Phytopharmacology of Holy basil (Ocimum tenuiflorum, Ocimum sanctum) Part 3
Phytopharmacology
of Holy basil
(Ocimum
tenuiflorum, Ocimum sanctum) Part 3
Phytochemistry
The chemical composition of Tulsi-Basil
(Ocimum sanctum) is highly complex.
The amount of biologically active compounds and nutrients may vary considerably
between strains and even among plants within the same field. The effect of the
way of growing, harvesting, processing and storage on the quantity of chemical
constituents have not yet well understood.
Phytochemicals found in Tulsi-Hloy Basil
(Ocimum sanctum) isolated by various
researchers are summarized here:
1. Eugenol, rosmarinic acid, apigenin,
myretenal, luteolin, Ī²-sitosterol, carnosic acid, flavonoids, orintin, vicenin.
[50]
2. From fresh leaves and stem of
Tulsi-Basil (Ocimum sanctum) phenolic
compounds such as cirsilineol, circimaritin, isothymusin, apigenin, rosameric
acid. The leaves of Tulsi-Basil (Ocimum
sanctum) contain 0.7 % volatile oil comprising of eugenol and methyl
eugenol. The oil also contains carvacrol and sesquiterpine hydrocarbon
caryophyllene. Recently two flavonoids orientin and vicenin have been isolated
from leaves of Tulsi-Basil (Ocimum
sanctum) [51], [52]
3. The volatile oil of leaf of Tulsi-Basil (Ocimum sanctum) eugenol (as it resembles
phenols it is called eugenic acid), ursolic acid, carvacrol, methyl carvicol
(also called Estragol), linalool, limatrol, caryophyllene, sugars and
anthocyans (anthocyanins: water-soluble pigments, that depending on their pH
may appear red, purple, blue or black)
In addition phenols: rosmarinic acid,
propanoic acid, apigenin, cirsimaritin, isothymucin, isothymonin and
water-soluble flavonoids: orientin and vicenin have been isolated.
The stem
and leaves contain a variety of
biologically active compounds of groups: phenols, organic acids, saponins,
flavonoids, triterpenoids and tannins.
The volatile oil of seeds have fatty acids, sitosterol, mucilage, some sugars (xylose)
and polysaccharides. [53]
4. Chemical constituents of Tulsi-Basil
(Ocimum sanctum) in different
solvents were found to be different. Here I summarize results of various
researchers.
a) Hexane (H): 1,2-dimethoxy-4-(2-propyl)benzene, 2-pentanone,
caryophyllene oxide, acetic anhydride, tricycloundecan, aromadendrene oxide and
benzodioxide
b) Ethyl acetate (EA): 1, 2-dimethoxy-4-(2-propenyl)benzene,
2-pentanone, 4-methyl-2-pentyl acetate, aromadendrene oxide, carbophyllene
oxide, benzenedicarboxylic acid and 10-heneicosene
c) Chloroform: 1, 2- dimethoxy-4-(2-propenyl) benzene,
4-methyl-2-phenyl acetate, ledene oxide, longipinocarvone, phtalic acid, butyl
hexyl ester, 2-pentanone, eugenol, 1.2-benzene dicarboxylic acid and
2-4-(lpropyl)-(E)-phenol
There are many essential
oil constituents in Tulsi-Basil
(Ocimum sanctum) such as camphor,
vanillin, naphthalene, ledene oxide, caryophyllene and eugenol. But when
extracted with different solvents the extract contains only borneol, vanillin,
naphthalene and eugenol. [54]
Seeds
In addition to mucilage, some sugars such as xylose
and polysaccharides seeds of Tulsi-Basil (Ocimum sanctum) contain ‘drying’oil with
physico-chemical properties similar to those of linseed oil. Other unsaturated
fatty acids include Ī±-linoleinic acid, linoleic acid and oleic acid. The most
abundant saturated fatty acids are palmitic acid and stearic acid.
