Extraction of Alkaloids from Colocynthis vulgaris (L.) Schrader harvested in Oued N’sa (Ouargla region)
Latra Benkhira1, Noura Chaouch2*
1Process Engineering Department, Faculty of Applied Sciences, Kasdi Merbah Ouargla University,
Ouargla 30000, Algeria.
2Process Engineering Laboratory, Process Engineering Department, Faculty of Applied Sciences,
Kasdi Merbah University, Ouargla 30000, Algeria.
*Corresponding Author E-mail: amirchaouch@gmail.com
ABSTRACT:
Plants constitute a vital component of the biodiversity as they play a key role in maintaining earth’s environmental equilibrium and ecosystem stability. Herbal medicine is known to be the oldest form of healing, dating back to ancient Greece (1600 BC). It involves the use of plant to improve, maintain or restore health and wholeness. Phytochemicals are bioactive chemicals of plant origin. They are regarded as secondary metabolites synthesized in all parts of the plant body. The quantity and quality of phytochemicals present in plant parts may differ from one part to another. In fact, there is a lack of information on the distribution of the biological activity in different plant parts mainly due to the difference in distribution of active compounds which are more frequent in some plant parts than in others. Colocynthis vulgaris (L) Schrader is one of the medicinal plants of the Cucurbitaceae family, widespread in Algeria mainly in Ouargla in the region of Oued N'sa. This plant is used in traditional medicine for the treatment of several diseases. Its multiple pharmacological properties are due to its richness in active substances, especially alkaloids. The main objective of this study is the comparison between two extraction protocols of alkaloids, one in acid medium and the other in an alkaline medium. The results obtained confirm the plant's richness in alkaloids and the efficiency of the extraction protocol in acid medium.
The use of plant in herbal medicine has a long history and is currently experiencing a resurgence of public interest. According to the World Health Organization, approximately 65-80 % of the world's population uses traditional herbal medicine to meet their primary health care needs3-5.
The role of medicinal plants in disease prevention or control has been attributed to bioactive properties of their constituents, usually associated to a wide range of molecules, including alkaloids, flavonoids, lipids, steroids, terpenoids, carotenoids, glycosides, tannins, resins and saponins. These compounds are commonly found in, and often responsible for actions of most medicinal plants6–8.
For several years, many phytotherapy products have been marketed, and in different forms: tablets, capsules, fluids in ampoules or vials, etc. This makes the choice difficult. The quality of the plant used: fresh plant, dry plant, organically grown or not, determines its content and richness in active ingredients. Moreover, to be optimally effective, a plant-based product must restore all the molecular complexity of the plant that is at the origin of its therapeutic activity. Particular attention must therefore be paid to the process used for the extraction of the active compounds9-11.
The growing interest in secondary metabolites of plants has directed attention to methods for their extraction. They are extracted by conventional methods such as Soxhlet12,13 and room temperature solvent extraction14,15 or by ultrasound16,17, microwaves18-20 and supercritical solvents21,22.
Alkaloids are compounds that contain nitrogen at a negative oxidation level23. They are of considerable importance in phytotherapy. Their physiological action is very varied, some act on the central nervous system which they excite, as antidepressants (morphine, scopolamine) or stimulants (strychnine, cafeine), others on the autonomic nervous system as sympathomimetics, sympatholytics, parasympathomimetic, parasympatholitycs, anticholinergics and genglioplegics24. We also note the existence of curariforms, antitumour local anaesthetics, antimalarials, amoebicudes and antiprotozoals. In most cases, they act at low doses, but even at this very low dose, they can be highly toxic25.
The biological properties of alkaloids are as varied as their structures; they are beneficial in the treatment of different diseases or dysfunctions of the human organism. Alkaloids also have anti-inflammatory, antioxidant and antibacterial properties26.
However, the alkaloids indoloquinoline, cryptolepin cause morphological changes in Staphylococcus aureus, but these effects may be due to other mechanisms since this compound is know to be a AND intercalator and an inhibitor of ADN synthesis through inhibition of topoisomerase. Berberine is endowed with bacteriostatic properties at low dose, bactericidal at higher dose, it is active on many germs (staphylococci, streptococci, but also salmonella, proteus, vibrios…etc.)27.
Colocynthis vulgaris (L.) Schrader is among the most abundant plant in the Ouargla region, especially in Oued N'sa, and also considered to be among the most popular and most used in phytotherapy. It's very rich in active principles. This plant is well known for its multiple biological activities. Its therapeutic efficacy is largely due to the presence of alkaloids. Thus several extraction methods have been recommended depending on amount of this active substance28.
The objective of this study is the selection of the best protocol for the extraction of alkaloids from Colocynthis vulgaris (L.) Schrader collected in the region of Oued N'sa.
MATERIALS AND METHODS:
Collection of plant material:
Colocynthis vulgaris (L.) Schrader plant used in this study (Fig.1) was harvested in the region of Oued N'sa-Ouargla during May 2019.
Figure 1: Colocynthis vulgaris (L.) Schrader.
