Experimental studies on Influence of different doses of Vermicompost on essential oil yield of Ocimum tenuiflorum var. CIM-Ayu

 

A. Ranganadha Reddy¹*, R. Bharath Kumar¹, T. C. Venkateswarulu¹, M. Indira¹, D. Ramyakrishna¹, M. Deepthi¹, K. P. Sastry²

¹School of Biotechnology, Vignan University, Vadlamudi, Guntur 522 213, Andhra Pradesh, India

²CIMAP Uppal, Hyderabad 500 039, Andhra Pradesh, India.

 

ABSTRACT:

Vermicompost is a naturally occurring organic biofertilizer and it contains 1.5 – 2.5 % Nitrogen, 0.9 – 1.7 % Phosphorus, 1.5 – 2.4 % Potash, 0.5 – 1.0 % Calcium, 0.2 – 0.3 % Magnesium, 0.4 - 0.5 % Sulphur. Ocimum tenuiflorum yields aromatic oil containing eugenol (71.3%). The present study focused on the influence of different doses of vermicompost on the essential oil yield of Ocimum tenuiflorum var. CIM-Ayu. In the experiment, six different doses of vermicompost (0- 3.0 tons/ha) was applied and during the course of investigation the essential oil from the herbs was collected at two different stages (i.e. at 45, 90 days after planting) and it was noticed that the essential content did not follow any particular pattern. Similarly, the chemical constituents of the oil were not influenced by the treatments. It was also observed that reduction in plant height and number of tillers/clump that was manifested in reduced herb yield was compensated by a higher oil concentration in the leaves. The highest content of Eugenol, was reported in addition to  Methyl Eugenol, Beta Elemene and Caryophyllene.

 

 

INTRODUCTION:

Ocimum tenuiflorum (Krishna Tulasi) belonging to the family Lamiaceae (labiate) [1] is a medicinal [2] and aromatic crop that is thought to have originated in north central India and now grows native throughout the eastern world tropics, which yields essential oil and aroma chemicals and find diverse uses in the perfumery and cosmetic industries as well as in indigenous systems of medicine.[3][4] Out of 160 species of Ocimum, recorded in India, Ocimum tenuiflorum Linn is cultivated in India on a commercial scale. It is grown in various parts of the country viz. in west Bengal, Maharashtra, Uttar Pradesh, Madhya Pradesh, Bihar, Jammu, Assam etc.[5]

 

Essential oil of Ocimum tenuiflorum contained higher amount of linalool (39.39-55.26%) and moderate amounts of methyl chavicol (0-6.66%), nerol (0.48-8.0%), geraniol (0.26-1.75%) and citral (4.26-6.79%).[6] Recent analysis of the chemical constituents in the essential oils of Ocimum tenuiflorum revealed the presence of α-humulene (0.71%), germacrene D (2.11-4.91%), β-elemene (1.59-6.35%), methyl eugenol (36.47-76.27%) and β-caryophyllene (8.71-56.63%).[7] The essential oils, mainly used in food industries, perfumery, and also possess anti-bacterial and insecticidal properties. It inhibits the in vitro growth of Mycobacterium tuberculosis and Micrococcus pyrogenes variety aureus. It has marked insecticidal activity against mosquitoes. Vermi compost improve herbs quality foliage yield, essential oil content, and oil yield Vermi compost contains Nitrogen, Phosphorous, Potassium, Calcium, Sulphur and Magnesium- Conserved Nutrients from Nature. Vermicompost is the product obtained by composting organic residues using earth worms.[8] Earth worms like Edudrilus eugeniae, Eistenia fetida and Perionyx erxcauatus have been identified as the organisms to process the biodegradable organic material.[9] The chemical composition of basil oil is reported as -pinene 0.1-0.4%, camphene 0.02-0.1%, ßpinene 0.07-0.8%, myrcene 0.12-0.8%, limonene 2.0-9.3%, cis-ocimene 0.1-0.6%, p-cymene 0.05-0.15%, cis-3-hexenol 0.02-0.08%, fenchyl acetate 0.1-0.5%,camphor 0.37-0.75%, linalool 40-54%, fenchyl alcohol 2-9%, methyl chavicol 23-26%, -terpineol 0.8-1.9%, citronellol 0.65-3.7%, geraniol 0.03-0.30%, methyl cinnamate 0.05-0.34% and eugenol 5-12%.[10]

 

Table-1: Physico-chemical properties of the sweet basil oil [11]

Assay of GLC:	Comparable to standard [methyl chavicol (71% to 75%) Linalout (18% to 24%) traces 4-10%]
Appearance:	Pale yellow liquid
Refractive index:	1.453 to 1.655
Saponification value:	2.0000 to 8.0000
Specific Gravity:	0.9320 to o.9830
Optical Rotatim:	- 4 to -8
Specific Gravity:	0.9320

 

MATERIALS AND METHODS:

Plant Material:

Fully grown slips of size 4-6” were planted in field at 60x60 cm spacing in 3x4 m plots as per the lay out plan.

