCMC (p-Chlorom-cresol), Carbendazim, Merkaptobenzothiazole, TCP (tri-chloro phenol), p-Nitro phenol, BMC, DIMTS, etc. be active fungicides that are commonly used in leather industry. However, these chemicals are generally harmful to human health and nature. They cause dermatitis, including itching, irritation, redness, and burns, in some cases, acute respiratory troubles were reported (Sirvaityte et al., 2012b). Because of this researcher are now showing their interest in the use of natural products also in leather tanning (Sirvaityte et al., 2012b). Different part of plant had been used for their antifungal, antimicrobial, insecticidal, activities (Rehman et al., 2015). When wet-blues are attacked by microbes, appearance of pigmentation occurs on it, which leads to a poor finished leather (Akpomie et al., 2012). Hence the main goal of this work is to investigate the possibility of using of product obtained by blending of trans esterified Calophyllum inophyllum with water extract from Bishop’s weed as antifungal fat liquor for leather industry which exhibit high activity when compared with currently used fungicide for leather.

2.1. Transesterification of oil and blending with water extract of Bishop's weed Transesterification reaction was carried out in a 200 ml spherical flask, provided with a thermostat, mechanical stirring, and sampling outlets. Thirty-three grams of oil was weighed and placed in a continuously stirred reactor. An (0.1%) p-Toluene sulfonic acid, catalyst was dissolved in the preferred amount along with proportionate amount of Poly ethylene glycol, and the resulting solution was added to the agitated reactor at 180-°C temperature. The reaction was carried out until it reached the desired reaction time. The addition of water extract of Bishop's weed with trans esterified fatliquor was done after fatliquor preparation with the help of stirrer at 50-°C temperature. 2.2. Physico-chemical characterizations 2.2.1. Particle size analysis of emulsion To analyze the size of the particle, the experiment was carried out on Zeta potential analyzer (Zeta sizer 3000, Malvean instruments HSA: 2004) at Industrial chemistry lab, CSIR-CLRI Chennai. 2.2.2. Spectroscopic and SEM analysis Spectroscopic analysis like Fourier Transform Infrared Spectrophotometer and nuclear magnetic resonance, of trans esterified Calophyllum inophyllum oil and water extract of Bishop’s weed were carried out at Tannery division and NMR lab of CSIR-CLRI respectively. A surface morphology of the leather after treatment with blended trans esterified Calophyllum inophyllum oil with water extract from Bishop’s weed was carried out by scanning electron microscopy (SEM) (A JEOL JSM-5300) at Biophysics lab, CSIR-CLRI.

2. Materials and methods Calophyllum inophyllum oil was purchased from local supplier, Chennai. p-Toluene sulfonic acid (98.5%) was supplied by SigmaAldrich. Poly ethylene glycol (96%) used was purchased from Vijaya scientific company, Chennai. All the other chemicals were obtained commercially and of analytical grade. Quantitative and qualitative analysis of oil was performed on a gas chromatograph with mass 105

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Fig. 1. FTIR of PEG- ester of Calophyllum inophyllum.

Fig. 2.

13

C Spectra of PEG- ester of Calophyllum inophyllum.

106

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Fig. 3. 1H Spectra of PEG- ester of Calophyllum inophyllum. Table 3 Chemical characterization of the Bishop’s weed. Name of compound

Retention time

% of compound

Thymol Gamma terpinene Para cymene Carvacrol pH of water extract of 2%w/v solution mean (n = 3) ± SD Loss on drying (%w/w) mean (n = 3) ± SD

12.70 13.11 15.23 14.22 3.5 ± 0.10 4.8 ± 0.36

60 12 13 10

Process

The amount of % of agent

Agent

Temperature (°C)

Time (min)

Remarks

Washing

200

water

35

15

pour bath

0.5

wetting agent water 35 Sodium formate Sodium bicarbonate water 55

60

ph-5.2

60

pour bath

Neutralization 100 0.5 0.5

Fat liquoring

150 10

Fig. 4. Major chemicals present in Bishop’s Weed.

