ArticlePDF Available

Effect of extraction conditions on total polyphenol contents, antioxidant and antimicrobial activities of Cannabis sativa L

Authors:

Abstract

Cannabis sativa L. (hemp) was extracted for 2, 8 and 18 h with absolute acetone, methanol and their 50% aqueous solutions. The extracts were screened for total polyphenol contents, antioxidant and antibacterial activities. The polyphenol content of the extracts was found to be in the range of 0.090-0.556 mg gallic acid equivalents (GAE)/g dry weight cannabis, depending on the solvent used and the length of the extraction process. In general, methanol extract displayed the highest polyphenol contents and antioxidant activity, while absolute acetone was the least efficient solvent. Antioxidant activities of C. sativa extracts tested using the reducing power method ranged from 0.202 to 0.866 and depending on the extraction conditions; and the antioxidant activities correlated well with the polyphenol concentrations. The extracts also possessed antibacterial activity, depending on the solvent used and microbial species tested. Staphylococcus aureus and Candida albicans were found to be the most sensitive in the antimicrobial assay. Cannabis sativa extracts were not effective against Aspergillius niger.
!!
ISSN:!1579*4377!
EFFECT OF EXTRACTION CONDITIONS ON TOTAL
POLYPHENOL CONTENTS, ANTIOXIDANT AND ANTIMICROBIAL
ACTIVITIES OF CANNABIS SATIVA L.
Victor N. Mkpenie1, Emmanuel E. Essien1* and Idongesit I. Udoh2
1Department of Chemistry, University of Uyo, Uyo, Nigeria.
2Department of Science Technology, Akwa Ibom Polytechnic, Ikot Osurua, Ikot Ekpene, Akwa Ibom
State.
emmaflowus1@yahoo.co.uk
ABSTRACT
Cannabis sativa L. (hemp) was extracted for 2, 8 and 18 h with absolute acetone, methanol
and their 50% aqueous solutions. The extracts were screened for total polyphenol contents,
antioxidant and antibacterial activities. The polyphenol content of the extracts was found to
be in the range of 0.090-0.556 mg gallic acid equivalents (GAE)/g dry weight cannabis,
depending on the solvent used and the length of the extraction process. In general, methanol
extract displayed the highest polyphenol contents and antioxidant activity, while absolute
acetone was the least efficient solvent. Antioxidant activities of C. sativa extracts tested
using the reducing power method ranged from 0.202 to 0.866 and depending on the
extraction conditions; and the antioxidant activities correlated well with the polyphenol
concentrations. The extracts also possessed antibacterial activity, depending on the solvent
used and microbial species tested. Staphylococcus aureus and Candida albicans were found
to be the most sensitive in the antimicrobial assay. Cannabis sativa extracts were not
effective against Aspergillius niger.
KEYWORDS
Cannabis sativa, Cannabaceae, extraction, solvent, antioxidant, polyphenol, antimicrobial
activity.
Essien, E.E. et al. EJEAFChe, 11 (4), 2012. [300-307]
!
301!
INTRODUCTION
Cannabis sativa L. (Hemp) is an annual plant in the Cannabaceae family. The flowers (and
to a lesser extent the leaves, stems, and seeds) contain psychoactive and physiologically
active chemical compounds known as cannabinoids that are consumed for recreational,
medicinal, and spiritual purposes [1]. The plant and its preparations have been used for its
sedative, narcotic, antispasmodic, analgesic and many other properties including its use for
photophabia, migrain, asthma and piles. Cannabis also induces an increase in heart rate,
lowers blood pressure due to vasodilatation, stimulates appetite, and causes dry mouth and
dizziness [2]. Free radicals may cause reversible or irreversible damages to biological
molecules such as DNA, proteins and/or lipids [3]. These damages may cause cancer, heart
diseases and arthritis, and could accelerate aging of organisms [4]. Factors influencing the
antioxidant activity of phenolic compounds include position and number of hydroxyl groups,
polarity, solubility and stability of phenolic compounds during processing [5]. The
antioxidant action of polyphenol compounds depends on their free radical scavenging
capacity and iron reducing ability [6]. Extracts of Cannabis grown in different parts of the
world have been shown to possess antibacterial [2, 7, 8] and antioxidant activity [9]. The
antibacterial inhibitory of constituents [10] and antioxidant compounds [11, 12] of C. sativa
have been studied. Also, the effect of extraction conditions on measured phenolics content in
olive oil [13], teas [14], Orthosiphon staminens leaf [15], red and black currant [16] and
seabuckthorn seeds [17] have been reported. The total polyphenol amounts determined from
the same plant and their corresponding antioxidant and antimicrobial activities may vary
widely, depending on extraction conditions applied [18]. This study was designed to assess
the antioxidant activities of C. sativa leaf extracts and their antimicrobial activities against S.
aureus (Gram-positive bacteria), Pseudomonas aeruginosa (Gram-negative bacteria),
Candida albicans (yeast) and A. niger (mold); and to identify the effects of different
extraction solvents of varying polarities and the extraction times on these two properties and
the polyphenol contents of C. sativa. The antioxidant activity of C. sativa extracts was
determined using reducing power (RP) due to the fact that iron-polyphenol complex was
demonstrated to cause the inhibition of formation of oxygen radicals associated with many
pathological conditions [6] and also probably growth of some pathogen microorganisms
[19].
MATERIALS AND METHODS
PLANT SAMPLE AND CHEMICALS
Cannabis sativa was purchased from a flower garden at Ikot Obioema, Etim Ekpo Local
Government Area of Akwa Ibom State in July 2007. A taxonomist, Dr. (Mrs.) M. E. Bassey
of the Department of Botany and Ecological Studies, University of Uyo, Akwa Ibom State
authenticated the plant where a voucher specimen was deposited. The air- dried leaves were
pulverized and stored in sealed bags before extraction. The acetone and methanol were of
analytical grade from Aldrich, USA. Folin-Ciocalteu’s reagent and other chemicals were
products of Merck (Darmstadt, Germany).
