(Tejpat) Essential
Natural Product Radiance, Vol. 8(2), 2009, pp.106-116 Research Paper
Chemistry, antimicrobial and antioxidant potentials of Cinnamomum tamala Nees & Eberm. (Tejpat) essential oil and oleoresins† I P S Kapoor1, Bandana Singh1, Gurdip Singh1*, Valery Isidorov2 and Lech Szczepaniak2 1Chemistry Department, DDU Gorakhpur University, Gorakhpur- 273 009, Uttar Pradesh, India 2Institute of Chemistry, Bialystok University, UI. Hurtowa 1, 15-399, Bialystok, Poland *Correspondent author, E-mail: [email protected]; Phone: +91-551-2200745 (R), 2202856 (O); Fax: +91-551-2340459 Received 6 November 2007; Accepted 26 November 2008 †Part 63 Abstract Gas chromatography-mass spectrometery (GC-MS) analysis of essential oil and oleoresins of Cinnamomum tamala Nees & Eberm. (Tejpat) revealed eugenol as major component of essential oil and oleoresins. The antioxidant activity of essential oil and oleoresins were evaluated against mustard oil by peroxide, p-anisidine, thiobarbituric acid and total carbonyl value method. In addition, their inhibitory action by FTC method, scavenging capacity by DPPH (2, 2′-diphenyl- 1-picrylhydrazyl radical) method was also studied. The antimicrobial potentials of essential oil and oleoresins were tested against various food born fungi and bacteria. Results showed that both the volatile oil and oleoresins have effective antioxidant and antimicrobial activities. Thus, they could be evaluated as natural food preservatives, however, essential oil is better than oleoresins. Keywords: Antimicrobial activity, Antioxidant activity, Cinnamomum tamala, Essential oil, Indian Cassia lignea, Oleoresins, Tejpat. IPC code; Int. cl.8 — A61K 36/54, A61P 31/00, A61P 39/06
Introduction of spices have been reported by few Antioxidants are being workers4-6. commonly used to prevent the fat rancidity Indian Cassia lignea, because these are substances that when Cinnamomum tamala Nees & Tejpat leaves added to food products, especially lipids Eberm. (Hindi—Tejpat)7 is an and lipid containing food can increase evergreen tropical tree, belonging to the with the chemistry, antioxidative and shelf-life by retarding lipid peroxidation. Lauraceae family. It is mainly used for antimicrobial behaviour of essential oil Various synthetic antioxidants such as flavouring food and widely used in and oleoresins (extracted in methanol, ascorbyl palmitate, butylated pharmaceutical preparation because of its ethanol, carbon tetrachloride and hydroxytoluene (BHT), butylated hypoglycemic, stimulant and carminative isooctane) of C. tamala. The objective hydroxyanisole (BHA) and propyl gallate properties8, 9. The leaves of this tree used of this paper is the comparative study of (PG) have been approved to control and as spice having clove like taste and pepper chemistry, antioxidative and antimicrobial delay the onset of rancidity and routinely like odour. Essential oil of Cinnamomum properties of tejpat essential oil and used as food protecting agent1. However, leaves has excellent inhibitory effect on oleoresins. recent concern over their adverse effects bacteria10. There are numerous studies on and toxicity2-3 has created a need and the composition of tejpat essential oil11; Materials and Methods prompt research for natural antioxidants however, tejpat oleoresins are not studied Thiobarbituric acid (TBA), (spices). Antioxidant and antimicrobial so vastly. As a part of our ongoing research diphenylpicrylhydrazyl (DPPH), linoleic properties of volatile oil and oleoresins programme12, 13 the present paper deals acid (Across, USA). BHT, BHA, PG and
