Journal of Chromatography & Separation Techniques
Open Access

Qualitative HPTLC Phytochemical Profiling of the Seeds of Swietenia mahagoni Jacq

Vigneshwaran L.V^1* and Lalitha K.G^{2 *}Correspondence: Vigneshwaran L.V, Department of Pharmacy, Malik Deenar College of Pharmacy, Kerala University of Health and Allied Sciences, Kerala, India, Email:

Author info »

Introduction

The Indian medical systems are regarded as a vast repository of knowledge from which many beneficial things can be learned. Native medications are quite significant from an economic and professional standpoint. Many of the drugs that are currently on the market have either been directly or indirectly produced from plants, which have historically been an excellent source of pharmaceuticals. Simple assertions do not satisfy the scientific mind unless they are supported by experimental data. A thorough research of indigenous pharmaceuticals would enable much more to be done to advance their cause and make them truly beneficial to the people of this nation [1].

Diabetes mellitus

Diabetes, frequently known as diabetes mellitus, is a set of metabolic ails characterized by persistently elevated blood sugar situations. Frequent urine, increased thirst, and increased hunger are signs of elevated blood sugar. Diabetes can lead to a wide range of consequences if ignored. Diabetic ketoacidosis and nonketotic hyperosmolar coma are exemplifications of acute complications. Cardiovascular complaint, stroke, habitual order failure, bottom ulcers, and eye damage are serious long term consequences [2]. A high frequency of threat factors was set up among the diabetic subject’s including pre-hypertension (37), hypertension (53.7), fat (14.8), rotundity (57.4), hypercholesterolemia (25.9) and hypertriglyceridemia (39.6).

Numerous indigenous Indian medicinal shops have been set up to be useful to successfully manage diabetes. One of the great advantages of medicinal shops is that these are readily available and have veritably low side goods. India has about 45,000 factory species and numerous of them have medicinal parcels [3].

Materials and Methods

Plant profile

Swietenia mahagoni Jacq. (Meliaceae) is a large, deciduous, and economically important timber tree native to the Central America and is commonly known as “Mahogany”. Commonly it is called as Puerto Rico, Spanish, Cuban or Jamaica Mahagony tree [4]. It is an important plant that is connected to the African genus Khaya and the source of one of the continent's most wellknown traditional remedies (Figure 1) [5].

Figure 1: Seed pods of S. mahagoni Jacq.

Preparation and extraction of plant material

Plant drying and size reduction: Small pieces of S. mahagoni Jacq. seeds were dried at room temperature in the shade. The seeds were reused into a fine, coarse form. The factory material was kept in a watertight vessel in a clean, dark position. Birth of factory material, identification of the phytoconstituents, quantitative computation of the total phenolic and flavonoid content, and HPTLC analysis are some of the possible way in phytochemical examinations of a factory (Figures 2-4).

Figure 2: Opened fruits of S. mahagoni JACQ.

Figure 3: Seeds of S. mahagoni Jacq. (with cotyledons), collection location: South Canara, India.

Figure 4: Seeds of S. mahagoni Jacq. (without cotyledons), collection location: South Canara, India.

Extraction

Using specific solvents in a typical extraction method, extraction includes separating the bioactive portion of the plant seed from the inert components. These preparation types include the decoction, fluid extraction, tinctures, semisolid extract, and powder categories. To obtain the concentrated form of the active chemical, extraction is done for a variety of reasons.

Extraction method

Using a Soxhlet extractor, 500 g of the air dried powder from the seeds of S. mahagoni Jacq. were repeatedly extracted with solvents of increasing polarity.

Petroleum ether extract of seeds of S. mahagoni Jacq: Using the continuous hot percolation method and a Soxhlet apparatus, 2 litres of petroleum ether were used to extract the dried coarse powder of S. mahagoni Jacq. The extraction was finished after 72 hours, and after taking an extract of petroleum ether, the solvent was again distilled. A desiccator was used to retain the extracted material, which was brown in hue.

Benzene extract of seeds of S. mahagoni Jacq: The marc that remained after the petroleum ether extraction was dried and then extracted using the continuous hot percolation process with 2 liters of benzene. The extraction was finished after 72 hours. After filtering, distillation at low pressure was used to get rid of the solvent. The marc was dried for further extraction, and the light brown extract was kept in a desiccator.

Ethyl acetate extract of seeds of S. mahagoni Jacq: The marc that remained after the benzene extraction was dried and then extracted using the continuous hot percolation process using 2 liters of ethyl acetate. The extraction was finished after 72 hours. After filtering, distillation at low pressure was used to get rid of the solvent. The marc was dried for additional extraction before the dark brown extract was kept in a desiccator.