Essien, E.E. et al. EJEAFChe, 11 (4), 2012. [300-307]
!
302!
EXTRACTION OF POLYPHENOLIC COMPOUNDS
The C. sativa leaves (1.0g) were extracted with methanol, acetone and their 50% aqueous
solutions (10 mL) on a horizontal shaker at room temperature for different times (2, 8, and
18 hr.). The samples were filtered through Whatman No.1 filter paper to remove rough
particles and then centrifuged (10 min, 10,000 × g). The supernatant was stored in a
refrigerator before analysis. Each solvent extraction was carried out in triplicate.
DETERMINATION OF TOTAL POLYPHENOLS
The amount of total polyphenol was determined using the Folin-Ciocalteu method [20]. A
gallic acid calibration curve ranging from 0.005 to 0.05 mg/mL was prepared and the results
determined from the regression equation of the calibration curve (y= 62.94x-0.67, R2=0.99)
were expressed as mg gallic acid equivalents (GAE)/g on dry weight basis. Cannabis
extracts (1 mL) diluted 10-75 times with de-ionized water (to obtain an absorbance in the
range of the prepared calibration curve) was mixed with 3-fold-diluted Folin-Ciocalteu
phenol reagent (1 mL). Sodium carbonate solution (2 mL, 35%) was added to the mixture,
which was shaken thoroughly and diluted to 6 mL by adding water (2 mL). The mixture was
allowed to stand for 30 min and the blue color formed was measured at 700 nm using a
double beam spectrophotometer (Unicam 8625).
ANTIOXIDANT ACTIVITY DETERMINATION BY REDUCING POWER
The reducing power (RP) of C. sativa extracts was determined according to the Yuan et al.
[21] with slight modifications. Cannabis sativa extract (0.5 mL) was mixed with phosphate
buffer (1.25 mL, 0.2 M, pH 6.6) and potassium ferric cyanide (1.25 mL, 1%) and the
mixture was incubated at 50 °C in a water bath for 20 minutes. The sample was then cooled
and mixed with 10% trichloroacetic acid (1.25 mL). Afterwards, a sample aliquot (1.25 mL)
was mixed with distilled water (1.25 mL) and 0.1% ferric chloride (0.25 mL) and then left to
react at room temperature for 10 min. Sample absorbance was read at 700 nm. An increase
in the absorbance (A) of the reaction mixture indicated an increase in the reducing power.
ANTIMICROBIAL ACTIVITY
Clinical strains of S. aureus, P. aeruginosa, C. albicans and A. niger were used as test
microbes. The bacteria and fungi were grown in Tyriptic Soy Broth (Merck, Germany) at 37
ºC for 24 h and fungi for 5 days. Test microorganisms were obtained from the culture
collections of the Department of Pharmaceutical Microbiology, University of Uyo, Akwa
Ibom State, Nigeria. Antibacterial activity was determined by the disc diffusion method [22,
23]. A sterilized 5 mm diameter antibacterial and antifungal susceptibility blank disc (Oxoid,
Basinstoke, UK) was loaded with 200 µL (4 mg) of each leaf extract (20 mg/mL) and left to
dry in an open sterile Petri dish in a laminar air flow hood (Forma Scientific, Turkey). The
test bacteria and fungi were transferred onto a 9 cm diameter Petri dish containing nutrient
agar and Sabouraud dextrose agar respectively (Merck, Germany) using a sterile cotton swab
and spread over the whole surface of the medium as a thin film. The inhibition of bacterial
growth was evaluated by measuring the diameter of the transparent inhibition zone around
Essien, E.E. et al. EJEAFChe, 11 (4), 2012. [300-307]
!
303!
each disc. Control disc was loaded with the same solvents and dried using the same method
as the treated disc.
RESULTS AND DISCUSSION
POLYPHENOL CONTENT
The polyphenol contents of C. sativa extracts were examined and the results are presented in
Table 1. The different solvent extraction systems showed a wide range of polyphenol
concentrations from 0.09 to 0.556 (GAE)/g dry weight Cannabis, depending on the solvent
used and the extraction times. The neat solvent extracts contained higher amounts of
polyphenols, compared to their corresponding 50% solvent ones, after 2, 8 and 18 h of
extraction indicating that the C. sativa leaf polyphenols were moderately polar compounds.
Among the solvents tested, the highest levels of polyphenols after 2 h of extraction were
achieved using methanol, followed by acetone, 50% methanol and 50% acetone. The orders
of polyphenol contents in C. sativa extracts at 8 and 18 h were same as that observed at 2 h.
On the whole, the polyphenol contents increased with the duration of the extraction process.
The results revealed that the total polyphenol content was influenced by solvent polarity and
extraction time as was reported previously for almond hulls and pine dust [24]. Similar
observations consistent with our present findings were reported by Brenes et al. [13] in
which N,N-dimethylformamide (DMF) as an extraction solvent resulted in the complete
extraction of phenols from olive oils, compared to methanol; and in the study of various
concentrations of ethanol ranging from 15% to 96% for the extraction of phenolic
compounds such as rosmarinic and caffeic acids from herbs, ethanol concentrations between
30% and 60% were found to give the highest extraction yields for both compounds.
However, 50% aqueous solvent extracts of black and black mate tea have been reported to
contain high concentrations of polyphenols relative to their absolute solvent extracts [14].
This variation could be due to the different polyphenol composition of materials
investigated.