The composition of Tulsi-Basil (Ocimum sanctum) seed oil suggests that
the oil would be suitable for industrial purposes, much in the same way as
linseed oil is used. [55]
Nutrients
and mineral contents (Fresh leaves) Per 100 G
Protein 4.2 g, Carbohydrate 2.3 g, Fat 0.5g, Omega 3
fatty acids 3%
Vitamin A 3%, Vitamin K 13%, Vitamin C 5%, Folate 4%
Calcium 4-5%, Iron 4-5%, Manganese 1.5-3 %, Copper 9%
[56]
Inorganic elements reported by other researchers are:
Sodium 0.74%, Potassium 6.62%, Sulphur 1.55%, Phosphorus 1.10% and Nitrogen
3.30%
Recent advances
on chemical composition of Tulsi-Holy Basil (Ocimum sanctum)
Leaves
The leaves contain high amount of phytochemicals which
include: Toluene, Camphene, Octane, Benzene, Citronellal, Sabinene, Limocene,
Ledol, Dimethylbenzene, ethyl-2-methylbuteryrate, Eugenol, Iso-eugenol,
Terpiniolene, Ī²-elemene, Iso-caryphyllene, Ī±-amorphene, Ī±-amorphene, Ī±-guaiene,
Ī±-humulene, humulene oxide, 14-hydroxy-Ī±-humulene, Ī±-terpenol, borneol,
Calamine, Nerolidol, Carvacrol, Geraneol, Elemol, Tetradecanol, (EZ)-famesol, Cissesquisainenehydrate,
Ī±-bisbolol, Selin-11-en-4-Ī±-ol, Ī±-murolene
The extraction of fresh leaves and stem yielded
phenolic compounds such as Apigenin, Circimaritin, Isothymusin, Eugenol and
Rosameric acid.
Tulsi-Basil (Ocimum sanctum) is also a good source of monoterpenes and
sesquiterpenes like Neral, Camphene, Cholesterol and Stigmasterol along with
Vitamin A, Vitamin C and Vitamin K [57]
Identity,
Purity and strength as per international guidelines
Foreign
matter: Not more than 2 percent
Total Ash: Not more than 10 percent
Acid-insoluble
Ash: Not more than 1.5 percent
Sulfated
ash: Not more than 20 percent
Alcohol-soluble
extractive: Not more than 4 percent
Water
soluble extractive: Not less than 8
percent
Loss on
drying: Not more than 14 percent [58], [59]
Heavy
Metal Analysis as per international guidelines
Element Permissible Limits
Arsenic Not more than 5 to 10 mg/kg
Cadmium Not more than 0.03mg/kg
Lead Not more than 5 to 10 mg/kg
Mercury Not more than 0.5 mg/kg
Chromium Not more than 0.3 mg/kg [60], [61]
Permissible
Microbial Load as per international guidelines
Microbial Limits:
Total bacterial count: Not more
than 105cfu/g
Total yeast and mould count: Not more
than 104cfu/g
Bile tolerant gram negative bacteria: Not more
than 104cfu/g [62], [63]
Specific Pathogens: (as
per international guidelines)
Salmonella species:
Absent in 25 g /none
Escherichia coli:
Absent in 1g / maximum 102 to 104
per gram
Staphylococcus aureus: Absent
in 1g
Pseudomonas aeruginosa: Absent
in 1g
Shigella species: Absent in 1g
Enterobacter
species: maximum 104
per gram
Other
enterobacteria: maximum 103
per gram
Aerobic bacteria: maximum 105 to 107
per gram
Mould propagules: maximum 103 to 105 per
gram
Yeasts and Mould: maximum 103 to 104
per gram [64], [65]
Aflatoxins (as per international guidelines)
Aflatoxin B1, Aflatoxin
B2, Aflatoxin G1, Aflatoxin G2
Preferably Aflatoxins
should be below detectable limits (BLD) [66], [67]
Pesticide residues (as per
international guidelines)
In recent times various pesticides are used to protect and preserve the
food and medicinal values of plants.
To avoid toxicity of herbal medicine, International Society for
Standardization of Drugs and World Health Organization (WHO) have laid the
guidelines for permissible levels of pesticides in herbal medicines.
In general, the pesticide contamination in any herbal medicine should
be less than 1 percent of total intake from all sources, including food
and drinking water.
Aldrin and dieldrin are broad spectrum pesticides
commonly used in agriculture. The recommended maximum limit of these pesticides
is Not more than 0.05 mg/kg. [68],
[69]
Radioactive residues (as per
international guidelines)
A certain amount of exposure to ionizing radiation of plants cannot be
avoided since there are many sources, including radionuclides occurring
naturally in the ground and the atmosphere.
The World Health Organization (WHO), in close collaboration with
several international organizations, has developed guidelines for permissible
and acceptable limits for radioactive residues in herbal medicines.
The amount of radiation in plants depends on intake of radionuclides.
