Pharmacopoeia. Ayurveda Pharmacopoeia4. Distribution. An endemic species to
Sri Lanka. It is grown in Kandy, Kadugannawa and Kurunegala. It.
SRI
LANKAN
MEDICINAL
MONOGRAPHS
AND
VOL -
LAKSHMI ARAVINDA
PLANT
ANALYSIS
11
ARAMBEWELA WIJESINGHE
SRI LANKAN MEDICINAL
PLANT
MONOGRAPHS AND ANALYSIS VOL-11
PLECTRANTHUS
£ >om»2
ZEYLANICUS
Lakshmi Arambewela and Aravinda Wijesinghe
Edited by Dilmani Warnasuriya
Industrial Technology Institute (Ceylon Institute of Scientific and Industrial Research) 363, Bauddhaloka Mawatha Colombo 07 Sri Lanka
Published by National Science Foundation 2006
©Industrial Technology Institute & National Science Foundation, 2006. First published in 2006. ISBN 955-590-045-0 Published by National Science Foundation.
Arambewela, Lakshmi Sri Lankan Medicinal plant Monograph and Analysis: Plectranthus zeylanicus / Lakshmi Arambewela and Aravinda Wijesinghe.Colombo: National Science Foundation, 2006. Vol.11, iii,27p. ill.; 30 cm. -(Sri Lankan Medicinal Plants, Vol.11) ISBN 955-590-045-0
Price: Rs. 200.00
i. 615.321 DDC21 iii. Series iv. Arambewela, Lakshmi jt.au. v. Wijesinghe, Aravindajt.au. 1. Medicinal plants
ii. Title
2. Botany, Chemistry, Medical
The information found in this monograph is taken from available scientific literature. The authors accept no liability for any damages arising from any claims contained in this text except the work carried out by the authors.
ii
Preface
Studies on medicinal plants of Sri Lanka have been carried out in the Herbal Technology Division of Industrial Technology Institute (former Ceylon Institute of Scientific and Industrial Research) for almost two decades. This monograph which is the eleventh in this series incorporates information collected from literature surveys, researches and also experiences of the Herbal Technology Division staff. This monograph is intended for a varied reading public, herbal drug manufacturers who need to identify their herbal raw materials, Ayurvedic physicians who need some scientific information on medicinal plants, research workers requiring some quick background information on a plant, industrialists or entrepreneurs pondering on commercial ventures and the inquiring lay readers. W e hope this monograph fulfils some requirements of each of them. The authors wish to thank the members of the Herbal Technology Division for their contribution, the Information Service Center for providing information, Department of Plant Sciences and Department of Zoology of the University of Colombo for assisting in anatomical studies, Food Technology Division of the Industrial Technology Institute for helping in the analysis of powdered plant materials and the Microbiology Laboratory for photographing the slides. They also gratefully acknowledge the sponsor National Science Foundation for the research grant (RG / 2004 / T M / 01).
Herbal Technology Division Industrial Technology Institute P.O.Box 787 Colombo 07 Sri Lanka.
in
Plectranthus zeylanicus Benth Family Lamiaceae
Synonyms 1
Coleus zeylanicus (Benth) Cramer
Selected V e r n a c u l a r Names 1,2
Sinhala - Iriweriya 1,2
Sanskrit-Valakan
3
Tamil- Vettiyar
Pharmacopoeia 4
Ayurveda Pharmacopoeia
Distribution An endemic species to Sri Lanka. It is grown in Kandy, Kadugannawa and Kurunegala. It 1,2,3
can be easily grown in prevailing climatic and soil conditions in Sri Lanka.
Morphology
2
A herbaceous, perennial herb, about 60-90 cm tall, stems branching, somewhat prostrate, bluntly quadrangular, reddish and hairy, leaves simple, opposite, aromatic 5.3-6.5 cm somewhat succulent, dentate, light green on the upper surface, paler below, veins prominent beneath, petioles 2.5-3 cm long, reddish, hairy, faintly grooved above, flowers irregular, bisexual, light blue, 1 cm long and narrow in whorls, each containing about 10 flowers opening at different times along a terminal or branched raceme 15-23 cm long, pedicels 3 m m long reddish and hairy, sepal the largest, broadly ovate, apiculate, others lanceolate acuminate, acute, corolla 2-lipped consisting of 5 segments, upper lip of 4 short segments and a long, boat- shaped lower lip, stamens 4, didynamous, epipetalous, 2 slightly shorter, all included, ovary superior, 2-carpellary, 4-locular with a single ovule in
l
each loculus, style 7 m m long, stigma bifid; fruit of 4, one seeded achenes at the base of a persistent calyx.