Parameter
Extr. Time (h)
Extraction solvent
Acetone
Methanol
>99%
50%
>99%
Total
polyphenols
2
8
18
0.290±0.006
0.378±0.009
0.416±0.08
0.144±0.004
0.174±0.004
0.307±0.01
0.202±0.002
0.526±0.027
0.890±0.000
Antioxidant
activity (PR)
2
8
18
0.224±0.002
0.494±0.003
0.795±0.007
0.205±0.002
0.293±0.006
0.371±0.035
0.572±0.007
0.658±0.003
0.866±0.015
Table 1: Effect of different extraction solvents and extraction times on total polyphenol content (mg GAE/g
Cannabis) and antioxidant activity of C. sativa leaf. Data are expressed as means ± SE of triplicate
experiments.
ANTIOXIDANT ACTIVITY
The reducing power (RP) of C. sativa extracts using solvents of different polarities at
different extraction times was found to range from 0.202 to 0.866 (Table 1). The absolute
solvent extracts gave higher RP values than the corresponding 50% aqueous solvent extracts.
At 2 h of extraction, RP values of C. sativa extracts were affected by the extraction solvent
Essien, E.E. et al. EJEAFChe, 11 (4), 2012. [300-307]
!
304!
in the following order: methanol > acetone > 50% methanol > 50% acetone. At 8 h and 18 h,
the order of RP values from high to low were same as that observed for 2 h and the RP
values increased with increase in extraction time for each extraction solvent. It is apparent
that the RP values correlate with the polyphenols content at all different extraction times and
with respect to the polarity of the extraction solvent. These findings are in congruent with
the results of Akowuah et al. [15] who reported that using different solvent extraction
systems showed a wide range in the concentrations of individual phenolic compounds in
Orthosiphon stamineus leaf and the effect of different extraction times on their antioxidant
activity depended on solvents used. The moderately high reducing powers (a function of
their iron binding) of the absolute C. sativa extracts compared to the 50% aqueous ones may
be attributed to their molecular structure. Khokhar and Apenten [25] showed that a large
number of OH groups enhanced Fe-binding efficiency in phenolic compounds. Satoh et al.
[26] also observed that there was a strong correlation between the RP and total phenolic
content of different tea extracts. However, Psarra et al. [27] found a poor correlation
between RP and total polyphenols of wines and high correlation for radical scavenging
activity. These variations were attributed to the different polyphenol composition of
materials investigated.
ANTIMICROBIAL ACTIVITY
In this study, only absolute solvent C. sativa extracts at 2, 8 and 18 h were assayed for
antimicrobial activity against clinical strains of S. aureus, P. aeruginosa, C. albicans and A.
niger. The 50% aqueous extracts did not display significant antibacterial and antifungal
activity. The inhibitory effect of the tested extracts of C. sativa on the growth of selected
pathogens is shown in Table 2. Extraction solvents used as controls had no inhibitory effects
on the test microbes. It was found that extracts obtained at 18 h exhibited the strongest
inhibitory properties against S. aureus, P. aeruginosa and C. albicans compared to extracts
recovered after 2 and 8 h. All the tested extracts were inactive against the growth of A. niger.
The observed antimicrobial activity is attributed to the polyphenol contents of the extracts
obtained using different solvent systems and extraction times. It is also obvious that the
Gram-positive bacteria were more sensitive to C. sativa extracts than the Gram-negative
specie. Higher resistance of Gram-negative bacteria is attributed to the presence of
lipopolysaccharides in their outer membranes [28, 29]. In previous studies conducted by
Sakanaka et al. [30], Jayaprakasha et al. [31] and Ozkan et al. [32], the antibacterial activity
of plant extracts was attributed to individual phenolic compounds. Differences in
antibacterial activities of various solvent extracts have been reported in previous studies with
grape seed [29] and seabuckthorn seed [17]. The polyphenols in C. sativa leaf extract have
shown potent antimicrobial activity on all test microorganisms in the assay except for A.
niger and further research on the phenolic composition of potent extracts is necessary for a
comprehensive assessment of individual phenolic compounds exhibiting the observed
antimicrobial activity.
Essien, E.E. et al. EJEAFChe, 11 (4), 2012. [300-307]
!
305!
Microorganism
Extr. Time
(h)
Diameter of inhibited zone (mm)
Acetone (>99%)
Methanol (>99%)
S. aureus
2
8
18
12.00±0.00
16.00±0.00
20.00±0.10
10.00±0.00
14.00±0.01
20.00±0.00
P. aeruginosa
2
8
18
10.00±0.00
10.00±0.00
18.00±0.00
11.00±0.01
11.00±0.00
18.00±0.01
C. albicans
2
8
18
20.00±0.00
25.00±0.01
35.00±0.00
25.00±0.01
37.00±0.00
40.00±0.00
A. niger
2
8
18
nz
nz
nz
nz
nz
nz
Table 2: Antimicrobial activity of C. sativa. Data are expressed as means ± SE of triplicate experiments. nz: No
inhibition zone detected.
CONCLUSION
The different extraction solvents (absolute methanol, acetone, and their 50% aqueous
solvents) and extraction times of 2, 8 and 18 h showed significant effect on the polyphenol
contents of C. sativa leaf extract. For each parameter, with increasing extraction time
different trends were observed, depending on the solvent used. The observed antioxidant
activity by RP method and antimicrobial activity were attributed to the total polyphenols
concentration of the extracts with respect to the extraction solvent and time. Thus, the
highest concentration of total polyphenols, antioxidant and antimicrobial activities were
obtained with absolute methanol with extraction time of 18 h.
!
ACKNOWLEDGEMENTS
The authors are grateful to Dr. (Mrs.) M. E. Bassey for the authentication of the plant sample (C.
sativa).
!
REFERENCES
1. H. G. Pars, R. J. Razdan, J. F. Howes. Potential therapeutic agents derived from the
cannabinoid nucleus. Adv. Drug. Res. 11, 72-75 (1977).
2. K. Wasim, I. U. Haq, M. Ashraf M. Antimicrobial studies of the leaf of Cannabis sativa L.
Pak. J. Pharm. Sc. 8, 29-38 (1995).