Ocimum tenuiflorum var. CIM-Ayu were collected from the experimental garden of CIMAP, Hyderabad and other chemical are analytical grade, NSP chemicals.

 

Methods:

Experiment I: Different doses of Vermicompost were applied in the following composition   on herb Ocimum tenuiflorum var. CIM-Ayu”

 

Table-2: Different levels of vermicompost applications

S NO	Treatment	Vermicompost tons/ha	Vermicompost Kg/plot
1.	T1	Control	Control
2.	T2	3.00	7.20
3.	T3	1.50	3.60
4.	T4	1.00	2.40
5.	T5	0.75	1.80
6	T6	0.50	1.45

Note: The treatment composition of vermicompost was taken as 3.0, 1.5, 1.0, 0.75, 0.50 tons/ha.

 

Table-3:  Shows  the  replication  of  treatments  with Vermicompost  correspondingly  the    information  has  been  provided  in  parenthesis.

T1 Control
R1T1  (Control)	R2T1  (Control)	R3T1  (Control)	R4T1  (Control)
R1T2 7.20 Kg VC	R2T2 7.20 Kg VC	R3T2 7.20 Kg VC	R4T2 7.20 Kg VC
R1T3 3.60 Kg VC	R2T3 3.60 Kg VC	R3T3 3.60 Kg VC	R4T3 3.60 Kg VC
R1T4 2.40 Kg VC	R2T4 2.40 Kg VC	R3T4 2.40 Kg VC	R4T4 2.40 Kg VC
R1T5 1.80 Kg VC	R2T5 1.80 Kg VC	R3T5 1.80 Kg VC	R4T5 1.80 Kg VC
R1T6 1.45Kg VC	R2T6 1.45Kg VC	R3T6 1.45Kg VC	R4T6 1.45Kg VC

(VC: Vermicompost, T: treatment, R: replication)

 

Experiment –I

Treatment imposition:

The crop was planted in garden on 10.10.2010. Vermicompost  as per treatments was applied 10 days after planting. 

 

Maintenance:

The crop was kept weed free and regularly irrigated.

 

Observations on essential oil yield:

Observations were taken at different intervals starting from 10 days after vermicompost application. In each treatment five plants were removed at random from the plants and washed under tap water. The plants were partitioned   in to lamina, petiole and stem. Fresh and dry weights of individual components were taken separately. Later observations were recorded. Details about the observations recorded are presented here (see Table-4):

 

Table-4: Table showing the different stages of fresh plant samples was collected at regular intervals of time i.e., (30, 60, 90, 120 days)

S.No	Regular intervals	Code
1	30	Stage I
2	60	Stage II
3	90	Stage III
4	120	Stage IV

 

Oil content and quality:

At   45, 90 days after planting leaf samples were collected and separated in to small branches. 250 g leaves were weighed and using a Clevenger apparatus the oil content was estimated. .

 

Statistical Analysis:

The data generated in this study were statistically analyzed following the procedure described by Mandal and Numbiar (1999)^[11][16]. The results are presented and discussed at 5 % probability level uniformly. The non significant results are denoted as N.S and * indicates the significant results. In the statistical analysis, F'-Test was performed with the difference between the highest and

lowest average values. If the difference is more than C.D value, then it is significant and if it is less than C.D value, then it is non significant.