Fixing

1

Washing

200

fat liquoring agent Formic acid Water

28

30

28

15

Pour bath

2.3. Antifungal properties of oil and fat liquoring emulsion Antifungal efficacy of Calophyllum inophyllum oil, trans esterified Calophyllum inophyllum oil and different blending mixtures like, 1, 1.5 and 2% of wet blue of water extracts from Bishop’s weed with trans esterified Calophyllum inophyllum oil were tested according to standard method (Orlita, 2004). Followed by its comparative study against trans-

2.2.3. Methodology of wet blue leather fat liquoring After preparation of emulsion the resulted emulsion was applied on leather by following method. Methodology for wet blue leather fat liquoring. 107

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Fig. 5. FTIR of water extract from Bishop’s weed.

Fig. 6. 1H NMR of water extract from Bishop’s weed.

antifungal activities of Calophyllum inophyllum oil, trans esterified Calophyllum inophyllum oil and of blending mixtures of trans esterified Calophyllum inophyllum oil with 2% of the wet blue of water extracts from Bishop’s weed.

esterified vegetable oil fat liquor, synthetic and semisynthetic fat liquor, was also carried out. Potato Dextrose Agar (PDA) media of volume 20 ml was taken in petri plates under sterile condition and treated leather specimen with product was placed in the middle of each petri plates followed by incubated at 28 °C temperature, for 72 h. against four fungal species like Aspergillus Niger, Aspergillus flavus, Penicillium notatum, and Paecilomyces variotii. Zone of inhibition provides information about

2.4. Leather quality indexes Leather was dried in free state after fat liquoring with blending 108

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Fig. 7. SEM image of Vegetable oil treated leather: Magnificatication at a) 300 μm b) 100 μm and c) cross section.

Fig. 8. Shows the SEM image of leather treated with Fat liquor made from 10% weight of wet blue of trans esterified Calophyllum inophyllum oil based emulsion. a) 300 μm b) 100 μm and c) cross section.

2.4.1. Leathers shrinkage temperature Determination rules to consists of temperature measurement in which leather sample shrinkage starts in gradually heated glycerin solutions according to standard method- SATRA 17.

Table 4 Antifungal activity of Calophyllum inophyllum oil against four fungal species. Fungal Species

Zone of inhibition (ZOI) (mm) of Leather species without oil as control

Zone of inhibition (ZOI) (mm) of Calophyllum inophyllum oil

Aspergillus Niger Paecilomyces variotii Aspergillus flavus Penicillium notatum

0 0

7 5

0 0

4 3

2.4.2. Strength properties of the leather Strength characteristics of the fat liquored leathers such as tensile strength, (ISO 3376:2002/IUP 6/SATRA TM 43) and tear strength (SATRA TM 162 ISO 3377:2002/IUP 8 DOUBLE EDGE TEAR/IUP 40 SINGLE EDGE TEAR) and lastometer test were tested in CSIR-CLRI.

Table 5 Antifungal activity of Trans esterified Calophyllum inophyllum oil in comparison with synthetic fat-liquor, semi synthetic fat-liquor, trans esterified vegetable oil against fungal species such as AspergillusNiger. Fat liquor

Zone of inhibition (mm)

Trans esterified Calophyllum inophyllum oil Synthetic fat-liquor Semi synthetic fat-liquor Trans esterified vegetable oil (control) With Commercial fungicide Without Commercial fungicide

20 20 21 13 0 0

2.4.3. Analysis of fat content in leather Fat content of the leather was determined by soxhlet extraction method using petroleum ether as solvent according to standard method EN ISO 4048.

The% of fats in leather Weight of Fats by soxhlet method (gm) = × 100 Weight of leather sample taken for soxhlet analysis (gm)

mixtures of trans esterified Calophyllum inophyllum oil with 2% of the wet blue of water extracts from Bishop’s weed, then they were subjected for following testing. 109

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Table 6 Antifungal properties of products obtained by blending of 1, 1.5 and 2% of wet blue of water extracts from Bishop’s weed with trans Esterified Calophyllum inophyllum oil against Aspergillus Niger. Paecilomyces variotii and Aspergillus flavus on wet blue. Fungal species