3. G. Goldberg. Plants: Diet and Health. The Report of a British Nutrition Foundation Task
Force, Blackwell Science, Oxford, U.K (2003).
4. E. Cadenas, K. J. A. Davies. Mitochondrial free radical generation, oxidative stress, and
aging. Free Radical Biol. Med. 29, 222230 (2000).
5. E. A. Decker. Antioxidant mechanism. In Food Lipids (C. C. Akoh, D.B. Min, eds.), pp.
397-421. Marcel Dekker, New York (1998).
6. M. Yoshino, K. Murakami. Interaction of iron with polyphenolic compounds: application
to antioxidant characterization. Anal. Biochem. 257, 40-44 (1998).
Essien, E.E. et al. EJEAFChe, 11 (4), 2012. [300-307]
!
306!
7. L. Nissen, A. Zatta, I. Stefanini, S. Grandi, B. Sgorbati, B. Biavati, A. Monti.
Characterization and antimicrobial activity of essential oils of industrial hemp varieties
(Cannabis sativa L.). Fitoter. 81, 413-419 (2010).
8. J. Novak, K. Zitterl-Eglseer, S. G. Deans, C. M. Franz. Essential oils of different cultivars
of Cannabis sativa L. and their antimicrobial activity. Flav. Frag. J. 16, 259-262 (2001).
9. R. Abuzaytoun, F. Shahidi. Oxidative stability of flax and hemp oils. J. Am. Oil. Chem.
Soc. 83, 855861 (2006).
10. G. Appendino, S. Gibbons, A. Giana, A. Pagani, G. Grassi, M. Stavri, E. Smith, M. M.
Rahman. Antibacterial cannabinoids from Cannabis sativa: a structure-activity study. J. Nat.
Prod. 71, 1427-1430 (2008).
11. A.J. Hampson, M. Grimaldi, M. Lolic, D. Wink, R. Rosenthal, J. Axelrod.
Neuroprotective antioxidants from marijuana. Ann. New York Acad. Sci. 899, 274-282
(2000).
12. J. E. Koch. Delta 9-THC stimulates food intake in Lewis rats: effects on chow, high-fat
and sweet high fat diets. Pharmacol Biochem Behav. 68, 539543 (2001).
13. M. Brenes, A. Garcia, P. Garcia, A. Garrido. Rapid and complete extraction of phenols
from olive oil and determination by means of a coulometric electrode array system. J. Agric.
Food Chem. 48, 5178-83 (2000).
14.N. Turkmen , F. Sari, Y. S. Velioglu. Effects of extraction solvents on concentration and
antioxidant activity of black and black mate tea polyphenols determined by ferrous tartrate
and Folin-Ciocalteu methods. Food Chem. 99, 835-841 (2005).
15. G. A. Akowuah, Z. Ismail, I. Norhayati, A. Sadikun. The effects of different extraction
solvents of varying polarities on polyphenols of Orthosiphon stamineus and evaluation of the
free radical- scavenging activity. Food Chem. 93, 311-317 (2005).
16. B. Lapornik, M. Prosĕk, A. G. Wondra. Comparison of extracts prepared from plant by-
products using different solvents and extraction time. J. Food Eng. 71, 214-222 (2005).
17. P. S. Negi, A. S. Chauhan, G. A. Sadia, Y. S. Rohinishree, R.S. Ramteke. Antioxidant
and antibacterial activities of various seabuckthorn (Hippophae rhamnoides L.) seed extracts.
Food Chem. 92, 119-124 (2005).
18. A.Wach, K. Pyrzyńska, M. Biesaga. Quercetin content in some food and herbal samples.
Food Chem. 100, 699-704 (2007).
19. K. T. Chung, Z. Lu, M. W. Chou. Mechanism of inhibition of tannic acid and related
compounds on the growth of intestinal bacteria. Food Chem. Toxicol. 36, 1053-1060 (1998).
20. M. Obanda, P. O. Owuor. Flavanol composition and caffeine content of green leaf as
quality potential indicators of Kenyan black teas. J. Sci. Food Agric. 74, 209-215 (1997).
21. Y. V. Yuan, D. E. Bone, M. F. Carrington. Antioxidant activity of dulse (Palmira
palmate) extracts evaluated in vitro. Food Chem. 91, 485-494 (2005).
22. A.W. Bauer, W. M. M. Kirby, J. C. Sherries, M. Turck. Antibiotic susceptibility testing
by single 22. disk method. Am. J. Clin. Pathol. 45, 493496 (1966).
23. C. H. Collins, M. L. Patricia. Microbiological Methods, pp. 441-446. Oxford Press,
London (1994).
24. M. Pinelo, M. Rubilar, J. Sineiro, M. J. Nunez. Extraction of antioxidant phenolics from
almond hulls (Prunus amygdalus) and pine sawdust (Pinus pinaster). Food Chem. 85, 267-
273 (2004).
25. S. Khokhar, R. K. O. Apenten. Iron binding characteristics of phenolic compounds: some
tentative structure-activity relations. Food Chem. 81, 133-140 (2003).
26. E. Satoh, N. Tohyama, M. Nishimura. Comparison of the antioxidant activity of roasted
tea with green, oolong, and black teas. Int. J. Food Sci. Nutr. 56, 551-559 (2005).
27. E. Psarra, D. P. Makris, S. Kallithraka, P. Kefalas. Evaluation of the antiradical and
reducing properties of selected Greek white wines: correlation with polyphenolic
composition. J. Sci. Food Agric. 82, 1014-1020 (2002).
Essien, E.E. et al. EJEAFChe, 11 (4), 2012. [300-307]
!
307!
28. N. S. Alzoreky, K. Nakahara. Antibacterial activity of extracts from some edible plants
commonly consumed in Asia. Int. J. Food Microbiol. 80, 223-230 (2003).