Fig-1. Plectranthus zeylanicus plant 1. Leaf
2. Stem
3. Flower, lateral view
4. Inflorescence
(Source - Jayawcera, D.M.A, Medicinal plants (1982). part V, p 109)
PLANT MATERIAL OF INTEREST
Whole plant'
Official Drug Whole plant
3
Pharmacognostic Features* Anatomy
Fig-2. Cross section of I. Epidermis
Plectranthus zeylanicus
2. Palisade parenchyma
bundles 5. Parenchyma cells
leaf (stained with safranine (10x10))
3. Spongy parenchyma
6. Trichomcs
4. Vascular
Fi«-4. Cross section of Plectrantluts 1. Epidermal hairs 6. Cambium
7. Xylem
2. Epidermis
zeylanicus
stem (stained with safranine (10x10))
3. Collenchyma
8. Pith
4
4. Cortex
5. Phloem
Powder analysis Analyzed part
Aerial parts of the plant
Oruanoleptic properties Colour - Brownish black Odour - Aromatic Taste - No special taste
Microscopic characters
Fig-6. Powder of aerial parts of Plcctranthus 1. Vessel elements
zeylanicus
2. Trichome
under the microscope
3. Parenchyma cells
Fig-7. Schematic diagram of powder of aerial parts 1. Epidermal cells with stomata
2. Stomata with guard cells
4. Vessel segments with different thickenings
3. Trichomes
5. Epidermal cells
Analyzed part - Root Organolcptic properties Colour - Brownish black Odour - No special odour Paste - No special taste M icroseopic characters
Fig-8. Powder of root of'Plectranthus zeylanicus under the microscope. I. Vessel segments
2. Parenchyma cells
ft
Fig-9. Schematic diagram of powder microscopy of roots 1. Root hairs
2. Parenchyma cells
3. Vessel segments with different thickenings
• These analysis were carried out by the authors at Industrial Technology Institute and the Dept. of Plant Sciences and Dept. of Zoology of University of Colombo
6
Physico-chemical A n a l y s i s Extractable
12
matter
Crushed, dried plant material (about 4 g) was weighed to a glass-stoppered conical flask. Solvent (100 mL) was added, weighed, shaken well and allowed to stand for lh. It was then boiled for lh and cooled. The weight was readjusted with specified solvent and filtered. Filtrate (25 mL) was taken, solvent was evaporated and oven dried at 105 °C for 6 h, cooled in a desiccator and weighed.
Total ask
Crushed, air dried plant material (about 4 g) was weighed to a previously ignited crucible. The material was ignited by gradually increasing the temperature to 550 °C until free from carbon. The crucible was cooled and weighed.
Acid insoluble ash
Hydrochloric acid (25 mL, cone. -70 g/L) was added to the crucible containing total ash, covered with a watch glass and boiled gently for 5 min. The insoluble matter was collected on an ashless filter paper and washed with hot water until the filtrate was neutral. Thefilterpaper containing the insoluble matter was transferred to the original crucible and ignited to a constant weight.
Water soluble ash
Water (25 mL) was added to the crucible containing total ash, covered with a watch glass and boiled gently for 5min. The insoluble matter was collected on an ashlessfilterpaper and washed with hot water. Thefilterpaper containing the insoluble matter was transferred to the original crucible and ignited for 15 min. at a temperature not exceeding 450 °C. Water soluble ash is the calculated difference in weight between the total ash and the residue remaining after treatment of the total ash with water.
Moisture content of the samples was estimated and all the calculations were done on dry weight basis.
