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1346Current Trends in Biotechnology and PharmacyVol. 5 (3) 1346 -1352 July 2011, ISSN 0973-8916 (Print), 2230-7303 (Online)

AbstractMedicinal plants growing in wild cannot

ensure consistent quality due to geneticvariability found in the natural populations,geographical variations and season of collection.Therefore, there is a need to cultivate medicinalplants using known varieties for extraction ofhigh yields of active principles. This can beachieved only with organic farming practices andpost harvest technologies. The researchconducted by institutes like CIMAP and otherCSIR institutes, ICAR and Universities has ledto the release of a number of high yieldingmedicinal plant cultivars. Coleus forskohli is aherb with fleshy, fibrous roots that grows in wildin the warm, subtropical areas like in India,Burma, and Thailand. Coleus forskohlii is beinggrown in several parts of the country as acommercial crop. In the present investigation theeffect of nitrogen, phosphorus, potassium (NPK)and farm yard manure amendments on thegrowth of roots and accumulation of forskolinwas studied. The results revealed that 50% ofNPK and 10 tonnes of farm yard manure perhectare at a harvesting time 150 days are idealfor maximum yield of forskolin.

Key words: Forskolin, Farm Yard Manure, NPK,Coleus forskohlii

IntroductionCultivation of medicinal plants appears

to be an important strategy for research anddevelopment. More than 400 plant species usedfor production of medicines by the IndianIndustry. Cultivation of various varieties ofmedicinal plants is important to isolate the activeprinciples. The application of organic farmingpractices and post harvest technologies canenhance their yields so that it becomes profitableto the farmers. The potential returns to the farmerfrom cultivation of medicinal plants are reportedto be quite high if fertilizers are used at a properdose and time. The application of bio-fertilizersin combination with the inorganic fertilizerssubstitutes the required quantity of the fertilizersneeded by the medicinal and aromatic crops. Thebio-fertilizers due to its eco friendliness willchange the phase of medicinal and aromatic cropsproduction in the years to come (1). InChlorophytum borivallianum, the number oftubers, fresh and dry tuber yields and steroidcontent increased with the application ofvermicompost of 5 tonnes ha-1 (2).

When Coleus forskohli was grown in FYMamended soil with Paecilomyces lilacinus, therewas better root development due to control of

Effect of NPK and Farm Yard Manure on the Accumulation ofForskolin, an Anti-Obesity Diterpenoid Molecule in Coleus

forskohlii

S. Nageswara Rao, S. Gangadhar Rao, B. Bhargavi and K. Janardhan Reddy*Department of Botany, Osmania University, Hyderabad 500 007, India

*For correspondence - [email protected]

Effect of NPK and farm yard manure on the accumulation of forskolin


nematodes (3). Earlier studies on Coleusforskohlii, with a combination of 40 kg N, 60 kgP

2O

5 and 50 kg K

2O ha-1 resulted in optimum

fresh and dry tuber yields of 13.12 tonnes ha-1

(4). Raising the level beyond this was found toreduce the tuber yield significantly. Nitrogen at60 kg ha-1 produced higher tuber yield, high drymatter yield but maximum bulking rate alongwith 120 kg K

2O ha-1 in Coleus parviflorus (5).

Somanath et al. (6), in their study on influenceof FYM and NPK on uptake of nutrients andgrowth of Coleus forskohlii found that maximumincrease in fresh weight of roots was recorded inall the stages of growth (at 30 DAP 0.135 kgplant-1, at 90 DAP 0.347 kg plant-1 and at 150DAP 0.600 kg plant-1).

Application of N at 100 kg ha-1 (half basal+ half one month after transplanting) along witha basal application of 100 kg ha-1 P2O5 and 20tonnes FYM ha-1 increased the fresh weight ofrhizomes in Acorus calamus (7). In Aswagandha(Withania somnifera), high dry weight of roots(1.297 g plant-1) were obtained with 15 kg N ha-

1 (8). In Catharanthus roseus, 100 kg N, 40 kgP

2O

5 and 50 kg K ha-1 gave maximum root yields

(9). Application of N (30kg N ha-1) increased theroot yields (2702 kg ha-1) in Rauwolfia serpentina(10). Highest root yield of Aswagandha wasobtained with the application of 20 kg N and 40kg P

2O

5 ha-1 (11).