29. N. G. Baydar, G. Ozkan, O. Sagdic. Total phenolic contents and antibacterial activities of
grape (Vitis vinifera L.) extracts. Food Control 15, 335-339 (2004).
30. S. Sakanaka, L. R. Juneja, M. Taniguchi. Antimicrobial effects of green tea polyphenols
on thermophilic spore-forming bacteria. J. Biosci. Bioeng. 90, 81-85 (2000).
31. G. K. Jayaprakasha, T. Selvi, K. K. Sakariah. Antibacterial and antioxidant activities of
grape (Vitis vinifera) seed extracts. Food Res. Int. 36, 117-122 (2003).
32. G. Ozkan, O. Sagdic, N. G. Baydar, Z. Karamahmutoglu. Antibacterial activities and
total phenolic contents of grape pomace extracts. J. Sci. Food Agric. 84, 807-1811 (2004).
... 47 This seems to ignore the bactericidal effect produced by cannabis extract in vitro against gram-positive microorganisms and mycobacterium. [48][49][50][51] The results of the study then recommended the use of cannabis extract for the treatment of festering wounds, swelling accompanied by pus, inflammation of the auditory canal, and sinusitis. 52 Based on data collected by Krejci 53 Regarding the use of cannabis in the health care sector, especially in the dental and oral department, cannabis extract has been shown to relieve herpes labialis, gum pocket pain caused by inflammation and cysts in dental tissue, gums covering wisdom teeth, inflammation of the alveolar bone, ulcers in the mouth or tongue (aphtha), and inflammation and ulcers of the gums. ...
... 47 57 Research on extracts and preparations of cannabis and cannabinoids is also ongoing. The study used extracts from the leaves of, 47,48,48,49,50,51 seeds, 52 flowers, 53 and all parts of the cannabis plant. 56 Furthermore, the type of preparation used is essential oil 54,55,56,57 and hashish. ...
... 56 Furthermore, the type of preparation used is essential oil 54,55,56,57 and hashish. 50 In addition, the constituents of the studied cannabinoids are cannabinol (CBN), cannabidiol (CBD), 58,59 cannabichromene (CBC), cannabigerol (CBG), 61 delta-9-tetrahydrocannabinol (Δ 9 -THC), 60,61 delta-8tetrahydrocannabinol (Δ 8 -THC). 62 Although not all research results show the potential of cannabis as an antimicrobial agent 63 and not all showed significant results against gram-negative bacteria, 50,64,65 These results are very likely influenced by the lack of cannabinoids contained in the extract and preparation of cannabis. ...
Article
Full-text available
Cannabis sativa, or widely known as ganja and other names in Indonesia, is a plant that could not be used in the health care system in Indonesia. It is because cannabis is still a Class 1 Narcotic. However, trends within the last few years show that cannabis has medicinal properties, and is already approved as medicine and (even) for recreational means in several nations/countries-Thailand is the first nation in Southeast Asia that did. This article aims to emphasize the antimicrobial potential that cannabis has. Since conventional antibiotic is considered unable to provide, it is important to find another source to counter the high rates of antibiotic (antimicrobial) resistance. Moreover, this article is also aimed to introduce one of the potentials that cannabis have to Indonesia since scientific article written in Bahasa is rare.
... Notably, the level of sensitivity of the extracts in qualitative tests is not equipollent since their polarity and solubility change their diffusivity through media [69,70]. However, in many other investigations, the activity of C. sativa was shown against P. aeruginosa [62,[71][72][73][74][75], E. coli [62,67,68,72,[76][77][78][79][80][81][82], Salmonella species [76,80,83,84], Shigella species [76,82], K. pneumoniae [82], Acinetobacter calcoaceticus [79], Morganella morganii [62] and Serratia marcescens [84]. ...
... C. albicans is part of a natural microflora that forms asymptomatic colonies on the skin and inside the body and can proliferate if the host has an immunosuppressed condition and cause superficial mucosal and dermal infections [120,121]. Activity against Candida species [67,73,74,105,107] Fusarium spp. [68], Candida neoformans [73] and Aspergillus [68,105,122] are documented. ...
... Bioactivity of C. sativa EOs and extracts depend on the concentration of active compounds, which is associated with many extrinsic factors such as geographical origin, sowing time, plant age, collection time, and soil composition, along with many intrinsic factors including genetic information, cultivars, accessions, maturity, and even the aging of the extract itself [60,79,83,204]. Appropriate selections of solvents, their concentration, extraction method and extraction parameters are crucial for biocidal property of respective extracts [74,205]. Apart from these, the choice of antimicrobial tests also produces variability in results [206,207]. ...
Article
Full-text available
Abstract: Antimicrobial resistance has emerged as a global health crisis and, therefore, new drug discovery is a paramount need. Cannabis sativa contains hundreds of chemical constituents produced by secondary metabolism, exerting outstanding antimicrobial, antiviral, and therapeutic properties. This paper comprehensively reviews the antimicrobial and antiviral (particularly against SARS-CoV-2) properties of C. sativa with the potential for new antibiotic drug and/or natural antimicrobial agents for industrial or agricultural use, and their therapeutic potential against the newly emerged coron-avirus disease (COVID-19). Cannabis compounds have good potential as drug candidates for new antibiotics, even for some of the WHO's current priority list of resistant pathogens. Recent studies revealed that cannabinoids seem to have stable conformations with the binding pocket of the M pro enzyme of SARS-CoV-2, which has a pivotal role in viral replication and transcription. They are found to be suppressive of viral entry and viral activation by downregulating the ACE2 receptor and TMPRSS2 enzymes in the host cellular system. The therapeutic potential of cannabinoids as anti-inflammatory compounds is hypothesized for the treatment of COVID-19. However, more systemic investigations are warranted to establish the best efficacy and their toxic effects, followed by preclinical trials on a large number of participants.