Significant risk is associated only with consumption of quantities over 20 kg
of plant material per year so that the risk to health is most unlikely to be
encountered given the amount of medicinal plant materials need to be ingested.
Additionally, the level of contamination might be reduced during the
manufacturing process. Therefore World Health Organization (WHO) has not
proposed strict limits regarding the acceptability for radioactive
contamination. [70]
Genetic
Identity/ DNA sequencing
The genus Ocimum is highly variable. It includes
countless species. Further the genus
possesses wide genetic diversity at intra-species and inter-species levels. It
is very difficult to identify species on the basis of morphology of the leaf
alone. The recent technique of genome sequencing has offered a helping hand in
identifying each species accurately. [71]
Gene sequencing will
now facilitate identification of not yet identified genes involved in the
synthesis of important secondary metabolites in Tulsi-Basil
(Ocimum sanctum).
According
to the scientists, the development of molecular tools and genomic resources
will accelerate molecular breeding and utility of Tulsi-Basil (Ocimum sanctum) for medicinal purpose.
[72]
Tulsi-Basil
(Ocimum sanctum) is reported to have
a karyotype of 2n=36, which is the lowest among members of Ocimum genus. [73]
PCR
Analysis and SSR Sequencing
Recently the expression
of six important genes identified from genome sequencing data were validated by
scientists by performing q-RT-PCR in different tissues of five different
species, which shows the high content of urosolic acid-producing genes in young
leaves of Rama Tulsi (Ocimum sanctum)
subtype. In addition, the presence of eugenol and ursolic acid, implied as
potential drugs in the cure of many diseases. These findings were confirmed by
using mass spectrometry.
Expression of
anthocyanin biosynthesis-related genes in leaf samples of Krishna Tulsi (Ocimum tenuiflorum) were found to be
high explaining the purple coloration of Krishna Tulsi (Ocimum tenuiflorum). [74]
By suing SSR method,
efforts are now made by researchers towards the development of molecular
markers in order to study genetic diversity of Ocimum species. The data generated from SSR study would be helpful
in providing insights to the plant breeders and geneticists for evaluation of
desired genotypes with varied essential oil compositions and also development
of new Ocimum species. These efforts
will fling open the door of goldmine of synthesis of new drugs.
RAPD
analysis
RAPD (Random
Amplification of Polymorphic DNA) analysis has helped scientists to identify Ocimum species more accurately. [75]
AFLP
Analysis
AFLP (Amplified
Fragment Length Polymorphism) analysis also helps to identify Ocimum species accurately. The
advantages of AFLP (Amplified Fragment Length Polymorphism) over other genetic
markers are: high reproducibility, resolution, sensitivity at whole genome
level and no need of prior sequence information for amplification. AFLP (Amplified
Fragment Length Polymorphism) also has capability to amplify between 50 and 100
fragments at a time. [76]
HPTLC
analysis
By HPTLC analysis
quantitative estimation of various phytochemicals such as eugenol, ursolic
acid, rosmarinic acid and many others in Tulsi-Basil (Ocimum sanctum) has been done.
Chromosomal
Identity
Ocimum
species
are characterized by the different basic chromosome numbers
X=8, 10 12 or 16. In
case of Ocimum tenuiflorim individuals
chromosome numbers reported are 2n= 32, 2n=36 and 2n=76, however
internationally accepted chromosome number of Ocimum tenuiflorim is 2n=36.
As of today
internationally accepted number of chromosomes in other Ocimum species are:
Ocimum
gratissimum:
2n= 40
Ocimum
basilicum:
2n=48
Ocimum
americanum: 2n=72
Lemon basil: 2n=52 [77]
Ocimum canum Sims (hoary basil): 2n=26
Ocimum
kilimandscharicum: 2n=76 [78]
Ocimum viride Willd:
2n=40
Ocimum suave Willd:
2n=48
Vij and Kashyap (1976) have counted chromosome number
2=64 for Ocimum americanum collected
from North India, while Singh et al (1980) reported chromosome count of 2n=24,
26, 72 and 84 in Ocimum americanum L.
Morton (1962) has observed chromosome number 2n= 72
for Ocimum canum collected from North
India.
Pushpangdan and Sobti (1982) have reported chromosome
number 2n=24 for Ocimum canum introduced
from Kenya and 2n=26 for Ocimum canum
growing in South India. Mukherjee and
Datta (2006) also reported chromosome number 2n=26 for Ocimum canum.