 

Gas Chromatography parameters:

Lemon grass short method:

GC analysis was carried out using Varian CP-3800 with Galaxie chromatography data system fitted with flame ionization detector (FID) and an electronic integrator. Separation of the compounds was achieved employing a Varian CP-Sil 5CB capillary column (ID: 50 m X 0.25 mm; film thickness 0.25 µm) with 5% dimethyl polysiloxane. Nitrogen was the carrier gas at 0.5 ml/min constant flow rate. The column temperature program was: 120⁰C (2 min) to 240⁰C (6 min) at 8⁰C/min ramp rate. The injector and detector temperature were 250⁰C and 300⁰C, respectively. Samples (0.2 µL) were injected with a 20:80:20 split ratio. Retention indices were generated with a standard solution of n-alkanes (C₆-C₁₅). Peak areas and retention times were measured by an electronic integrator. The relative amounts of individual compounds were computed from GC peak areas without FID response factor correction. ^[13][14][15]

 

RESULTS AND DISCUSSION:               

Essential Oil characteristics: During the course of investigation the essential oil from the herbs was collected at three different stages and it was noticed that the essential content did not follow any particular pattern (Table-5 and Figure-1). Similarly, the chemical constituents of the oil were not influenced by the treatments (Table-6, 8, and10).

 

Table-5: Influence of different levels of Vermicompost on the Oil content (%) at different stages.

Treatment	days after planting
	I	II
T1	0.21	0.24
T2	0.23	0.28
T3	0.25	0.30
T4	0.25	0.34
T5	0.25	0.30
T6	0.21	0.31
F'- Test	*	*
C.D.(P=0.05)	0.027	0.036
C.V.%	8.18	8.575

Quantity of essential oil volume yielding in different stages at 45,90 days after vermicompost application. It has been observed that reduction in plant height and number of leaves that was manifested in reduced herb yield was compensated by a higher oil concentration in the leaves.

	Stge-II

 

Figure-1: Influence of different levels of Vermicompost on the essential Oil content (% ) of Ocimum tenuiflorum var. CIM-Ayu at different days after planting

 

An increase in essential oil content was noticed upto 90 days in all the treatments. The essential oil content noticed at observation three was less because the observation was taken during growth period.  It was observed that the essential oil content did not follow any particular pattern. 

 

Chemical constituents of the oil:

Influence of different levels of vermicompost on the chemical constituents of essential oil. Stage I– Chemical constituents of the oil after every 45 days 

 

It has been also observed that, the oil content and chemical composition of oil did not change with regard to influence of the age of the crop.( after every 45 days)

 

 

Compounds	T1	T2	T3	T4	T5	T6
Methyl Eugenol	6.027	0.177	0.159	0.698	0.917	0.161
Linalool	0.302	0.195	0.324	0.537	0.391	0.266
Limonene	0.175	0.356	0.586	1.368	0.328	0.289
Methyl Chavicol	0.733	0.540	0.611	0.816	0.691	0.514
Beta Elemene	6.748	13.251	11.682	8.867	11.783	11.861
Caryophyllene	7.591	10.994	14.805	13.006	11.829	12.197
Eugenol	45.024	70.080	66.441	65.614	65.334	69.580

 

Figure-2: GC Analysis for Vermicompost application of Plant samples after 45 days

 

In the above graph, the highest content of the Eugenol , Limonene, Linalool, Methyl Chavicol, Methyl Eugenol, Beta Elemene and Caryophyllene was present in the essential oil component. It has been represented where as in X-axis Retention Time (RT), min.and Y-axis Height of the peak (µV) (refer to   Figure-2).

 