... Figure 2a-c show that India, Sudan, Nigeria, and Pakistan express the highest temperatures, rainfall, and humidity levels. In relationship, the highest antimicrobial growth occurred in hemp oils sourced from Sudan, Nigeria, and Pakistan [9,42,43]. Therefore, the rationale underpinning this objective is that if hemp plants experience optimal growing conditions of mild climates, humid atmosphere, and a rainfall of at least 635-762 mm per year, the plants will obtain high quantities of antimicrobial compounds. HSO cultivated in Niala, South Darfur, and Sudan possessed the highest activity against Gram-positive Staphylococcus aureus with 28 mm of ZGI. ...
... In addition to the high humidity of 93% to 100%, high rainfall of 40-324 mm daily, high soil fertility, and high content of CBD, the hemp leaf oil possessed the most increased antifungal activity up to 40 mm of growth inhibition against Candida albicans. The hemp plant that shows the highest activity against Gram-negative bacteria in Pakistan was similar to the hemp plant grown in Nigeria [42,43]. The hemp plant was raised in an environment with high temperatures and humidity of 27 °C and 80%, respectively (Figure 2a,c). ...
... The findings were due to the higher content of CBD in HLE compared to HEO. Both hemp plants grown in Pakistan and Nigeria were grown on garden soils and hemp plants in Sudan on woodland savanna/semi-arid soils, while Indian hemp was found on brown forest [31,42,43]. ...
Article
Full-text available
The bioactive Hemp Seed Oil (HSO) is becoming very popular in the medical and research fields due to its antimicrobial properties against several diseases caused by bacteria and fungi. However , the effect of hemp-growing factors and extraction methods on the bioactivity of HSO does not receive adequate research attention. Therefore, this review aims to investigate the effect of growth factors and extraction methods on the antimicrobial activity of HSO. Articles were retrieved from Google Scholar and the Scopus database and screened against inclusion and exclusion criteria. The study revealed that HSO prefers warm climates and favorable humidity ranging from 20 to 39 °C and 79-100% per year, respectively, and rainfall of 324 mm daily. The multivariate linear regression shown excellent prediction (R 2 = 0.94) with climates upon Zone of Growth Inhibition (ZGI) of Gram-positive bacteria. Temperature is the strongest predictor (p < 0.01) followed by humidity and rainfall (p < 0.05). Furthermore, well-drained loam soil rich in organic matter seems to stimulate the antimi-crobial activity of HSO. The major constituents that influence HSO's antimicrobial ability to Staphylococcus aureus were cannabidiol (CBD), β-caryophyllene, and limonene. The extraction methods showed less influence on the HSO bioactivity. HSO did not show significant antioxidant activity, but Hemp Seed Hull (HSH), Hemp Seed Flour (HSF), and Hydrolyzed Hemp Seed Protein (HPH), expressed promising DPPH scavenging ability.
... Thus, studies essentially focused on determining the recovery potential of these molecules from cannabis by-products such as seed meals or leaves. The majority of the extractions were conducted at room temperature [48][49][50][51]53,54], though an increase in temperature leads to a higher PC recovery [54]. Only one study optimized the extraction temperature carried out with a new eco-friendly solvent, an aqueous solution of 2-hydroxypropyl-β-cyclodextrin (CD) [52]. ...
... Optimization studies to find the optimum operating conditions are lacking. Mkpenie et al. (2012) [51] investigated the influence of the extraction time during maceration at room temperature. They found that the recovery of PC increases by 60% between 2 and 8 h of extraction. ...
... Optimization studies to find the optimum operating conditions are lacking. Mkpenie et al. (2012) [51] investigated the influence of the extraction time during maceration at room temperature. They found that the recovery of PC increases by 60% between 2 and 8 h of extraction. ...
Article
Full-text available
Cannabis sativa L. is a controversial crop due to its high tetrahydrocannabinol content varieties; however, the hemp varieties get an increased interest. This paper describes (i) the main categories of phenolic compounds (flavonoids, stilbenoids and lignans) and terpenes (monoterpenes and sesquiterpenes) from C. sativa by-products and their biological activities and (ii) the main extraction techniques for their recovery. It includes not only common techniques such as conventional solvent extraction, and hydrodistillation, but also intensification and emerging techniques such as ultrasound-assisted extraction or supercritical CO2 extraction. The effect of the operating conditions on the yield and composition of these categories of phenolic compounds and terpenes was discussed. A thorough investigation of innovative extraction techniques is indeed crucial for the extraction of phenolic compounds and terpenes from cannabis toward a sustainable industrial valorization of the whole plant.
... The methanol extract, being the most polar, had the highest content of phenolics. In a study conducted by Mkpenie et al. (2012), the polyphenol content of the acetone and methanolic extracts of C. sativa was found to be in the range of 0.090 -0.556 mg GAE/g dry weight. We therefore attribute the higher content of total phenolics observed in the present study to the different extraction times; 2, 8 and 18 hours in the study by Mkpenie et al. (2012) compared to the extraction time of 48 hours in the current study. ...
... In a study conducted by Mkpenie et al. (2012), the polyphenol content of the acetone and methanolic extracts of C. sativa was found to be in the range of 0.090 -0.556 mg GAE/g dry weight. We therefore attribute the higher content of total phenolics observed in the present study to the different extraction times; 2, 8 and 18 hours in the study by Mkpenie et al. (2012) compared to the extraction time of 48 hours in the current study. ...
... In addition, several studies have quantified individual phenolic compounds, such as caffeic acid, gallic acid, rosmarinic acid, p-OH-benzoic acid, ferulic acid, 3,4-dihydroxybenzoic acid, p-coumaric acid, syringic acid, quercetin, luteolin, canniprene, cannflavin A, cannflavin B, catechin, naringenin, isorhamnetin, resveratrol, rutin trihydrate, apigenin, and apigenin 7-glucoside in the extracts [95][96][97][98]. Of note, most conventional extraction was conducted at room temperature and not under optimal conditions [99][100][101][102][103]. Furthermore, an aqueous solution of 2-hydroxypropyl-β-cyclodextrin (a green solvent) has also been applied to recover phenolic compounds from the by-products of industrial hemp oil processing [103]. ...