7
Table 1. Physico-chemical parameters of Plectranthus zeylanicus**
Physico-chemical parameter Ethanol extract of aerial parts Ethanol extract of root Water extract of aerial parts Water extract of root
A mo unt % 14.4-15.1 3.5-3.8 33.3-35.4 7.0-7.5
Total ash content of aerial parts
13.6-15.9
Total ash content of root
13.2-13.6
Acid insoluble ash content of aerial parts
0.9-1.0
Acid insoluble ash content of root
6.8-6.9
Water soluble ash content of aerial parts Water soluble ash content of root
11.1-14.9 0.2-0.3
(Results arc expressed as percentages on dry weight basis)
T H I N L A Y E R C H R O M A T O G R A P H I C PROFILE
Plectranthus zeylanicus water extract of aerial parts Sample preparation
: P. zeylanicus
aerial parts (4 g) were boiled for one hour with
water (100 mL) and the extract was filtered and evaporated to dryness. Ten microliters (10 uL) of the diluted extract (0.1 g in 5 mL) was spotted on TLC plate. Adsorbent
: Silica g e l - G F 5 4
Solvent system
: Ethyl acetate : Acetic acid : Water (4.6 : 0.2 : 0.2)
2
Detection Direct evaluation
:
Scanning
: Densitometer at 254 nm (before spraying) and 450 nm (after
UV
2 5
4nm,
Rf values - 0.39. 0.52. 0.93
spraying) Spray reagent
Fig-10.
T
L
: Vanillin sulphate
C
f i n g e r
p r i n t
p r o f i l e
o f
w a t e r
e x t r a c t
9
o f
Plectranthus zeylanicus
a e r i a l
p a r t s
a. ?.
oo
l> . 250
i"
0 . 20 j
•:7*
"ii;!
a -15 c
•
i!'.2
•
•
•) f n
.... t.c. :
i \ . osa
3
'
!
Table 2. Description of densitogram (Fig-V1)
:
Peak no.
Y(mm)
1
8.10
Relative area % 20.87
2
12.12
5.31
3
20.05
3.45
4
65.45
31.21
r
; ^ T \ ; V
. ooo
.
c
o
Table 3. Description of densitogram (Fig-12)
O
Peak no.
Y(mm)
1
9.61
Relative area % 24.16
2
20.94
5.10
3
28.25
7.44
4
34.18
7.02
5
66.80
32.67
1;!J0 - •
O.01C.i)iO.(.'.".O.O-y. Or'.'.OiO.r,?}.
,
,
Fig-12. Densitogram of TLC finger print profile of water extract of Plectranthus zeylanicus aerial parts at 450 nm
10
Plectranthus
zeylanicus ethanol extract of aerial parts
Sample preparation
: P. zeylanicus aerial parts ( 4 g) were boiled for one hour with 9 5 % ethanol ( 1 0 0 mL) and the extract was filtered and evaporated to dryness. Ten microliters ( 1 0 uL) of the diluted extract ( 0 . 9 g in 5 mL) was spotted on TLC plate.
Adsorbent
: Silica g e l - G F 5 4
Solvent system
: Chloroform : Methanol ( 4 . 9 : 0 . 1 )
2
Detection Direct valuation
:
Scanning
: Densitometer at 2 5 4 nm (before spraying) and 4 5 0 nm (after
UV
2 5
4nm,
K t
values - 0
. 1 9 , 0 . 4 8 , 0 . 6 7 , 0 . 8 0 ,
0 . 9 6
spraying) Spray reagent
: Vanillin sulphate
Fig-13. TLC finger print profile of ethanol extract of Plectranthus zeylanicus aerial parts
11
Table 4. Description of densitogram (Fig-14)
-it .
1 O" Stage
Y(mnO
Peak no.
Y(ram)
1
10.06
Relative area % 38.67
2
23.73
10.35
3
30.00
4.24
4
71.49
25.12
Fig-14. Densitogram of TLC finger print profile of ethanol extracl of Plectranthus zeylanicus aerial parts at 254 nm
Table 5. Description of densitogram (Fig-15) Peak no.
Y (mm)
1
10.29
Relative area % 16.21
2
23.44
7.93
3
36.04
5.28
4
55.59
3.73
5
63.98
3.39
6
71.58
9.21
2.
• .". . • ;i i 3 -
4
.v/
5 ! '• • (i
jt.iijc V : inn
Fig-15. Densitogram of TLC finger prim profile of ethanol extract of Plectranthus zeylanicus aerial parts at 450 nm
12
Plectranthus zeylanicus Sample preparation
w a t e r
e x t r a c t
o f
r o o t s
: P. zeylanicus roots (4 g) were boiled for one hour with water (100 mL) and the extract was filtered and evaporated to dryness. Nine microliters (9 uL) of the diluted extract (0.9 g in 5 ml.) was spotted on TLC plate.
Adsorbent
: Silica gel-GFISJ
Solvent system
Detection
: Ethyl acetate : Acetic acid : water (4.6 : 0.2 : 0.2)
Direct evaluation Scanning : Spray reagent
UV
2
5
4nm,
Rf values - 0.32, 0.47. 0.96
: Densitometer at 254 nm (before spraying) and 450 nm (after spraying) : Vanillin sulphate
Fig-16. TLC finger print profile of water extract of Plectranthus zeylanicus roots
iI
Table 6. Description of densitogram (Fig-17)
C O
2 0 . 0
A 0. C
60.O
Peak no.