Ethanolic extract of seeds of S. mahagoni Jacq: After ethyl acetate extraction, the remaining marc was dried and then extracted with 2 liters of ethanol using the continuous hot percolation method. The extraction was finished after 72 hours. After filtering, distillation at low pressure was used to get rid of the solvent. The extract, which was semisolid in colour and yellowish brown, was kept in a desiccator. All of the aforementioned extracts were employed for pharmacological and biological screening, identification of plant ingredients, and phytochemical content analysis.

Phytochemical screening: To identify the plant's active ingredients, phytochemical experiments were performed on the various extracts of S. mahagoni Jacq. The outcomes were displayed in Table 2.

HPTLC analysis of ethanol extract S. mahagoni Jacq plant sample with oleanolic acid standard

Preparation of test solution: The provided plant sample of S. mahagoni Jacq's ethanolic seed extract was centrifuged at 3000 rpm for 5 minutes. For an HPTLC analysis, this solution served as a test solution.

Preparation of standard solution: For the analysis, 1 mg of the prescribed amount of oleanolic acid was combined with 1 ml of chloroform.

Mobile phase: Toluene-ethyl acetate (8:2)

Spray reagent: 10% H₂SO₄ in methanol

Sample application and spot development: The samples were spotted in bands of 5 mm using a Hamilton 100- l hype on an aluminum plate 60 GF₂₅₄ that had been carpeted in silica gel (20 cm × 10 cm × 250 m; E. Merck). Prior to chromatography, the plates were actuated at 110â?? for 5 twinkles after being prewashed with methanol. The distance between each band was kept at 6 mm. The CamagLinomet 5 automatic TLC sample applicator was used at a constant operation rate of 100 nl/s. The slit size was held to 5 mm × 0.45 mm, and the scanning speed used was 10 mm/s. Each track was tri-sectionally scrutinized, the monochromator band range was set to 20 nm, and birth correction was applied. Linear thrusting development was carried out in a 20 cm × 10 cm binary through glass chamber impregnated with the mobile phase. The optimized chamber achromatism time for mobile phase was 30 min at room temperature (25â?? ± 2â??) at relative moisture of 60 ± 5. The length of chromatogram run was 9 cm. posterior to the development, HPTLC plates were dried in current of air with the help of air teetotaler in a rustic chamber with acceptable ventilation. On a CAMAG REPROSTAR 3 scanner running Palm pussycats software (winCATS1.3.4 interpretation), densitometry scanning was carried out in the reflectance absorbance mode at visible light, UV 254 nm, and UV 366 nm and the images were taken. Deuterium lights furnishing a nonstop UV diapason between 190 nm and 400 nm were used as the radiation source [6].

Derivatization: The formed plate was sprayed with the appropriate spray reagent and baked in a hot air oven at 100â?? for drying. The plate was photographed utilising the photodocumentation (CAMAG REPROSTAR 3) chamber in visible light mode.

Scanning: Derivatization was followed by fixing the plate to the scanner stage of the CAMAG TLC scanner 3 and performing 500 nm scanning. It was noticed the peak table, peak display, and peak densitogram. The sample loaded plate was held in the TLC twin trough development chamber with the appropriate mobile phase up to 90 mm (after being saturated with solvent vapour).

Photo-documentation: To remove solvents from the created plate, hot air was used to dry it. The plate was held in a photodocumentation chamber (CAMAG REPROSTAR 3) while images were taken at visible, UV 254 nm, and UV 366 nm wavelengths.

Results and Discussion

Phytochemical investigations of Swietenia mahagoni Jacq

The extraction of plant material and subsequent identification of the phytoconstituents are steps in the phytochemical analysis of a plant. 500 gm of the air-dried powder from the seeds of S. mahagoni Jacq. Were extracted using a solvent extractor in steps with increasing polarity, and the extracts were evaporated using a rotor evaporator to obtain the concentrate of active ingredient. The following solvents produced the highest extract yields: Petroleum ether (1.45%), benzene (1.68%), ethyl acetate (10.24%), and ethanol (12.86). The ethanol extract of HN was determined to have the highest yield (12.86%). In the Table 1 below, the % yield of various S. mahagoni Jacq seed extracts is reported [7].

Table 1: Successive solvent extraction of Swietenia mahagoni Jacq.