Article
Full-text available
Cannabis is well-known for its numerous therapeutic activities, as demonstrated in pre-clinical and clinical studies primarily due to its bioactive compounds. The Cannabis industry is rapidly growing; therefore, product development and extraction methods have become crucial aspects of Cannabis research. The evaluation of the current extraction methods implemented in the Cannabis industry and scientific literature to produce consistent, reliable, and potent medicinal Cannabis extracts is prudent. Furthermore, these processes must be subjected to higher levels of scientific stringency, as Cannabis has been increasingly used for various ailments, and the Cannabis industry is receiving acceptance in different countries. We comprehensively analysed the current literature and drew a critical summary of the extraction methods implemented thus far to recover bioactive compounds from medicinal Cannabis. Moreover, this review outlines the major bioactive compounds in Cannabis, discusses critical factors affecting extraction yields, and proposes future considerations for the effective extraction of bioactive compounds from Cannabis. Overall, research on medicinal marijuana is limited, with most reports on the industrial hemp variety of Cannabis or pure isolates. We also propose the development of sustainable Cannabis extraction methods through the implementation of mathematical prediction models in future studies.
Chapter
Full-text available
Bal arıları meyve, sebze ve tohum oluşumu için çok önemli hayvanlardır. Arılar, çiçekli bitkilerin erkek yapılarındaki polenleri dişi kısımlarına aktararak bitkilerde meyve ve tohum oluşumunu sağlar. Ayrıca bal yapmaları nedeniyle de tarih boyunca bu böceklere çok önem verilmiştir. Biz insanlar her zaman bal arısına hayran olmuşuzdur. Afrika, Avrupa ve Asya’daki en eski atalarımız, yüz binlerce yıl boyunca, bu arının bal depolama ve balmumu yapma konusundaki şaşırtıcı endüstrisine, çok değerli iki maddeye kesinlikle hayran kaldılar. Daha yakın zamanlarda, son 10.000 yılda, karmaşık arıcılık zanaatını icat ettik ve bal arıları üzerine bilimsel çalışmalarımıza başladık. Örneğin, bu arının «çiçek değişmezliği» uygulamasını ilk kez tanımlayan antik filozof Aristotales’ti: Bir işçi arı, yiyecek toplamanın verimliliğini artırmak için yiyecek arama gezisi boyunca genellikle bir tür çiçeğe yapışır. Bal arısının doğal dünyasında nasıl yaşadığını bilmek, geniş bir bilimsel araştırma yelpazesi gerekmektedir. Bunun nedeni, Apis mellifera’nın biyolojideki, özellikle davranışla ilgili temel soruları araştırmak için model sistemlerden biri haline gelmesidir. Bu arıları ister hayvan bilişindeki, ister davranışsal genetikteki veya sosyal davranışlardaki bazı gizemleri çözmek için çalışıyor olun, birinin deneysel araştırmalarını tasarlamadan önce doğal biyolojilerine aşina olmak kritik derecede önemlidir.
Article
In this work, different extraction techniques (soxhlet extraction, hydrodistillation, subcritical water extraction and supercritical carbon dioxide extraction followed by conventional extraction) were employed for the isolation of bioactive compounds from the areal parts of industrial hemp (Cannabis sativa L.). The extraction process parameters, time and temperature for subcritical water extraction and pressure, temperature and time for supercritical carbon dioxide extraction, on the extraction yield and the content of bioactive compounds from hemp were examined. As the plant material after supercritical carbon dioxide still contains hydrophilic compounds, conventional extraction was used for isolation of these. The content of cannabidiol, the main cannabinoid present in hemp, in supercritical carbon dioxide extracts was between 71.84–163.11 mg/g, while in soxhlet extract it was much lower (64.40 mg/g). In comparison to these the significantly lower cannabidiol content was detected in subcritical water extracts, ranging from 0.0039 to 0.0183 mg/mL. Comparing all applied extraction techniques, supercritical carbon dioxide followed by conventional extraction was selected as the most valuable process for bioactive compounds isolation for hemp.
Article
Full-text available
Marijuana (Cannabis sativa) has long been known to contain antibacterial cannabinoids, whose potential to address antibiotic resistance has not yet been investigated. All five major cannabinoids (cannabidiol (1b), cannabichromene (2), cannabigerol (3b), Delta (9)-tetrahydrocannabinol (4b), and cannabinol (5)) showed potent activity against a variety of methicillin-resistant Staphylococcus aureus (MRSA) strains of current clinical relevance. Activity was remarkably tolerant to the nature of the prenyl moiety, to its relative position compared to the n-pentyl moiety (abnormal cannabinoids), and to carboxylation of the resorcinyl moiety (pre-cannabinoids). Conversely, methylation and acetylation of the phenolic hydroxyls, esterification of the carboxylic group of pre-cannabinoids, and introduction of a second prenyl moiety were all detrimental for antibacterial activity. Taken together, these observations suggest that the prenyl moiety of cannabinoids serves mainly as a modulator of lipid affinity for the olivetol core, a per se poorly active antibacterial pharmacophore, while their high potency definitely suggests a specific, but yet elusive, mechanism of activity.