Zone of inhibition (mm) Fat liquor Blended trans esterified Calophyllum inophyllum Oil with 1% of wet blue Of water Extract of Bishop’s weed

Blended trans esterified Calophyllum inophyllum oil with 1.5% of wet blue water Extract of Bishop’s weed

Blended trans esterified Calophyllum inophyllum oil with 2% of wet blue water Extract of Bishop’s weed

25 20 14 15

27 21 14 16

36 29 15 20

Aspergillus Niger Paecilomyces Variotii Aspergillus Flavus Penicillium notatum

particle size of the give proper lubrication of leather. Emulsion particle size values shown by Table 2 indicates that trans esterified Calophyllum inophyllum oil can also be used as fat- liquor in leather processing due to its comparable particle size with pores of the leather.

Table 7 Antifungal properties of trans esterified Calophyllum inophyllum oil, and its blending with 2% of wet blue of Bishop’s weed. against Aspergillus Niger. Paecilomyces variotii Penicillium notatum and Aspergillus flavus on wet blue. Zone of inhibition (mm) Aspergillus Niger

Zone of inhibition (mm) Paecilomyces variotii

Zone of inhibition (mm) Aspergillus flavus

Zone of inhibition (mm) Penicillium notatum

Trans esterified Calophyllum Inophyllum Oil

20

11

15

12

Blended trans Esterified Calophyllum Inophyllum oil with 2% of wet blue of water extract of Bishop’s weed

36

29

15

20

Fat liquor

3.3. Spectroscopic analysis of trans esterified Calophyllum inophyllum oil In Fig. 1 FTIR spectra of PEG- ester of Calophyllum inophyllum. Peak no 17 at 1735 cm−1, correspond to the stretching of C]O, typical of esters. The peak no 21 at 1538 cm−1 corresponds to the asymmetric stretching of eCH3 present in the trans esterified oil. The stretching of OeCH3, represented by the peak at 1196 cm−1 seen in peak no-25, is typical of trans esterified oil. Peak no 12 and 13 at 2857–2925 cm−1 corresponds to −CH2 symmetric and anti-symmetric stretching mode of vibration. In Fig. 2. 13C Spectra of PEG- ester of Calophyllum inophyllum. Peak in the region of 22–33 ppm is due to aliphatic carbon. Peak in the region of 61–72 ppm is due to glycerol carbon. Peak in the region of 127–129 ppm is due to unsaturated carbon. Peak in the region of 173.25 ppm is due to carbonyl carbon confirms the presence of ester and transesterification process of oil. In Fig. 3 1H Spectra of PEG- ester of Calophyllum inophyllum. Peak in the region of 0.6351 ppm is due to methyl protons. Peak in the region of 1.3 ppm is coresponds to methylene protons. Peak in the region of 1.7 ppm is represented by group CH3eCH2eCH2eCH2e. Peak in the region of 2.0 ppm is due to eCH2eCH2eCH]CH⋯. Peak at 2.5 ppm is due to ]CHeCH2eCH]. Peak at 3.5 ppm is due to eOeCH2eCH2e. Peak at 5.08 ppm coresponds to eCH]CHe group. Peak in the region of 7.3 ppm is due to solvent. All the values of Figs. 1–3 confirm the transesterification process of Calophyllum inophyllum oil.

3. Result and discussion 3.1. Chemical specification of Calophyllum inophyllum oil by gas chromatogram mass spectrometer and determination of Iodine and Saponification value Presence of high percentages of unsaturated acid (Oleic acid and Linoleic acid) shown in Table 1 and high iodine value indicates the better possibility for its modification and presence of more number of double bond. High value of saponification indicates short chain fatty acid present in the oil.