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2,4-dinitrophenylhydrazine were extractor (The reason for using different acquisition parameters: transfer line held purchased from s. d. fine-chem Ltd, solvents is that each oleoresin extracted at 280oC and detector was held at 300oC. Mumbai, India. Ampicillin of Ranbaxy in a particular solvent shows different Detection was performed in the full scan Fine Chemicals Ltd., New Delhi, India was component with varying percentage). The mode from m/z 41 to 450. used. Crude mustard oil was purchased solvent was evaporated under reduced A hexane solution of C8-C32 from local oil mill, Gorakhpur, and all pressure. The different viscous oleoresins n-alkanes was previously separated under solvents used were of analytical grade. of C. tamala leaves were stored under the above conditions, and their retention Tejpat leaves were purchased cold condition until further use. times were determined. Linear from local market of Gorakhpur, Uttar temperature programmed retention Pradesh, during July 2006 and voucher Chemical investigation indices (LTPRI) were calculated from the specimens were kept at the Herbarium of The chemical analysis of volatile results of the separation of the essential the Science faculty, DDU Gorakhpur oil and oleoresins were undertaken by gas oil and oleoresins (eq. 1):
University, Gorakhpur. chromatography-mass spectrometry LTPRI = 100(tx - tn)/(tn+1 - tn) + 100n In order to determine the (GC-MS) technique: ……..(1) antimicrobial activity of the volatile oil where tx, tn and tn+1 are the retention times and oleoresins, various food-born GC-MS of component x, and n-alkanes with the pathogenic fungi and bacteria were taken. The analysis of the volatile oil and number of carbon atoms in the molecule ≤ ≤ Aspergillus niger, A. flavus, A. oleoresin extracts were performed using n and n+1, respectively (tn tx tn+1). oryzae, A. awamori, A. solani and gas chromatograph HP 6890 with mass After integration the fraction of each Fusarium monoliforme were taken as selective detector MS 5973 (Agilent component in the total ion current (TIC) fungi and the test bacteria were: Technologies, USA) fitted with a HP-5MS was calculated. Escherichia coli, Staphylococcus fused silica column (30 m × 0.25 mm; aureus, Pseudomonas aeruginosa, 0.25 µm film thickness), with electronic Identification of components Proteus vulgaris Klebsiella pressure control (EPC) and splitless Components of essential oil pneumoniae and Bacillus cereus. injector. Helium flow rate through the (Table 1) and oleoresins (Table 2) were These fungi and bacteria were purchased column was 1 ml/min in constant flow identified with the aid of an automatic from Microbial Type Culture Collection mode. The initial column temperature system of processing data of GC-MS MTCC, Chandigarh, India. was 40oC rising 250oC at a rate 3o/min and supplied by NIST mass spectra library. The the higher temp. 250oC was maintained MS library was performed by using PBM Extraction of oil and oleoresins for 15 minutes. The MS detector (Probability-Based Matching) algorithm. The powdered leaves of C. Identification tamala were subjected to hydrodistillation was considered in a Clevenger apparatus for 6 h reliable if the in accordance with European measured Pharmacopoeia14 procedure to get yellow values of volatile oil (yield 2.2%) of characteristic retention odour and sharp taste. Further it was dried indices over anhydrous sodium sulphate and confirmed the stored in a refrigerator. results of Oleoresins were obtained by computer extracting 25 g of powdered leaves with search at mass 250 ml of each solvent (MeOH, EtOH, Essential oil and oleoresins of spectra library. Cinnamomum tamala leaves iso-octane and CCl4) for 3 h in a Soxhlet
Vol 8(2) March-April 2009 107 Research Paper
Table 1: Relative composition of essential oil of Tejpat by GC-MS
Compound Rel. comp. (%) aRIExp Compound Rel. comp. (%) aRIExp