But there are no reports on the influence ofnutrients (inorganic and bio- fertilizers) on thecontent of forskolin, the main chemicalconstituent of Coleus forskohlii. Keeping thisview in mind, the present investigation was takenup to enhance the forskolin levels under differentfertilizer amendments. The aim of the presentinvestigation is to increase the yields of rootsand forskolin content under different fertilizeramendments so as to make it profitable to thefarming community.

Materials and methodsPreparation of soil bed :The experimental areawas repeatedly ploughed; finally, it was leveledand divided into plots as per the layout. Theexperiment was laid out in a randomized blockdesign with 6 treatments and replicated thrice. Aplot size of 3 X 2.5 m was taken for experimentalwork.

Treatments of fertilizers and farm yard to thesoil for cultivation of Coleus forskohliiThe details of the treatments are:T

1: Control – No manures

T2

: 100% NPK recommended doseT

3: 50% NPK + 10 tonnes of farm yard manure / ha

T4

: 50% NPK + 5 tonnes of vermicompost/ haT

5: 20 tonnes of farm yard manure / ha

T6

: Vermicompost 10 tonnes/ha

Extraction of forskolin : Coleus forskohlii roots(10 kg) were extracted with methanol (25 x 3) atroom temperature. After this, methanol wasdistilled out to give a residue of 1.5 kg. Thisresidue was further extracted with Petroleum-Ether (5 x 5) to give another residue of 230 gcontaining waxes and fats. After this, defattedresidue was extracted with ethyl acetate to givea residue of 270 g, which contains 27% forskolin.

Fresh and dry weights of the root : The influenceof different fertilizer amendments on freshweight and dry weights of the root [kg plant -1]at 30, 90 and 150 Days after Planting (DAP) ofgrowth in Coleus forskohlii was recorded.

HPLC analysis of forskolinConditions:Mobile phase : Mehanol :Water (65:35 v/v)Column : Hypersil C-18 (250 mm* 4.6 mm )

Detection : Refractive indexFlow rate : 1 µl/minInjection volume : 20 micro lit.

Coleus forskohlii (roots) was cultivatedunder different treatment conditions, and

Nageswara Rao et al


forskolin was extracted in a soxhlett apparatuswith methanol and concentrated. TLC solventsystem (Rf 0.45, 20% EtOAc + petroleum ether)showed the presence of forskolin. The extractswere subjected to HPLC analysis.

ResultsFresh weight of the root : The crop washarvested at 30, 90 and 150 days aftertransplantation. Light irrigation was given oneday before harvesting for easy lifting up of roots.Harvesting was carried out manually by diggingand uprooting the individual plants. The rootswere separated and washed to remove the soil.Forskolin content was determined in roots 150days after transplantation. The influence ofdifferent fertilizer amendments on fresh weightof the root [kg plant -1] at 30, 90 and 150 DAP ofgrowth in Coleus forskohlii is shown in table 1.Maximum increase in fresh weight of roots wasrecorded in treatment number 3 in all the stages(at 30 DAP 0.135 kg plant-1, at 90 DAP 0.347 kgplant-1 at 150 DAP 0.600 kg plant-1) of growth.

Dry weight of the root and content of forskolin: Influence of various fertilizer amendments ondry weight of the root [kg plant -1] at 30, 90 and150 DAP of growth in Coleus forskohlii is shown

in Table 2. In treatment number 3, maximumfresh weight was observed i.e., at 30 DAP 9.1 gplant-1, at 90 DAP 18.9 g plant-1 at 150 DAP 60.0g plant-1. The fresh and dry weights of roots weremeasured under different nitrogen regimes andare shown in Table 3. The treatment 50% NPK +10 tonnes of farm yard manure gave the best dryroot weight among the different treatments (Table3). Samples were collected under differenttreatments for quantification of forskolin.Forskolin was extracted in a soxhlett apparatus,concentrated and used for HPLC analysis (Fig.1). HPLC analysis revealed that application of50% NPK + 10 tonnes of farm yard manure ha-1

gave maximum yields of forskolin after 150 daysof harvest.