3.4. Chemical and spectroscopic specification of Bishop’s weed by gas chromatogram mass spectrometer and FTIR respectively

3.2. Emulsion particle size analysis Particle size of the emulsion plays important role in fat liquoring of leather. If their size exceeds the size of the pores of the leather their penetration deep into the leather would become difficult. Appropriate

Presence of high amount of four phenolic compounds thymol carvacrol para cymene and gamma terpinene was confirmed by

Table 8 Antifungal activity in terms of zone of inhibition of blended trans- esterified Calophyllum inophyllum oil with 2% of wet blue of water extracts from Bishop’s weed in comparison with synthetic fat liquor, semisynthetic fat liquor, trans- esterified vegetable oil against four following fungal species on wet blue. Fat liquor

Fungal Species Aspergillus Niger

Aspergillus flavus

Paecilomyces variotii

Penicillium notatum

Trans- esterified Calophyllum Inophyllum oil(blended) with 2% of wet blue of water extracts of Bishop’s weed

36

15

29

20

Trans-esterified Vegetable oil

10

11

11

13

Synthetic fat-liquor

9

11

8

10

Semisynthetic fat-liquor

8

6

9

4

110

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Fig. 9. The antifungal activity of blended trans- esterified Calophyllum inophyllum oil with 2% of wet blue of water extracts of Bishop’s weed against fungal species a) Aspergillus Niger b) Aspergillus flavus.

Figs. 7 and 8. Fig. 7 is SEM photograph of vegetable oil tanned leather sample, and Fig. 8 is the SEM photograph of processed leather sample fat liquoring with 10% solution of wet blue of trans esterified Calophyllum inophyllum oil. SEM photographs given in Figs. 7 and 8 show the voids and fibers in collagen matrix of two different leather samples. Fig. 7a (×300) and Fig. 7b (×100) shows the grain surface of the leather tanned with vegetable oil and Fig. 8a (×300) and Fig. 8b (×100) leather tanned with 10% solution of trans esterified Calophyllum inophyllum oil. Figs. 7c & 8c of Scanning electron microscope (SEM) shows the cross section of tanned leather with vegetable oil and with 10% weight of wet blue of Trans esterified Calophyllum inophyllum oil. The result shown by Figs. 7 and 8 confirms that the leather tanned with 10% weight of wet blue of trans esterified Calophyllum inophyllum oil which exhibited a soft grain without any fatty-spew. it is evident that leather treated with 10% emulsion of trans esterified Calophyllum inophyllum oil, shows the lubrication and deep penetration into the gaps of the leather fibers network which is very similar to vegetable oil treated leathers. The pore size distribution of wet-blue was carried out from skin of goat and sheep and reported that the average throat pore diameter (the most constricted diameter of through pore) has found to be around 1000 nm. Hence, the antifungal fatliquor can even fill up the constricted diameter of wet-blue leathers.

Table 9 Properties of fat liquored leather. Parameters of leather

Shrinkage temperature (−°C) Unbound fat content (%) Tensile strength (NM2) Tear strength (N) Lastometer (mm)

Experiments I

II

Trans- esterified Calophyllum inophyllum oil (blended) with 2% of water extract of Bishop’s Weed

Trans-Esterified Vegetable oil

111

120

11

13

36.29 172.82 10.64

42.72 163.06 11.40

GCMS analysis which is shown in Table 3 and further confirmed by FTIR which is shown in Fig. 4. In Fig. 5 FTIR spectra of water extract from Bishop’s weed, the absorption at 3359.39 cm−1 was due to the stretching of hydroxyl groups that were present in the extract. The band at 1729.83 cm−1 was assigned to C]O stretch. The band at 1729.83 cm−1 confirmed the presence of benzene ring in the sample. The bands at 1223.61 cm−1 were assigned to the strong intensity CeO skeletal vibrations. The molecular structure was similar to thymol as bands of functional group OeH, CeH, CeC, isopropyl and CeO have similar band wavelength to that of ajwain. In Fig. 6 1H Spectra of water extract from Bishop’s weed observed one proton multiplet at 5.34 ppm clearly assignable to olefinic protons. Another one proton multiplet observed at 3.6 ppm was assigned to carbonyl protons. The signals for all the methyl groups appeared a singlet between 1.9–0.8 ppm clearly indicated the attachment to saturated carbon atoms. Values given in Table 3 suggested that active component responsible for antifungal activity of Bishop’s weed is thymol, and carvacrol shown in Fig. 4. This was further confirmed from the spectroscopic analysis such as Fourier transform infrared spectroscopy (FTIR) and Nuclear magnetic resonance (NMR) of Bishop’s weed as shown in Figs. 5 and 6.