α-Thujene 0.02 923 (E)-β-Caryophyllene 1.9 1416 α-Pinene 0.07 932 Sesquiterpene 0.2 1430 β-Pinene 0.03 974 Aromadendrene 1.5 1436 6-Methyl-5-heptene-2-one trace 987 α-Guaiene 0.1 1439 Myrcene 0.02 990 α-Humulene 0.4 1449 α-Phellandrene 0.4 1001 Alloaromadendrene 0.5 1456 3-Carene 0.02 1006 Ethyl vanillin trace 1459 p-Cymene 0.6 1021 γ-Muurolene 0.6 1473 Limonene trace 1028 D-Germacrene 0.5 1476 1,8-Cineole 0.4 1030 β-Eudesmene (β-selinene) trace 1481 p-Mentha-2,4(8)-diene trace 1082 Mixture of sesquiterpenes 0.4 1483 Terpinolene 0.03 1084 Viridiflorene 2.9 1492 p-Cymenene trace 1086 γ-Cadinene 0.2 1509 Linalool 0.04 1100 Sesquiterpenoid C15H26O 0.2 1511 trans-Pinocarveol trace 1134 δ-Cadinene 1.1 1520 Camphor trace 1139 Eugenolacetate 0.1 1529 cis-Verbenol trace 1141 α-Cadinene 0.07 1533 Borneol 0.1 1161 α-Calacorene 0.1 1538 Terpinene-4-ol 0.1 1173 Sesquiterpenoid C H O 0.09 1546 m-Cymen-8-ol trace 1180 15 26 Sesquiterpenoid C15H24O 0.2 1550 p-Cymen-8-ol 0.08 1183 Sesquiterpenoid C H O 0.2 1554 α 15 26 -Terpineol 0.5 1187 Elemicin trace 1558 trans-Piperitol trace 1205 Sesquiterpenoid C15H26O 0.3 1561 iso-Dihydrocarveol 0.03 1211 trans-Nerolidol + 0.2 1564 (Z)-Cinnamaldehyde trace 1216 sesquiterpenoid C15H24O p-Cumic aldehyde 0.03 1236 Spathulenol 4.8 1576 Carvone trace 1241 Caryophyllene oxide trace 1578 Chavicol trace 1257 Globulol + 1.6 1580 (E)-Cinnamaldehyde 0.5 1278 sesquiterpenoid C15H24O Anethol 0.2 1282 Viridiflorol 0.6 1586 p-Cymen-7-ol trace 1289 Guaiol 0.5 1596 Thymol 0.2 1295 4-Allyl-2,6-dimethoxy phenol 0.2 1602 Carvacrol trace 1297 10-epi-γ-Eudesmol 0.3 1617 3-Methoxyacetophenone 0.1 1303 Sesquiterpenoid C H O 1.5 1634 α-Elemene 0.06 1333 15 24 δ-Cadinol (torreyol) 0.2 1641 α-Cubebene 0.06 1345 α-Cadinol 0.6 1650 β-Terpineol acetate trace 1347 Sesquiterpenoid C H O 0.2 1656 Eugenol 66.1 1381 15 24 Coniferol 0.2 1734 β-Elemene 0.3 1392 Hexahydrofarnesyl acetone 0.04 1842 Vanillaldehyde 0.2 1401 Farnesyl acetone 0.03 1914 α-Gurjunene 0.1 1406 Methyleugenol 1.3 1409 Total 94.12%
*Trace: below 0.01% of tic; Percentages are the mean of three runs and were obtained from electronic integrations measurements using flame ionization detection (fid).
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Table 2 : Relative composition of oleoresins of Tejpat in different solvents
a Exp Compounds Methanol Ethanol Isooctane CCl4 RI
α-Phellandrene - - 0.03 - 1001 p-Cymene - - 0.05 - 1021 Limonene - - trace - 1025 1,8-Cineole 0.06 trace 0.04 --- 1030 3,5-Dihydroxy-6-methyl-2,3-dihydro-4H-pyran-4-one 0.09 trace ------1141 α-Terpineol 0.2 0.2 0.5 0.03 1187 Monoterpenoid 0.08 trace ------1198 Pyrocatechol 0.2 trace ------1214 2,3-Dihydrobenzofuran (coumaran) --- 0.01 ------1226 Anethol ------0.04 --- 1282 Thymol 0.3 trace ------1296 p-Vinylguiacol 0.2 0.2 ------1313 Eugenol 69.3 63.9 28.4 17.3 1359 α-Copaene ------0.2 0.05 1370 Vanillaldehyde 0.2 trace --- 0.03 1398 Methyleugenol 0.4 0.5 0.1 0.04 1404 (E)-β-Caryophyllene 0.06 0.3 0.5 0.2 1416 Aromadendrene --- 2.2 0.4 0.1 1439 Isoeugenol 0.2 trace - --- 1446 Alloaromadendrene ------0.1 --- 1456 γ-Muurolene ------0.1 0.02 1471 D-Germacrene ------0.2 0.06 1476
Sesquiterpene C15H24 0.5 0.5 1.8 0.02 1488 γ-Cadinene ------0.1 trace 1508 δ-Cadinene 0.2 0.2 0.3 0.07 1518 Spathulenol 1.5 1.9 1.9 1.4 1572 Globulol 0.3 0.3 0.3 0.2 1584 Viridiflorol 0.2 0.2 0.1 0.06 1595 n-Hexadecane ------0.4 --- 1600 4-Allyl-2,6-dimethoxy phenol ------0.4 0.3 1602 Methoxy eugenol 1.1 1.0 ------1604 10-epi-γ-Eudesmol ------0.05 0.05 1617
Sesquiterpenoid C15H24O 0.5 --- - 0.2 1632 Homovanillic acid 0.3 0.3 - --- 1644 α-Cadinol ------0.3 --- 1650 Syringaldehyde 0.3 0.3 - --- 1657 Coniferol 0.7 0.8 - 1733 Neophytadiene (3,7,11,15-tetramethyl-2-hexadecene) --- 0.5 0.4 0.6 1836