Y(mm)
Relative area %
1
8.41
35.70
2
47.41
21.01
3
69.78
34.67
Fig-17. Densitogram of TLC finger print profile of ethanol extract of Plectranthus zeylanicus roots at 254 nm
Table 7. Description of densitogram (Fig-18)
0
. 'j
0y
Peak no.
Y (mm)
Relative area %
1
9.60
21.18
2
16.93
8.76
3
25.69
11.22
4
66.89
45.14
'
:o . o2n
. c J r>. o .">
a
. o 6
v>.
o" 0 . &
Fig-18. Densitogram of TLC finger print profile of ethanol extract of Plectranthus zeylanicus roots at 450 nm
14
Plectranthus
zeylanicus ethanol extract of roots
Sample preparation
: P. zeylanicus
roots (4 g) were boiled for one hour with 9 5 %
ethanol (100 mL) and the extract was filtered and evaporated to dryness. Eight microliters (8 u.L) of the diluted extract (0.8 g in 5 mL) was spotted on TLC plate. Adsorbent
: Silica
Solvent system
rChloroform : Methanol (4.9 : 0.1)
gel-GF254
Detection Detection evaluation
: UV
Scanning
: Densitometer at 254 nm (before spraying) and 450 nm (after
25
4nm,
Revalues - 0.37, 0.49, 0.84
spraying) Spray reagent
: Vanillin sulphate
Fig-19. TLC finger print profile of ethanol extract of Plectranthus zeylanicus roots
15
Table 8. Description of densitogram (Fig-20)
' , 2
Y(mm)
Relative area %
1
8.31
56.25
2
23.20
7.50
3
30.01
9.81
'
11,
0 .000
3
Peak no.
I
r-....v^..
S Liige Y ijrmi i
Fig-20. Densitogram of TLC finger print profile of ethanol extract of Plectranthus zeylanicus roots at 254 nm
Table 9. Description of densitogram (Fig-21)
Peak no.
Y (mm)
Relative area %
1
8.42
23.80
?
22.74
37.00
3
49.09
16.07
4
68.49
23.13
i. -31-:,; Y ( :-,-n
Fig-21. Densitogram of TLC finger print profile of ethanol extract a(~ Plectranthus zevlanicus roots at 450 nm
16
High Pressure Liquid Chromatographic Profile** Plectranthus zeylanicus Sample preparation
water extract of aerial parts : P. zeylanicus aerial parts (4 g) were boiled for one hour with water (100 mL) and the extract wasfilteredand evaporated to dryness. The diluted extract (5.2 mg in 5 mL) was purified using Sep-pak CI8 cartridge.
Injection volume
: 20 ixh
Apparatus
: Shimadzu LC - 10 ADvp pumps and Shimadzu SPD - M 10 Avp uv / vis photodiode array detector.
Column
: Inertsil 5U ODS - 2 reverse phase column, (250 m m x 2.6 mm).
Solvent system
: Acetonitrile : Water (50 : 50)
Flow rate
: 1 mL/min
Detection
: 254 nm
Table 10. Retention times of main peaks
Peak no.
Retention
Relative
time
area %
(min) 1
2.32
22.64
2
2.46
16.94
3
2.82
30.00
4
5.08
8.74
5
6.22
21.49
Fig - 22. HPLCfingerprint profile of water extract of Plectranthus zeylanicus aerial parts
17
Plectranthus zeylanicus Sample preparation
ethanol extract of aerial parts
: P. zeylanicus aerial parts (4 g) were boiled for one hour with 95% ethanol (100 mL) and the extract was filtered and evaporated to dryness. The diluted extract (7.9 mg in 5 mL) was purified using Sep-pak CI8 cartridge.
Injection volume
: 20 /xL
Apparatus
; Shimadzu LC - 10 ADvp pumps and Shimadzu SPD - M 10 Avp uv / vis photodiode array detector.
Column
: Inertsil 5U ODS - 2 reverse phase column, (250 m m x 2.6 mm).
Solvent system
: Methanol : Water (85 : 15)
Flow rate
: 0.8 mL/min
Detection
: 254 nm
Table 11. Retention times of main peaks
0
5
1