3.6. Antifungal activity of Calophyllum inophyllum oil and trans-esterified Calophyllum inophyllum oil It was observed from the data obtained as given in Table 4 that Calophyllum inophyllum oil shows higher zone of inhibition against four fungal species, which is satisfactory antifungal activity of oil against control (leather specimens without oil). It was also observed from the data presented in the Table 5 that trans-esterified Calophyllum inophyllum oil shows 20 mm zone of inhibition which is higher than control value of 0 mm and almost equal to synthetic fat-liquor and semi synthetic fat-liquor. But this antifungal activity of trans-esterified Calophyllum inophyllum oil was resistant only against Aspergillus Niger. 3.7. Blending of different ratios of water extract of Bishop’s weed with trans esterified Calophyllum inophyllum oil Trans esterified Calophyllum inophyllum oil was resistant only against Aspergillus Niger. Some common fungal species such as Paecilomyces variotii, Aspergillus flavus, and Penicillium notatum may also grow on leather after chrome tanning. To enhance the antifungal activity of fat

3.5. Scanning electron microscope (SEM) analysis of leather Scanning electron microscope (SEM) photographs are shown in 111

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Acknowledgments

liquor against some other fungal species, water extracts from Bishop's weed in different proportions was added. This was confirmed from the data given by Table 6 that out of three compositions, blending of 2% of water extracts of Bishop’s weed with trans esterified Calophyllum inophyllum oil was more effective than other two such 1% and 1.5% compositions

Financial support from CSIR-CLRI, is gratefully acknowledged. The authors thank Dr. Geetha Baskar and Dr. Suguna Lakshmi M. Principal scientist, Industrial chemistry lab, and Dr. A. Tamil Selvi Principal scientist, CHORD department CSIR-CLRI for their intellectual inputs and colleagues of EXECL lab CSIR-CLRI for their constant encouragement and support. The authors thank Director, CSIR-CLRI for his support and encouragement.

3.8. Comparison of Antifungal properties of trans esterified Calophyllum inophyllum oil and its blending with water extract of 2% of wet blue of Bishop’s weed

References It was found from the Table 7 that blended trans esterified Calophyllum inophyllum oil with water extract of 2% of wet blue of Bishop’s weed forms a stable and effective homogenous emulsion against four fungal species and shows better result than trans esterified Calophyllum inophyllum oil itself.