Table -7: Chemical Composition of oil after 45 days of Vermicompost application

S.No	Name	Time [Min]	Quantity [% Area]	Height [µV]	Area [µV.Min]	Area % [%]
1	Unknown	0.03	0.03	69.7	1.8	0.027
2	Unknown	5.73	0.03	61.8	2.2	0.034
3	Unknown	5.82	0.49	937.6	32	0.492
4	Unknown	5.9	0.01	11.2	0.7	0.01
5	Unknown	6.87	0.33	441.7	21.5	0.331
6	Unknown	7.24	0.02	54.4	1.5	0.023
7	Unknown	7.29	0	13.6	0.2	0.002
8	Unknown	7.65	0.02	31.2	1.2	0.018
9	Unknown	7.72	1.17	2541.8	76.2	1.172
10	Unknown	7.81	0.07	126.2	4.3	0.066
11	Unknown	7.89	0.47	994.8	30.6	0.471
12	Unknown	8.02	0.38	663.9	24.9	0.384
13	Unknown	8.14	0.37	787.2	24.4	0.375
14	Unknown	8.46	0.05	89.6	3.2	0.049
15	Limonene	8.63	1.37	2041.3	88.9	1.368
16	Unknown	8.74	0.3	594.1	19.6	0.302
17	Unknown	8.98	0.03	47.6	2.1	0.033
18	Linalool	9.19	0.54	1028.6	34.9	0.537
19	Unknown	9.25	0.07	146.4	4.6	0.07
20	Unknown	9.35	0.02	22.4	1.3	0.02
21	Unknown	9.99	0.04	63.7	2.3	0.035
22	Unknown	10.21	0.02	26.5	1.2	0.018
23	Methyl chavicol	10.32	0.82	1545.1	53.1	0.816
24	Unknown	10.51	0.49	817.6	32.1	0.494
25	Unknown	11.05	0.04	65.3	2.5	0.038
26	Unknown	11.42	0.07	145.5	4.8	0.074
27	Unknown	11.95	0.14	150.8	8.9	0.136
28	Unknown	12.57	0.16	191.6	10.6	0.163
29	Eugenol	12.77	65.61	122785.4	4263.3	65.614
30	Methyl eugenol	13.26	0.7	1341.4	45.4	0.698
31	Unknown	13.65	0.55	1042.6	35.9	0.552
32	Beta elemene	13.76	8.87	16650.1	576.1	8.867
33	Caryophyllene	14.35	13.01	24559.7	845.1	13.006
34	Unknown	14.5	0.01	14.2	0.8	0.012
35	Unknown	14.83	1.14	1440.4	74	1.14
36	Unknown	15.04	0.12	150.5	7.8	0.12
37	Unknown	15.17	0.07	52.3	4.6	0.071
38	Unknown	15.29	0.21	356.2	13.5	0.208
39	Unknown	15.4	0.27	418.2	17.4	0.268
40	Unknown	15.67	0.02	54.2	1.5	0.023
41	Unknown	15.74	0.03	46.1	1.7	0.026
42	Unknown	15.79	0.02	40.2	1.2	0.019
43	Unknown	16.61	0.12	195.5	7.8	0.12
44	Unknown	16.77	0.16	75.8	10.3	0.158
45	Unknown	16.99	0.06	50.2	4	0.062
46	Unknown	17.53	0.05	72	3.2	0.05
47	Unknown	17.68	0.16	98.6	10.1	0.156
48	Unknown	20.75	1.27	821.9	82.6	1.271
Total			100	183976.5	6497.6	100

The chemical composition contents was present in above mentioned table shows that the out of 48 unknown samples the only seven components (viz. Eugenol ,Limonene, Linalool, Methyl Chavicol, Methyl Eugenol, Beta Elemene and Caryophyllene) were shown the considerable enhancement in terms of quantity after adding the required quantity (2.40 kg/Plot) vermicompost application.( see Table-7)

 

Stage II -Chemical constituents of the oil after every 90 days 

Table-8: Quantity of oil after every 90 days of  Vermicompost application.

Compounds	T1	T2	T3	T4	T5	T6
Linalool	0.363	0.418	0.424	0.341	0.302	0.330
Limonene	0.665	0.673	1.410	0.555	0.474	0.490
Methyl Chavicol	0.720	0.576	0.695	0.568	0.463	0.533
Beta Elemene	11.031	13.394	8.110	9.282	11.748	10.521
Caryophyllene	18.322	17.822	19.967	15.205	11.288	18.012
Methyl Eugenol	1.049	0.098	28.126	8.307	0.127	0.099
Eugenol	61.163	60.665	33.558	61.145	71.509	65.250

 

Figure-3: GC Analysis for Vermicompost application of Plant samples after 90 days 

 

It has been also observed that, the oil content and chemical composition of oil did not change with regard to  influence of the age of the crop.( after every 45 days.)

 

In the above graph, the highest content of the Eugenol ,Limonene, Linalool, Methyl Chavicol, Methyl Eugenol, Beta Elemene and Caryophyllene was present in the essential oil omponent. It has been represented where as in X-axis Retention Time (RT), min.and Y-axis Height of the peak (µV).( refer to Figure-3)

 