Vol 8(2) March-April 2009 109 Research Paper
a Exp Compounds Methanol Ethanol Isooctane CCl4 RI
Methylpalmitate 0.3 ------0.1 1924 Dibutyl phthalate ------0.2 1957 Palmitic acid 0.6 2.8 --- 1.9 1962 4-Methoxystilbene 0.1 0.1 ------1987 Ethyl hexadecanoate --- 6.5 --- 0.1 1992 Methyl linoleate 1.3 trace 0.3 0.4 2090 Methyl oleate 0.2 ------0.1 2096 Phytol ------0.1 2109 Ethyl vanillin trace trace --- trace 2120 Linoleic acid 1.0 6.5 12.0 11.9 2137 Oleic acid --- 2.7 1.5 1.1 2141 Ethyl linoleate --- 2.4 0.8 0.9 2158 Ethyl oleate --- 0.8 0.5 0.3 2164 n-Tricosane ------0.3 0.1 2300 n-Tetracosane ------0.2 0.2 2400 n-Pentacosane ------0.3 0.3 2500 bis(2-Ethylhexyl)-phthalate --- 3.2 27.3 27.9 2549 Capsaicine, (E)-8-methyl-N-vanillyl-6-nonenamide - 0.3 0.04 0.04 2555 n-Heptacosane --- 0.3 0.6 0.5 2700 n-Nonacosane --- 0.6 1.0 0.9 2900 n-Triacontane ------0.5 0.5 3000 n-Hentriacontane ------0.6 0.6 3100 α-Tocopherol trace 0.6 1.1 1.1 3128 β-Sitosterol ------2.6 3289 Stigmastenone ------2.8 >3300
Total 80.4 93.8 86.6 78.1
*Trace: below 0.01% of TIC; Percentages are the mean of three runs and were obtained from electronic integrations measurements using flame ionization detection (fid);
a: the retention index was calculated for all volatile constituents using a homologous series of n-alkanes C8-C32.
Antioxidant activity having initial peroxide value 43.2 meq/ with crude mustard oil at 200ppm Antioxidant potential of kg was taken for present investigation. concentration. They were subjected to Cinnamomum oil and oleoresins for Schaal oven test15 in 100 ml open beakers mustard oil have been evaluated by Sample preparation at 90°C. A control sample was prepared different methods such as peroxide, The volatile oil and oleoresin of under similar condition without any p-anisidine, thiobarbituric acid and total tejpat along with synthetic antioxidants additive. carbonyl value method. Crude mustard oil such as BHA, BHT and PG were mixed The antioxidant activity of oil and
110 Natural Product Radiance Research Paper oleoresins was examined by comparing the oleoresins, the pathogenic fungus technique, the required dose (2, 4 and activity of known antioxidants such as Aspergillus niger, A. flavus, A. 6 µl) of the diluted sample (in methanol) BHA, BHT and PG by peroxide16, TBA3, 17, oryzae, A. awamori, A. solani and were mixed with the 20 ml of culture p-anisidine18 and total carbonyl values19 Fusarium monoliforme were medium. Each test was replicated for according to the method reported earlier. undertaken. Cultures of each of the fungi three times and fungi toxicity was were maintained on Czapek (DOX) agar measured after every 6 days in terms of Complementary antioxidant assays media with adjusting pH 6.0-6.5 and per cent mycelial zone inhibition using Further determination of slants were stored at 4°C. The antifungal the following formula: antioxidant activity of essential oil and activity of the volatile oil and oleoresins tejpat oleoresins of in linoleic acid against fungi were undertaken using Per cent mycelial= dc-dt system and scavenging effect on DPPH inverted petriplate23 and poison food zone inhibition dc were determined by the methods reported techniques24. In inverted petriplate earlier20-22. method, the required dose (2, 4 and where dc and dt are average diameters of 6 µl) of undiluted sample were soaked mycelial colony of control and treated Antimicrobial activity on a small piece (diam. 12 mm) of sets, respectively. The results recorded Antifungal investigations Whatmann No. 1 filter paper and was kept by inverted petriplate and food In order to determine the on the lid of petriplate which is in inverted poison technique are shown in antifungal efficacy of the volatile oil and position at 37°C whereas in poison food Tables 3 & 4.