DiscussionSomanath et al. (6), in their study on

influence of FYM and NPK on uptake ofnutrients and growth of Coleus forskohliiobserved maximum increase in fresh weight ofroots in treatment number 3 (at 30 DAP 0.135kg plant-1, at 90 DAP 0.347 kg plant-1 at 150 DAP0.600 kg plant-1). Application of N at 100 kgha-1 (half basal + half one month aftertransplanting) along with a basal application of

Table 1. Effect of various fertilizer amendments on fresh weight of root (g per plant) at differentstages of growth in Coleus forskohlii

Treatments Days after planting

30 90 150

T1

: Control – No manures 58 120 267

T2

: 100% NPK recommended dose (N- 40 kg ha-1) 83 340 490

T3

: 50% NPK + 10 tonnes of farm yard manure/ha 149 468 600

T4

: 50% NPK + 5 tonnes of vermicompost/ha 100 375 510

T5

: 20 tonnes of farm yard manure/ha 69 160 310

T6

: Vermicompost 10 tonnes/ha 73 190 385



Table 2. Effect of various fertilizer amendments on dry weight of root (g per plant) at differentstages of growth in Coleus forskohlii

Treatments Days after planting30 90 150

T1: Control – No manures 3.8 8.2 26.8

T2: 100% NPK recommended dose (N- 40 kg ha

-1) 7.3 16.5 47.5

T3: 50% NPK + 10 tonnes of farm yard manure/ha 9.1 18.9 60

T4: 50% NPK + 5 tonnes of vermicompost/ha 6.2 16.7 55

T5: 20 tonnes of farm yard manure/ha 4.5 11.7 29.4

T6: Vermicompost 10 tonnes/ha 5.1 14.9 44

Nageswara Rao et al


100 kg ha-1 P2O5 and 20 tonnes FYM ha-1

increased the fresh weight of rhizomes in Acoruscalamus (7). Sheu et al. (12) also observed higheryield (23 tonnes ha-1) in Dioscorea doryophorawith organic and inorganic fertilizeramendments. In treatment number 3, maximumfresh weights were recorded (Table 3 and Fig.4.). In Aswagandha (Withania somnifera), dryweight of roots (1.297 g plant-1) were obtainedwith 15 kg N ha-1 (Maryada et al., 2001). InCatharanthus roseus, 100 kg N, 40 kg P

2O

5 and

50 kg K2O) ha-1 gave maximum root yields

(Hegde, 1988). Highest root yield of aswagandhawas obtained with the application of 20 kg Nand 40 kg P

2O

5 ha-1 (11). In aswagandha,

application of 40 kg N ha-1 and 60 kg P2O

5 ha-1

and 20 kg K2O ha-1 gave the highest dry root

yields of 770.37 kg ha-1 (13).

Root characters like number of roots perplant and length of roots increased inChlorophytum borivallianum, along with thenumber of tubers, fresh and dry weights of tuberyield and steroidal content with the applicationof vermicompost of 5 tonnes ha-1. The steroidalcontent was the highest with vermicompostapplication (2). Maheshwari et al. (1988) alsoobserved increase in the root yield in Rauwolfiaserpentaina with the application of N (30 kg Nha-1). Application of NPK at 30:30:0 kg ha-1

recorded the highest dry root yields ofAswagandha but further increasing the levels of

Table 3. Fresh and weights of roots grown under different nitrogen regimes. N = Replicates, M= Mean, SE = Standard error

30 days 90 days 150 days

Treatment N M-SE M+SE M-SE M+SE M-SE M+SE

T1 Fresh weight of root 3 58 59.33 120 121.33 267 267.67

Dry weight of root 3 3.73 3.8 8.2 8.27 26.8 27.27




Dry weight of root 3 9.24 10.03 18.85 19.09 60 60.67

T4 Fresh weight of root 3 100 100.67 374.33 375 510 510.67


T5 Fresh weight of root 3 69 69.67 160 160.67 309.33 310




T1 = control; T2 = 100% NPK; T3 = 50% NPK + 10 tonnes of farm yard manure;T4 = 50% NPK + 5 tonnes of vermicompost; T5 = 20 tonnes of farm yard manure;T6 = 10 tonnes of vermicompost.



fertilizers had no effect on the yields of dry roots(14). Highest root yield of aswagandha wasobtained with the application of 20 kg N and 40kg P

2O

5 ha-1 (11). HPLC analysis revealed that

application of 50% NPK + 10 tonnes of farmyard manure ha-1 gave maximum yield offorskolin. The statistical analysis revealed thatfresh and dry weights of roots, number of rootsplant-1 and root length were significant comparedto controls and positively correlated with eachother. It is suggested that appropriate dose ofapplication of fertilizers at appropriate time willresult in higher tuber yield and forskolin content.Our results also suggest that time of harvest foroptimum yields of tuber is also important.