Akpomie, O., Ejila, A., Okocha, M.D., Opara, N.E., 2012. Preservation of wet-blues (Leather) using Cocos oil and organotin compounds. Int. Res. J. Microbiol. 3 (9), 296–300. Bajza, Zeljko, Vinkovic Vrcek, Ivana, 2001. Fat liquoring agent and drying temperature effects on leather properties. J. Mater. Sci. 36, 5265–5270. Dweck, A.C., Meadowsy, T., 2002. Tamanu (Calophyllum Inophyllum)—the African, Asian, Polynesian and Pacific Panacea. Int. J. Cosmet. Sci. 24, 1–8. Dwivedi, S.N., Mishra, R.P., Alava, Sangeeta, 2012. Phytochemistry, pharmacological studies and traditional benefits of Trachyspermumammi (Linn.). Int. J. Pharm. Life Sci. 3 (5), 1705–1709. Hassan Shahian, Mehdi, Bayat, Zeinab, Saeidi, Saeide, Shiri, Yasub, 2014. Antimicrobial activity of Trachyspermumammi essential oil against human bacterial. Int. J. Adv. Biol. Biomed. Res. 2 (1), 18–24. Kedzia, Anna, Mscisz, Alina, Meissner, Henry O., 2011. The effect of Tamanu oil (Calophyllum Inophyllum) on anaerobic bacteria isolated from respiratory tract. Borgis Post. Fitoter. 3, 159–163. Liu, Cheng-Kung, Latona, N.P., Dimaio, G., Cooke, P., 2007. Milling effects on mechanical behaviors of leather. JALCA 102, 191–197. Mathela, C.S., Singh, K.K., Gupta, V.K., 2010. Synthesis and In Vitro antibacterial activity of thymol and carvacrol derivatives. Acta Poloniae Pharm.—Drug Res. 67, 375–380. Meher, L.C., Vidya Sagar, D., Naik, S.N., 2006. Technical aspects of biodiesel production by transesterification—a review. Renew. Sustain. Energy Rev. 10 (3), 248–268. Morshed, S., Alam, M.K., Begum, A., Shahriar, S.M.S., Sharmin, K.N., Akther, S., 2012. Physicochemical properties and chemical constituents of oil from joan seed (Trachyspermumammi L.). J. Environ. Sci. Nat. Resour. 5 (2), 15–21. Orlita, A., 2004. Microbial biodetereorisation of leather and its control: a review. Int. Biodeterior. Biodegr. 53, 157–163. Rao, Mala B., Tanksale, Aparna M., Ghatge, Mohini S., Deshpande, Vasanti V., 1998. Molecular and biotechnological aspects of microbial proteases. Microbial. Mol. Biol. Rev. 62 (3), 597–635. Rehman, A., Rehman, Ali, Ahmad, I., 2015. Antibacterial, antifungal, and insecticidal potentials of Oxalis corniculata and its isolated compounds. Int. J. Anal. Chem. 2015, 1–5. Schuchardt, Ulf, Sercheli, Ricardo, Matheus Vargas, Rogério, 1998. Transesterification of vegetable oils: a review. J. Braz. Chem. Soc. 9 (1), 199–210. Shankar Mishra, U., Narasimha Murthy, P., Choudhury, P.K., Panigrahi, G., Ohapatra, S., Pradhan, D., 2010. Antibacterial and analgesic effects of the stem barks of Calophyllum inophyllum. Int. J. ChemTech Res. 2, 973–979. Sirvaityte, J., Siugzdaite, J., Valeika, V., 2012a. Commercial essential oils of Eucalyptus and Lavender as natural antibacterial agents in leather tanning industry. Rev. Chim. (Bucharest) 62 (9), 884–893. Sirvaityte, Justa, Siugzdaite, J., Valeikac, V., Dambrauskiened, Edita, 2012b. Application of essential oils of thyme as a natural preservative in leather tanning. Proc. Estonian Acad. Sci. 61 (3), 220–227. Sivakumar, V., Rao, P.G., 2001. Application of power ultrasound in leather processing: an ecofriendly approach. J. Clean Prod. 9 (1), 25–33. Zarlok, Jan, Smiechowski, Krzysztof, Mucha, Katarzyna, Tecza, Agnieszka, 2014. Research on application of flax and soya oil for leather fat liquoring. J. Clean Prod. 65, 583–589.

3.9. Antifungal activity in terms of zone of inhibition of blended transesterified Calophyllum inophyllum oil with 2% of wet blue of water extracts of Bishop’s weed in comparison with synthetic fat liquor, semisynthetic fat liquor, trans-esterified vegetable oil against four following fungal species on wet blue The results obtained from Table 8 and Fig. 9 indicates that the antifungal activity in terms of zone of inhibition of blended transesterified Calophyllum inophyllum oil with 2% of wet blue of water extracts of Bishop’s weed is more than that of synthetic fat liquor, semisynthetic fat liquor and trans- esterified vegetable oil against four following funguses on wet blue. 3.10. Fat liquored leather properties After fat liquoring, the leather was subjected to some tests like shrinkage temperature, tensile strength and tear strength. The obtained results shown by Table 9 indicates that the properties of the leather fat liquored with trans- esterified Calophyllum inophyllum with 2% of wet blue of water extracts from Bishop’s weed are similar to properties of the leather fat liquored with other vegetable oil and have considerable improved property specially with respect to tear strength (N). 4. Conclusion Fat liquor based on trans esterified Calophyllum inophyllum oil shows antifungal activity and possesses satisfactory stability. Blending of water extract of Bishop’s weed with trans Esterified Calophyllum inophyllum oil enhances the antifungal activity of fat liquor against fungal species such as Aspergillus Niger, Aspergillus flavus, Penicillium notatum, and Paecilomyces variotii. Therefore, use of this new blended fat liquor can be a progressive step towards the cleaner production.

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