Table 3 : Antifungal activity of volatile oils and oleoresins of Cinnamomum tamala leaves by food poisoned method
Test samples Dose (µl) Food poisoned method (per cent mycelial inhibition zone#)
Aspergillus Aspergillus Aspergillus Fusarium Aspergillus Aspergillus niger flavus awamori moniliferum oryzae solani
Cinnamomum oil 2 14.2±0.1 62.2±1.3 24.3±0.0 38.0±2.2 56.8±0.6 58.5±1.2 4 57.1±0.0 72.9±0.0 40.5±0.5 69.0±0.0 67.5±0.0 68.4±2.2 6 100 83.8±0.0 56.7±1.2 100 78.4±0.6 80.2±0.0 MeOH oleoresin 2 5.8±0.5 32.7±0.5 12.6±0.0 5.1±0.6 16.1±2.2 7.1±0.5 4 9.3±1.2 37.2±1.0 26.6±1.3 7.4±0.0 21.2±0.0 19.3±0.0 6 12.8±0.2 41.6±1.6 40.5±0.6 9.6±0.0 26.1±0.6 31.5±0.8 EtOH oleoresin 2 6.2±1.2 29.8±2.3 15.3±0.8 16.0±0.0 22.4±0.0 2.8±0.8 4 8.5±0.0 39.9±0.0 29.5±0.0 18.6±0.1 28.6±0.0 9.0 ±0.0 6 10.9±0.4 50.0±1.2 43.6±0.0 21.2±0.2 34.8±0.0 15.3±0.0 Iso-octane oleoresin 2 8.7±0.5 5.0±0.0 12.1±0.5 9.6±0.5 12.8±1.2 30.2±0.8 4 15.4±0.6 21.4±1.2 27.5±0.7 14.8±0.0 16.8±0.0 43.9±1.2 6 22.1±0.0 37.8±0.0 42.9±1.8 18.7±0.8 20.7±0.6 57.6±2.3
CCl4 oleoresin 2 7.5±0.0 12.6±0.0 5.0±2.3 14.8±0.9 8.3±0.6 10.7±1.2 4 9.2±0.0 17.6±0.3 13.4±0.0 16.3±1.2 18.7±0.7 21.9±0.6 6 10.9±1.2 22.6±0.2 21.8±0.0 17.7±0.2 29.0±0.8 32.6±0.9