Table-9:Chemical Composition of oil after 90 days of Vermicompost application

S.No	Name	Time [Min]	Quantity [% Area]	Height [µV]	Area [µV.Min]	Area % [%]
1	Unknown	5.87	0.25	2994.8	113.4	0.249
2	Unknown	6.08	0.01	127.8	5.9	0.013
3	Unknown	6.67	0.01	111.2	4.5	0.01
4	Unknown	6.87	0.16	1581.4	72.9	0.16
5	Unknown	7.16	0.01	110.2	3.9	0.009
6	Unknown	7.24	0.01	149.5	5.1	0.011
7	Unknown	7.59	0.01	138.8	4.3	0.009
8	Unknown	7.72	1.17	16570.4	534.8	1.174
9	Unknown	7.81	0.05	636.4	21.6	0.047
10	Unknown	7.88	0.35	4934.8	161	0.353
11	Unknown	8.02	0.38	4684	171	0.375
12	Unknown	8.14	0.28	3866.1	125.4	0.275
13	Unknown	8.46	0.07	973.7	32.9	0.072
14	Limonene	8.63	1.41	10661.1	642.6	1.41
15	Unknown	8.74	0.76	10224	344.1	0.755
16	Unknown	8.86	0.03	192.4	11.5	0.025
17	Unknown	8.98	0.03	258.7	13.1	0.029
18	Linalool	9.19	0.42	5086.8	193.1	0.424
19	Unknown	9.25	0.1	1361.4	47.2	0.104
20	Unknown	9.33	0.02	138.4	7.2	0.016
21	Unknown	9.67	0.01	126	4.2	0.009
22	Unknown	9.77	0.02	187.6	8.1	0.018
23	Unknown	9.99	0.07	710	33	0.072
24	Unknown	10.21	0.03	259.2	11.9	0.026
25	Methyl chavicol	10.32	0.69	8964.5	316.5	0.695
26	Unknown	10.43	0.03	314.1	13.8	0.03
27	Unknown	10.51	0.36	2862	164.5	0.361
28	Unknown	11.42	0.02	240.1	9.2	0.02
29	Unknown	11.95	0.02	247.2	8.5	0.019
30	Unknown	12.02	0.01	72.1	3.2	0.007
31	Unknown	12.15	0.02	305.8	10.6	0.023
32	Unknown	12.24	0.01	103.5	5.6	0.012
33	Eugenol	12.79	33.56	351512.5	15289.6	33.558
34	Unknown	13.03	0.06	147	27.8	0.061
35	Methyl eugenol	13.29	28.13	316007.4	12814.6	28.126
36	Unknown	13.58	0.02	140.9	8.6	0.019
37	Unknown	13.65	0.49	6419.8	224.1	0.492
38	Beta elemene	13.76	8.11	103937.9	3695.2	8.11
39	Unknown	14.12	0.03	369.4	15.6	0.034
40	Caryophyllene	14.36	19.97	249302.5	9097.2	19.967
41	Unknown	14.58	0.15	685.1	67.9	0.149
42	Unknown	14.83	1.11	13976.6	504.4	1.107
43	Unknown	15.05	0.04	389.9	16.7	0.037
44	Unknown	15.12	0.01	90.9	3.7	0.008
45	Unknown	15.17	0.03	190.8	12	0.026
46	Unknown	15.29	0.1	1159.2	46.2	0.101
47	Unknown	15.4	0.11	1349.4	50	0.11
48	Unknown	15.45	0.04	375.3	16.3	0.036
49	Unknown	15.58	0.03	248.8	14.7	0.032
50	Unknown	15.67	0.05	648.1	24.5	0.054
51	Unknown	15.74	0.05	401	21.1	0.046
52	Unknown	15.79	0.03	333.7	12.8	0.028
53	Unknown	15.85	0.04	274.6	18.6	0.041
54	Unknown	16	0.02	195.7	9.9	0.022
55	Unknown	16.38	0.01	77.4	2.7	0.006
56	Unknown	16.55	0.03	356.1	12.7	0.028
57	Unknown	16.61	0.25	3003.7	113.1	0.248
58	Unknown	16.76	0.1	839.3	45.9	0.101
59	Unknown	16.94	0.05	279.1	23.8	0.052
60	Unknown	17.26	0.05	372.4	24.1	0.053
61	Unknown	17.53	0.07	445.6	33.9	0.074
62	Unknown	17.64	0.02	135.1	7.8	0.017
63	Unknown	17.93	0.01	108.5	6	0.013
64	Unknown	18.21	0.03	262.1	12.8	0.028
65	Unknown	18.47	0.09	652.2	39.1	0.086
66	Unknown	19.37	0.3	673.1	135.5	0.297
67	Unknown	19.85	0.02	96.7	7.9	0.017
Total			100	1134652	45561.4	100

 

The chemical composition contents was present in above mentioned table shows that the out of 67 unknown samples the only seven components (viz. Eugenol ,Limonene, Linalool, Methyl Chavicol, Methyl Eugenol, Beta Elemene and Caryophyllene) were shown the considerable enhancement in terms of quantity after adding the required quantity (2.40 kg/Plot) vermicompost application.( refer Table-9)

 

CONCLUSIONS:

The influence of vermicompost on the essential oil yield of ocimum at two different stages (i.e. 45 and 90 days) was studied by using standardization method of oil analysis it was observed that the essential oil content did not follow any particular pattern.  The eugenol content of the oil was higher due to vermicompost application.