Pushpangdan and Sobti (1982) have reported chromosome
number 2n=48 for Ocimum basilicum
growing in West India while Paton and Putievsky (1996) have counted 2n= 64, 72,
74 and 76 for Ocimum basilicum. For Ocimum basilicum var. crispum Mukherjee
et al (2005) reported a chromosomal count 2n=52 and Eder and Aikpokpodion 2n=48
and 60. [79]
References
50. Baliga M. S. et al, Ocimum sanctum L
(Holy Basil or Tulsi) and its phytochemicals in the prevention and treatment of
cancer, Nutr Cancer, 2013; 65 Suppl 1:26-35
51. Uma Devi P. et al, Radioprotective,
anticarcinogenic and antioxidant properties of the Indian holy basil, Ocimum
sanctum (Tulsi). Indian J Exp Biol. 2001; 39:185-190
52.
Gupta et al, Validation of traditional claim of Tulsi, Ocimum sanctum Linn. as
a medicinal plant. Indian J Exp Biol. 2002; 5:765-773
53. Priyabrata Pattanayak et al, Ocimum sanctum Linn. A reservoir plant
for therapeutic applications: An overview. Pharmacogn Rev. 2010 Jan-Jun; 4(7):
95-105
54. N. Dev et al, Chemical Compositions of
different Extracts of Ocimum basilicum, Journal of Scientific Research, 3(1),
197-206 (2011)
55. Paul Angers et al, Basil seed oils
https://
hort.purdue.edu/newcrop/proceedings1996/V3_598.html
56. Priyanka Pattanayak et al, Ocimum sanctum Linn. A reservoir plant
for therapeutic applications: An overview Pharmacogn Rev 2010 Jan; 4(7): 95-105
57. Bano N et al, Pharmcological Evaluation
of Ocimum sanctum, Journal of bioquivanance & Bioavailability, Volume 9(3):
387-392 (2017)
58. Quality control methods for
medicinal plant materials. Geneva, World
Health Organization, 1998.
60. Quality control methods for
medicinal plant materials. Geneva, World
Health Organization, 1998.
62. Quality control methods for
medicinal plant materials. Geneva, World
Health Organization, 1998.
64. Quality control methods for
medicinal plant materials. Geneva, World Health
Organization, 1998.
66. Quality control methods for
medicinal plant materials. Geneva, World
Health Organization, 1998.
68. Guidelines for predicting dietary
intake of pesticide residues, 2 nd review edition, Geneva, World Health
Organization, 1997
69. European pharmacopoeia, 3rd
edition Strasbourg, Council of Europe, 1996
70. Quality control methods for medicinal
plant materials, Geneva, World Health Organization, 1998
71.
Atul K. Upadhyay et al, Genome sequencing of herb Tulsi (Ocimum tenuiflorum) unravels key genes behind its strong medicinal
properties, BMC Plant Biol. 2015; 15:212
Scientists-sequence-tulasi-genome.html
73.
Felix Bast et al, Chloroplast DNAPhylogeography of Holy Basil (Ocimum
tenuiflorum) in Indian Subcontinent, The Scientific World Journal, Volume 2014,
Article ID 847482
74.
Atul K. Upadhyay et al, Genome sequencing of herb Tulsi (Ocimum tenuiflorum) unravels key genes behind its strong medicinal
properties, BMC Plant Biol. 2015; 15:212
75.
Carovic-Stanko et al. Genetic relations among basil taxa (Ocimum L.) based on molecular markers, nuclear DNA content and
chromosome number, Plant Syst. Evol. 285, 13-22. IJPPR, Volume 8, Issue 2:
February 2016
76.
Carovic-Stanko et al. Genetic relations among basil taxa (Ocimum L.) based on molecular markers, nuclear DNA content and
chromosome number, Plant Syst. Evol. 285, 13-22. IJPPR, Volume 8, Issue 2: February
2016
IJPPR,
Volume 8, Issue 2: February 2016
78. Moumita Mukherjee and Animesh K. Datta,
Secondary Chromosome Association in Ocimum spp., Cytologia 71(2):149-152,
2006
79. Johnson Ademola Idowu and Matthew
Oziegbe, Mitotic and Meiotic Studies on Two Species of Ocimum (Lamiaceae) and
their F1 Hybrids, Botanica Lithuanica 2017, 23(1): 59-67
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