# Average of three replicates; Values are means ± S.D. of three independent determinations.
Vol 8(2) March-April 2009 111 Research Paper
Table 4: Antifungal activity of volatile oils and oleoresins of Cinnamomum tamala leaves by inverted petriplate method
Test samples Dose (µl)* Inverted petriplate method (per cent mycelial inhibition zone#)
Aspergillus Aspergillus Aspergillus Fusarium Aspergillus Aspergillus niger flavus awamori moniliferum oryzae solani
Tejpat oil 2 38.0±0.4 85.9±1.3 30.2±0.2 49.7±0.0 100 45.5±1.1 4 69.6±0.6 92.9±0.0 43.9±0.5 74.8±1.2 100 72.8±0.0 6 100 100 87.6±0.0 100 100 100 MeOH oleoresin 2 5.62±1.2 3.0±0.2 18.2±0.2 22.7±0.0 27.4±0.5 27.8±0.5 4 15.6±0.0 12.2±0.6 22.6±0.0 25.9±0.0 31.8±0.5 41.0±0.1 6 56.2±0.5 21.3±0.3 36.5±0.4 29.1±0.2 36.3±0.0 54.2±0.2 EtOH oleoresin 2 32.1±0.8 20.9±0.6 5.0±0.1 4..5±0.5 11.3±0.5 59.4±0.0 4 36.5±0.5 30.9±0.8 15.5±0.6 18.3±1.4 18.4±1.5 60.6±0.0 6 59.2±0.2 41.0±1.2 26.7±0.0 32.1±0.6 25.4±0.1 65.9±0.1 Iso-octane oleoresin 2 5.02±0.2 10.1±0.0 6.72±0.5 25.7±0.6 22.9±0.2 31.2±0.2 4 18.6±0.0 20.3±0.6 9.82±1.3 30.5±0.5 29.6±1.5 34.7±0.5 6 27.8±0.2 30.5±0.9 19.8±1.5 35.2±0.0 36.4±0.3 38.2±0.9
CCl4 oleoresin 2 9.06±1.2 13.6±0.0 10.2±0.5 4.84±1.2 19.6±0.9 17.9±1.2 4 19.5±0.0 17.8±1.2 16.5±0.5 16.9±0.0 31.6±0.1 19.9±0.0 6 32.6±0.0 21.9±0.0 27.6±0.0 29.1±0.5 43.5±0.0 21.8±0.6 # Average of three replicates; *Samples were diluted with methanol; Values are means ± S.D. of three independent determinations.
Antibacterial investigations solution were inoculated on to the surface 81 components representing 94.1% of the Six pathogenic bacteria of well settled sterilized culture medium. total amount (Table 1), and eugenol Escherichia coli, Staphylococcus The wells (10 mm diameter) were cut was major component (66.1%) followed aureus, Pseudomonas aeruginosa, from agar, and 0.2 ml of sample (2, 4 by spathulenol (4.8%), viridiflorene Proteus vulgaris, Klebsiella and 6 µl of essential oil or oleoresins (2.4%), methyleugenol (1.9%), pneumoniae and Bacillus cereus have diluted in 1 ml of DMSO) was delivered aromadendrene (1.5%),with other been used. They were sub-cultured on into them. For standard, 0.2 ml of aqueous constituents (in minor amounts). From nutrient agar broth (Hi-media) and stored solution of ampicillin (1 mg/ml) was literature survey it is clear that eugenol is at 4oC. Active cultures for experiments used. After incubation for 24 h at 37oC, the major component in the essential oil were prepared by transferring one loopful all plates were examined for any zones of of C. tamala leaves26-28. of cells from stock cultures to flask of growth inhibition and the diameters of The oleoresins (MeOH, EtOH, nutrient agar broth, which were incubated these zones were measured in millimeters iso-octane and CCl4) showed the presence without agitation for 24 h at 37oC. and averages of three replicates are of 28 components (80.4%), 34 In order to determine the reported in Table 5. components (93.8%), 42 components antibacterial activity of the essential oil (86.6%) and 41 components (78.1%) of and oleoresins, agar well diffusion Results and Discussion the total amounts, respectively. The major method25 was used. Bacterial strain, GC-MS analysis of Cinnamomum components of the oleoresins as observed 0.1 ml of 1010 times diluted in ringers' leaf volatile oil showed the presence of from GC-MS analysis are:
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Table 5 : Antibacterial activity of volatile oils and oleoresins of Cinnamomum tamala by agar well diffusion method