Processing of plant material:
Fresh fruits of Colocynthis vulgaris (L.) Schrader were air-dried at room temperature. The dry matter obtained was ground to a powder using a mortar, and then sieved using a sieve of granulometry (0.1mm). All of these steps are summarized in Fig.2.
Figure 2: Preparation of the plant material.
Test for alkaloids:
The alkaloids proofing test consists of the following steps:
· Extract 10g of the pulverized drug with 50 ml of diluted HCl (1%);
· After basifying the extract with NH3, the mixture is subjected to three extracts with 20ml of CHCl3 each time and those until complete exhaustion;
· This organic phase is evaporated and the precipitate is dissolved in 2ml of dilute HCl (1%);
· Three drops of Mayer's reagent are added to the previous solution. The appearance of a white precipitate indicates the presence of alkaloids28,29.
Extraction of alkaloids:
a) Extraction of alkaloids in alkaline medium:
The extraction protocol shown in Fig.3 begins with degreasing, by mixing 50g of the plant powder with 300 ml of petroleum ether. After 24 hours of maceration, the mixture is filtered, the marc obtained is degreased then air-dried, and the filtrate is discarded.
The delipidated powder is taken up in 120ml of NH40H (25%) for a 24-hour maceration to release the alkaloids from their saline combinations.
The released bases are dissolved in 100ml of CHCl3 for 24hours. The mixture is filtered, the marc is removed and the filtrate is concentrated until reaching a volume of 40ml.
The organic phase is extracted with H2SO4 at a rate of 15 ml per extraction, in our case we carried out three extracts. The recovered acid phase is mixed with 10ml of NH40H (25%), and then subjected to three successive extractions with CHCl3 at a rate of 20ml per extraction.
After this operation, the exhausted acid phase is removed and the organic phase is washed with distilled water, dried with Na2SO4 and left at room temperature for the time necessary to allow the evaporation of the organic solvent. The precipitate obtained represents the total alkaloids as salts.
Figure 3: Extraction of alkaloids in alkaline medium.
Figure 4: Extraction of alkaloids in acid medium
b) Extraction of alkaloids in acid medium:
The extraction protocol shown in Fig. 4 is carried out as follows:
· Mix 2g of the plant powder with 10ml of H2SO4 0,1 N, stir for 10 minutes and filter through paper after washing the filtrate with H2SO4 0,1 N until 20ml of filtrate is obtained;
· Add 1ml of NaOH 0,1N to check the pH, which must be alkaline, extract with 2 times 10ml of CH2Cl2, dry the organic phase recovered over anhydrous Na2SO4, filter and evaporate the solution obtained. The precipitate obtained represents the total alkaloids in salt form.
RESULTS AND DISCUSSION:
Detection of alkaloids:
The phytochemical test for the presence of alkaloids was positive, which confirms the existence of alkaloids in the fruit of Colocynthis vulgaris (L.) Schrader.
However, the fruit pulp is the organ richest in alkaloids compared to the epicarp and seeds as shown in the table below.
Table 1: Test for the detection of alkaloids in the fruit of colocynth
|
Organ |
Test |
|
Pulp |
+++ |
|
Epicarp |
++ |
|
Grain |
+ |
Key: +++: important Presence, ++: average Presence and +: weak presence
Extraction of alkaloids:
The results presented in Fig.5 show that the rate of alkaloids obtained following the application of an extraction protocol in acid medium is significantly higher than that obtained during the application of protocol in alkaline medium.
Figure 5: Extraction rates of alkaloids in two different mediums.
The protocol in acid medium is carried out in one hour with a limited number of steps while the protocol in alkaline medium is carried out in four days and requires a series of daily operations.
The protocol in acidic medium consumes less chemical product and can be performed with only 2grams of plant material as opposed to the protocol in alkaline medium which requires at least 40grams of plant material for it to be feasible.
Thus, we can see that the extraction protocol in acid medium is the most suitable for the extraction of alkaloids from Colocynthis vulgaris (L) Schrader.
We recommend it because of its simplicity, short production time, economy in terms of plant material and chemical consumption. All these advantages make it a safe process for multiple manipulations.
CONCLUSION:
The Colocynthis vulgaris (L) Schrader was chosen in this study on the basis of its abundance in southern Algeria as well as its use in local traditional medicine for the treatment of certain diseases. Indeed, this plant contains various secondary metabolites including alkaloids.
The present study is devoted to the extraction of alkaloids from Colocynthis vulgaris (L) Schrader by maceration using two different extraction protocols.
The results of this study confirm the existence of alkaloids in the fruit of study plant and show that the best extraction rate (33.5%) is obtained by using an extraction in acid medium.
CONFLICT OF INTEREST:
The authors have no conflicts of interest regarding this study.
ACKNOWLEDGMENTS:
The authors would like to thank the staff of Applied Sciences Faculty Laboratory/ Kasdi Merbah Ouargla University, Algeria, for their valuable cooperation in carrying out the experimental tests
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Received on 31.07.2022 Modified on 24.12.2022
Accepted on 08.03.2023 ©AJRC All right reserved
Asian J. Research Chem. 2023; 16(2):128-132.
DOI: 10.52711/0974-4150.2023.00021