References1. Srinivasappa, K.N., Anuradha, M.N.,

Farooqi, A.A., Kathiresan, C., Suresh, H.C.,and Shivananda, T.N. (2007). Use of bio-fertilizers in cultivation of medicinal andaromatic plants - a review. BIOMED, 2:235-256.

2. Paturde, J.K., Wankhade, S.G., Khode P.P,Chatol, P.U, Deo, D.D. and Bhuyar, S.A.(2002). Effect of organic manures and plantpopulation on yield of safed musli(Chlorophytum borivallianum).Agricultural Science Digest, 22: 51-52.

3. Sivakumar, C.V., and Vidhyasekaran, P.(1990). Control of Meloidogyne incognitaon Coleus Forskohlii with Paecilomyceslilacinus in farm yard manure amended andnon-amended soil. Journal of BiologicalControl, 4: 68-69.

4. Veeraraghavathatham, D, Venkatachalam,R., and Sundararajan., S. (1988). Effect ofvarious levels of N, P and K on the tuberYield of Coleus forskohlii. South IndianHorticulture, 36: 252-257.

5. Geetha, K., and Madhavan Nair., K.P.(1993). Effect of N and K in Coleus. Journalof Tropical Agricultural, 31: 198-203.

6. Somanath, I., Bhaskar, S., andSreenivasmurthy C.A. (2007). Influence ofFYM and NPK on uptake of nutrients andgrowth of Coleus forskohlii. BIOMED, 2:265-271.

7. Tiwari, J.P., Tiwari, A.B., Tiwari, G., andSingh A.K. (2000). Effect of nitrogenapplication on growth and yield of Acoruscalamus. Proceedings of the NationalSeminar on the Research and Developmentin Aromatic Plants. Current trends inbiology, uses, production and marketing ofessential oils, Lucknow, India, 30-31 July1999. Journal of Medicinal and AromaticPlant Sciences, 22: 636-638.

8. Maryada, M, Kaushal, S.K and Kaistha,B.P. (2001). Response of aswagandha(Withania somnifera) to nitrogenfertilization under mid hill conditions ofHimachal Pradesh. Himachal Jourmal ofAgriculture Research, 27: 77-80.

9. Hedge, D.M. (1988). Response ofperiwinkle (Catharanthus roseus L.) tonitrogen, phosphorus and potassiumfertilization. Agricultural Research Journalof Kerala, 26: 227-233.

10. Maheshwari, S.K., Yadav, S, Gangrade, S.Kand Trivedi K.C. (1988). Effect of fertilizerson growth, root and alkaloid yield ofrauwolfia (Rauwolfia serpentina). IndianJournal of Agricultural Sciences, 58: 87-488.

11. Maheshwari, S.K. and Yadav, S. (1981).Response of Withania somnifera to variablelevels of N and P rainfed. Paper presentedat the IV All India Workshop on Medicinal

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12. Sheu, B.W., Juan Y.H. and Yan H.F. (2000).The effects of organic and inorganicfertilizers on the growth and yield of Herng-Chuen Yams, Dioscorea doryophora.Bulletin of National Pingtung Universityof Science and Technology, 8: 103-107.

13. Muthumanickam, D and Balakrishna-murthy G. (1999). Studies on nutritional

requirement for aswagandha (Withaniasomnifera) in sheuroy hills of Tamilnadu,India. Journal of species and AromaticCrops, 8: 179-183.

14. Nigam K.B., Rawat G.S. and Prasad B.(1984). Effect of methods of sowing, plantdensity and fertility level on aswagandha(Withania somnifera Dunal.). South IndianHorticulture 32: 356-359.


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