Test samples Conc. Inhibition zone (mm)# (ppm) * Klebsiella Staphylococcus Bacillus Escherichia Proteus Pseudomonas pneumoniae aureus cereus coli vulgaris aeruginosa
Tejpat oil 1000 72 ± 0.6 72 ± 0.5 56 ±1.5 31 ± 0.2 63 ± 0.5 76 ± 0.6 2000 88 ± 1.5 87 ± 0.5 75 ±1.1 66 ± 0.5 81 ± 0.3 88 ± 1.5 3000 (+) 95 ± 0.5 79 ± 1.1 (+) 86 ± 0.3 (+) MeOH oleoresin 1000 29 ± 1.5 48 ± 0.5 64 ± 0.5 17 ± 0.3 40 ± 0.5 22 ± 1.5 2000 32 ± 1.1 64 ± 1.1 81 ± 0.3 22 ± 0.5 53 ± 1.2 32 ± 1.1 3000 32 ± 1.1 67 ± 1.1 83 ± 0.3 (+) 58 ± 1.2 38 ± 1.1 EtOH oleoresin 1000 (-) 42 ±0.5 53 ± 1.1 (-) 13 ± 1.5 41 ± 0.5 2000 56 ±1.2 64 ±1.1 84 ± 1.2 59 ± 0.6 24 ± 0.3 56 ± 1.2 3000 58 ± 1.2 64 ±1.1 84 ± 1.2 82 ± 0.6 24 ± 0.3 57 ± 1.2 Iso-octane oleoresin 1000 (-) 44 ±0.5 64 ± 0.5 42 ±1.3 (-) 26 ± 0.3 2000 42 ± 1.1 64 ±1.1 81 ± 0.3 66 ± 0.5 34 ± 0.6 40 ± 1.1 3000 44 ± 1.1 74 ± 1.1 83 ± 0.3 69 ± 0.5 39 ± 0.6 44 ± 1.1
CCl4 oleoresin 1000 18 ± 0.5 (-) 36 ± 0.5 (+) (-) (-) 2000 22 ± 0.5 24 ± 0.3 48 ±0.2 13 ± 0.2 32 ± 0.6 23 ± 0.5 3000 22 ± 0.5 24 ± 0.3 48 ± 0.2 13 ± 0.2 32 ± 0.6 29 ± 0.5 Ampicillin 1000 30 ± 0.2 18 ± 0.7 25 ± 0.2 17 ± 0.6 21 ± 0.7 22 ± 0.4 2000 34 ± 0.3 32 ± 1.6 34 ± 0.1 37 ± 1.4 25 ± 0.7 26 ± 1.5 3000 41 ± 0.2 37 ± 0.2 38 ± 0.3 39 ± 0.6 28 ± 1.3 27 ± 1.1
# Average of three replicates; (+) indicates complete inhibition ; (-) indicates no inhibition; *samples were diluted in DMSO.
Methanol oleoresin : eugenol (69.3%), spathulenol(1.5%), methyl Terpenoids too play an important role in linoleate(1.3%), linoleic acid(1.0%), respectively. antioxidant activity30 (Tables 1 & 2). It Ethanol oleoresin : eugenol (63.9%), bis(2-Ethylhexyl)phthalate(3.2%), has also been reported that most natural ethyl hexade-canoate(6.9%), linoleic acid(6.5%) oleic antioxidative compounds often work acid(2.7%), respectively. synergistically with each other to produce Iso-octane oleoresin : eugenol (28.4%), bis (2-Ethylhexyl) phthalate (27.3%), a broad spectrum of antioxidative activities linoleic acid (12.0%), oleic acid (1.5%), respectively. that creates an effective defence system 31, 32 CCl4 oleoresin : eugenol (17.3%), bis (2-Ethylhexyl) phthalate (27.9%), against free radical attack . The linoleic acid (12.0%), oleic acid (1.1%), respectively. malondehyde formation of all the additives increases with storage time Regarding antioxidant activity, all oil and ethanol oleoresin showed better (Fig. 2). The oil showed a moderate samples with essential oil and oleoresins antioxidant effect even at the final stage inhibition at 0.02% concentration, and were able to control the oxidation rate of of the experiment. This could be due to was comparable to BHA and PG. The the mustard oil on heating at 60°C than the presence of phenolic compounds29 samples with volatile oil and all oleoresins the control, when assessed by the change such as eugenol, spathulenol in essential were found to be significantly (p < 0.05) in peroxide values (Fig. 1). The essential oil, methanol and ethanol oleoresins. more effective than the control. These
Vol 8(2) March-April 2009 113 Research Paper results were well correlated with p-anisidine and total carbonyl values, Peroxide value 180 linoleic acid system and scavenging effect 160 (Figs 3-6). 