 

REFERENCES:

1        Simon, J.E., A.F. Chadwick, and L.E. Craker. 1984. Basil. In:Herbs: An indexed bibliography, 1971-1980. Archon Books.

2        Bast F, Rani P, Meena D. Chloroplast DNA phylogeography of holy basil (Ocimum tenuiflorum) in Indian subcontinent.ScientificWorldJournal.2014;847–482.

3        Krishnan R. Natural outcrossing in sweet basil-Ocimum basilicum L. Indian Perfumer, 1981; 25(3and4): 74-77.

4        Joy, P.P., Thomas, J., Mathew, S., Jose, G. and Joseph, J. Aromatic plants. Tropical Horticulture Vol. 2. (eds. Bose, T.K., Kabir, J., Das, P. and Joy, P.P.). Naya Prokash, Calcutta, 2001;633-733

5        Sobti SN. and Pushpangadan P. Studies in the Genus Ocimum : to genetics, Breeding and Production of New Strains of economic Importance, in CK. Atal and BM Kapur (eds), Cultivation and Utilization of Aromatic Plants. RRL, CSIR, Jammu-Tawi, 1982 :457-472.

6        Omer E.A., Elsayed A-G.A., El-Lathy A., Khattab M.E., Sabra A.S., Effect of the nitrogen fertilizer forms and time of their application on the yield of herb and essential oil of Ocimum americanum L. Herba Pol. 2008; 54(1):34–46.

7        Vani, S.R., S.F. Cheng and C.H. Chuah, Comparative Study of Volatile Compounds from Genus Ocimum. Am. J. Applied Sci. 2009; 6(3): 523-528.

8        Gandhi, M., Sangwan, V., Kapoor, K.K. and Dilbaghi, N. Composting of household wastes with and without earthworms. Environment and Ecology., 1997; 15(2): 432-434.

9        Patil, S.L and Sheelavantar M.N. Effect of moisture conservation practices, organic sources and nitrogen levels on yield, water use and root development of rabi sorghum [Sorghum bicolor (L.)] in the vertisols of semiarid tropics. Annals of Agricultural Research., 2000; 21(21) :32–36.

10     Kumar, A., Sharma, A. and Virmani, O. P. Cultivation and utilization of rose geranium: A review. Cur. Res. Med. Arom.Plants., 1985; 7(3):137-147.

11     A. Ranganadha Reddy, R. Bharath Kumar, Dr.V.Ravi Kumar, M. Deepthi, T. Naga Lohita ,M. Sriharsha, K.P. Sastry Experimental Studies on effect of Vermicompost and NPK on Essential oil yield of Ocimum tenuiflorum var. CIM-Ayu Research Journal of Pharmacy and Technology. 2015; 8(11):1519-1525

12     Risto Uusitalo, et al,. Effect of gypsum on the transfer of phosphorus and other nutrients through clay soil monoiliths, Agriculture and Food Science Journal., 2012; 21:260 – 278.

13     Ranganadha Reddy, V. Ravi Kumar, M. Priyanka, K.P. Sastry Experimental Studies on Influence of Different Doses of Gypsum on Essential Oil Yield of Lemongrass var. Krishna (Cymbopogon flexuosus) Research Journal of Pharmacy and Technology. 2014; 7(4):439-446.

14     Ranganadha Reddy, V. Ravi Kumar, R. Bharath Kumar, M.Priyanka, K.P. Sastry Experimental Studies on Influence of Different Doses of Sulphur on Essential Oil Yield of Lemongrass Var. Krishna (Cymbopogon flexuosus) Research Journal of Pharmacy and Technology. 2014; 7(8):856-863.

15     Nambiar, P. T. N and mandal R.C, Agricultural statistics :techniques and procedures, Agro Botanica Bikaner, 1999; 200P..

 

 

Accepted on 10.08.2016        © RJPT All right reserved

Research J. Pharm. and Tech 2016; 9(12):2087-2093.