Article
Leaf powder of Orthosiphon stamineus was extracted with the following solvents; distilled water, 50% aqueous methanol, methanol, 70% aqueous acetone and chloroform, at 2, 4 and 8 h, respectively, on a water bath at 40 °C. The extracts were subjected to qualitative and quantitative HPLC analyses of the polyphenols, the most dominant chemical constituents in the leaf. Chloroform extraction from 4 to 8 h at 40 °C gave the highest amount of sinensetin and eupatorin. The extraction with 70% aqueous acetone extracts at 4 and 8 h gave a high yield of 3′-hydroxy-5,6,7,4′-tetramethoxyflavone. The yield of rosmarinic acid was high in 50% methanol extracts at 2, 4 and 8 h of extraction. The extracts were screened for free radical-scavenging potential, using a 1,1-diphenyl-2-picrylhydrazyl in vitro model system. The extracts exhibited significant radical-scavenging activity and the acetone extracts showed the highest activity.
Article
Lignocellulosic residues, such as pine sawdust and almond hulls, were solvent-extracted under different experimental conditions to optimize the yield of polyphenolic antioxidant compounds, which were quantified. The antioxidant power of extracts was evaluated by ability to scavenge the 1,1-diphenyl-2-picryl-hydrazyl (DPPH) radical. Both materials were found to be important sources of phenolic antioxidants, although the efficiency of the extraction varied with the experimental conditions. Among the three solvents used (ethanol, methanol and water), ethanol was the most favourable for total extractables, although methanol was more selective for extracting polyphenolics. For these latter, pine sawdust offered the best results, with a 3–10 times higher (0.1122 g/100 g in dry basis) total phenolics content than almond hulls but, despite this, phenols from hulls showed a higher antioxidant capacity (58 vs 34% of inhibition).
Article
Quercetin is a typical flavonoid ubiquitously present in vegetables and fruits, and its antioxidant effect is implied to be helpful for human health. The efficiency of extraction process and acidic hydrolysis parameters for HPLC analysis of quercetin present in glycosides and aglycone forms was investigated. Hydrolysis for 5min in the presence of 2.8M HCl as well as for 10min with 1.1M HCl efficiently released quercetin from rutin. The method developed in this study was applied for quantitative determination of quercetin in some food (onion, apple) and herbal (Hypericum perforatum and Sambucus nigra) products.
Article
The essential oils of five different cultivars of Cannabis sativa contained as main compounds α-pinene, myrcene, trans-β-ocimene, α-terpinolene, trans-caryophyllene and α-humulene. The content of α-terpinolene divided the cultivars in two distinct groups, an Eastern European group of cultivars of approximately 8% and a French group of cultivars of around 16%. Therefore, this compound might be suitable as a genetic marker for the two breeding centres for the fibre types of Cannabis sativa. The content of trans-caryophyllene was up to 19%. However, the content of caryophyllene oxide did not exceed 2%. The antimicrobial activity of the essential oil of Cannabis sativa can be regarded as modest. Nevertheless, cultivar differences were visible. Δ-9-tetrahydrocannabinol (THC) could not be detected in any of the essential oils and the amount of other cannabinoids was very poor. Copyright © 2001 John Wiley & Sons, Ltd.
Chapter
Chocolate (see also Chapter 9, Section 9.5 on cocoa)Herbs, spices and condimentsResearch recommendationsKey points
Article
Cannabidiol and other cannabinoids were examined as neuroprotectants in rat cortical neuron cultures exposed to toxic levels of the neurotransmitter, glutamate. The psychotropic cannabinoid receptor agonist Δ9-tetrahydrocannabinol (THC) and cannabidiol, (a non-psychoactive constituent of marijuana), both reduced NMDA, AMPA and kainate receptor mediated neurotoxicities. Neuroprotection was not affected by cannabinoid receptor antagonist, indicating a (cannabinoid) receptor-independent mechanism of action. Glutamate toxicity can be reduced by antioxidants. Using cyclic voltametry and a fenton reaction based system, it was demonstrated that Cannabidiol, THC and other cannabinoids are potent antioxidants. As evidence that cannabinoids can act as an antioxidants in neuronal cultures, cannabidiol was demonstrated to reduce hydroperoxide toxicity in neurons. In a head to head trial of the abilities of various antioxidants to prevent glutamate toxicity, cannabidiol was superior to both a-tocopherol and ascorbate in protective capacity. Recent preliminary studies in a rat model of focal cerebral ischemia suggest that cannabidiol may be at least as effective in vivo as seen in these in vitro studies.
Article
The aim of this study was to determine the total phenolic contents and antibacterial effects of grape pomace extracts (cultivars Emir and Kalecik karasi) against 14 bacteria, and the effects of the extracts on the growth and survival of two of the bacteria during storage. The total phenolic contents of grape pomace of Emir and Kalecik karasi cultivars extracted with acetone/water/acetic acid (90:9.5:0.5) were 68.77 and 96.25 mg GAE g−1, respectively. The agar well diffusion method was used to test the antibacterial activity of the extracts at 1, 2.5, 5, 10 and 20% (w/v) concentrations in methanol on spoilage and pathogenic bacteria including Aeromonas hydrophila, Bacillus cereus, Enterobacter aerogenes, Enterococcus faecalis, Escherichia coli, Escherichia coli O157:H7. Mycobacterium smegmatis, Proteus vulgaris, Pseudomonas aeruginosa, Pseudomonas fluorescens, Salmonella enteritidis, Salmonella typhimurium, Staphylococcus aureus and Yersinia enterocolitica. All the bacteria tested were inhibited by extract concentrations of 2.5, 5, 10 and 20%, except for Y enterocolitica which was not inhibited by the 2.5% concentration. However, pomace extracts at 1% concentration had no antibacterial activity against some of the bacteria. According to the agar well diffusion method, E coli O157:H7 was the most sensitive of the bacteria. Generally, using the serial dilution method, while the extracts at 0.5% concentration had bacteriostatic activities on E coli O157:H7 and S aureus, the extracts appeared to have bactericidal effects at 1 and 2.5% concentrations. In accordance with this method, S aureus was more sensitive than E coli O157:H7 to the extracts. Copyright © 2004 Society of Chemical Industry