140 control The volatile oil was found to be 120 BHA 100 BHT PG 100% active against Fusarium 80 MeOH moniliforme, Aspergillus niger, A. 60 EtOH Peroxide value 40 Iso-octane oryzae and A. solani but not for A. (meq of oxygen/kg) 20 CC14 awamori in inverted petriplate method, 0 Cinnamomum oil though food poisoned method revealed 71421 28 100% activity for A. niger and Fusarium Incubation time (days) moniliforme at 6 µl dose. It was found to be highly effective in controlling the Fig. 1 : Inhibitory effect of C. tamala volatile oil and its various oleoresins on the primary oxidation of mustard oil using peroxide value method growth of A. flavus, A. solani and A. oryzae as more than 65% mycelial zone inhibition at 4 µl was observed. For other Thio barbituric acid value tested fungi, it was found to be less 7 effective. Since oleoresins have less or no control 6 BHA vapour action (compare to essential oil), 5 BHT therefore they were found to be less 4 PG 3 MeOH effective which is clear from the 40% EtOH
TBA (meq/kg) 2 mycelial zone inhibition. Moreover, the CC14 1 Iso-octane data for both essential oil and oleoresins 0 Cinnamomum oil were found to be statistically significant 71421 28
(p< 0.05). It is inferred that as the oil Incubation days concentration increased, inhibitory effects also increased. The oils and oleoresins Fig. 2 : Inhibitory effect of C. tamala volatile oil and its various oleoresins on though less effective at lower the secondary oxidation of mustard oil using TBA value method concentration, gave rise to complete inhibition of fungal growth when their concentration is increased. p-Anisidine value Antibacterial investigation 80 control (Table 4) of tejpat essential oil exhibited 70 BHA 60 strong inhibition against all the tested BHT microorganisms at various concentrations 50 PG 40 MeOH in the present study. Methanol, ethanol, 30 EtOH
-Anisidine value 20 Iso-octane iso-octane and CCl oleoresins have shown p 4 CC1 10 4 strong inhibition against E. coli at 3000 Cinnamomum oil 0 ppm. Essential oil and oleoresins have 71421 28 shown good to moderate activity against Incubation days other tested bacterial strains. Concluding these results, it can be said that the volatile Fig. 3 : Inhibitory effect of C. tamala volatile oil and its various oleoresins on oil and oleoresins of tejpat leaf possess the secondary oxidation of mustard oil using p- Anisidine value method
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excellent antibacterial properties even at Total carboyl value very low concentration against tested 18 bacterial strains. 16 14 Most of the antimicrobial activity 12 control in volatile oils derived from spices appears BHA 10 BHT to derive from phenolic compounds such 8 PG as eugenol, spathulenol, etc. while other MeOH TCV (meq/g) 6 EtOH constituents are believed to contribute 4 Iso-octane little33, though it is well known fact that 2 CCl4 Cinnamomum oil 0 the activity of an essential oil or oleoresins 71421 28 could be due to their major components. Incubation days
Fig. 4 : Inhibitory effect of C. tamala volatile oil and its various oleoresins on Conclusion the secondary oxidation of mustard oil using total carbonyl value method To conclude tejpat essential oil and oleoresins possess considerable Linoleic acid value antioxidant capacity. They have also shown
1.2 broad spectrum of antimicrobial activity
1 control against tested fungi and bacteria. Hence, BHA 0.8 BHT they could be used as natural food PG 0.6 MeOH additives. EtOH 0.4 Iso-octane CCl 0.2 4 Acknowledgement absorbance at 517 nm Cinnamomum oil 0 Authors are thankful to Head 212468 10 (Prof. K.D.S Yadav), Chemistry
Incubation days Department, DDU Gorakhpur University, Gorakhpur for providing laboratory and Fig. 5 : Inhibitory effect of C. tamala volatile oil and its various oleoresins on spectral facility. Thanks are due to CST, the oxidation of linoleic acid system using ferric thiocyanate method U.P. for providing financial assistance to one of the author (Bandana Singh).
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