Phytochemical screening: Below are the findings of the preliminary phytochemical screening tests conducted on the various solvent extracts of S. mahagoni Jacq. seeds. Alkaloids, carbohydrates, flavonoids, phytosterols, saponins, phenolic compounds, tannins, lignins, proteins, free amino acids, gums, and mucilage were all present in the petroleum ether extract, but glycosides, fixed oils, and fats were not. Alkaloids, carbohydrates, flavonoids, phenolic compounds, tannins, lignins, proteins, free amino acids, gums, and mucilage were all present in the benzene extract, but glycosides, fixed oils and fats, phytosterols, and saponins were not. Alkaloids, carbohydrates, glycosides, flavonoids, phytosterols, saponins, phenolic compounds, tannins, lignins, proteins, free amino acids, gums, and mucilage were all present in the ethanol acetate extract, but fixed oils and fats were not present. Alkaloids, sugars, glycosides, flavonoids, phytosterols, fixed oils and fats, saponins, phenolic compounds and tannins, lignins, proteins and free amino acids, gums, and mucilage are all present in ethanol extract (Table 2) [8].

Table 2: Preliminary phytochemical screening of Swietenia mahagoni Jacq. seed extracts.

HPTLC analysis of ESM-ethanol extracts of S. Mahagoni jacq. with oleanolic acid standard

For the HPTLC examination of S. mahagoni Jacq. samples and oleanolic acid with high resolution and repeatable results, mobile phases of various compositions were examined. Toluene: Ethylacetae (8:2) was used as the mobile phase to accomplish a satisfactory separation of the phyto constituents, which produced a peak for oleanolic acid at RF 0.34. Figure 3 shows the HPTLC plate of the ESM before derivatization, and Figure 3 shows the HPTLC plate of the ESM after derivatization. After derivatization (10% H₂SO₄ in ethanol was sprayed on plates, dried at room temperature, and plates were heated for 5 minutes in an oven at 105â??), it was possible to see that there was a pink coloured zone in the visible mode of the SEM track, which indicated the presence of oleanolic acid (Table 3 and Figures 5-11).

Table 3: HPTLC data of ethanolic seed extract of S.mahagoni Jacq (ESM).

Figure 5: HPTLC plate of ESM before derivatization.

Figure 6: HPTLC plate of ESM after derivatization.

Figure 7: Sample KGL plant extracts sample-Baseline display (Scanned at 500 nm).

Figure 8: Sample KGL plant extracts sample peak densitogram display (scanned at 500 nm).

Figure 9: Track STD-Oleanolic acid standard baseline display (scanned at 500 nm).

Figure 10: Track STD-Oleanolic acid standard peak densitogram display (scanned at 500 nm).

Figure 11: 3D display of all tracks.

Figures 10 and 11, respectively, depict the densitometric peak of the ethanolic seed extract of S. mahagoni Jacq (ESM) and oleanolic acid. Figure 5 displays the ESM and oleanolic acid 3D chromatogram peaks. Using toluene-ethylacetate (8:2), an ethanolic extract of Swietenia mahagoni Jacq. seeds was created. The HPTLC chromatogram displayed a total of 12 peaks with various Rf values and peakareas @500 nm. Both the standard Oleonolic acid and the ESM had the same Rf value (0.34) in the chromatogram. This verifies that S. mahagoni Jacq's (ESM) ethanolic seed extract contains oleonolic acid.

Conclusion

The current study focuses on evaluating the qualitative phytochemical potential and HPTLC analysis of S. mahagoni Jacq. (Meliaceae), a plant used in traditional Chinese medicine to treat diabetic mellitus. Using a Soxhlet apparatus and the continuous hot percolation process, the plant components were shade dried, shrunk in size, and simultaneously extracted with a variety of solvents, including petroleum ether, benzene, ethylacetate, and ethanol. The following solvents likely contributed to the percentage yields of the extracts: petroleum ether: 1.45%; benzene: 1.68%; ethyl acetate: 10.24%; and ethanol: 12.86. The ethanol extract of ESM (12.86%) had the highest percentage yield. For this research study, the ethanolic extract was preferred because it produced a high yield. It was determined whether there were any flavonoids, amino acids, tannins, steroids, glycosides, or reducing sugars in the ethanolic extract of S. mahagoni Jacq seeds. Alkaloids, flavonoids, fixed oils, and lipids have all been detected in the ethanolic extract from this plant. When compared to the standard oleanolic acid, HPTLC analysis of the seeds of S. mahagoni Jacq reveals the presence of oleanolic acid. We need additional analysis in wet lab investigations based on the findings from the current study to evaluate and identify new medication candidates. Therefore, we can anticipate that using the seeds' extract of S. mahagoni Jacq could offer a natural diabetes treatment that avoids the drawbacks of synthetic drugs.

References

1. Tohir D, Wuyung PE, Farida R. Anti-cancer activity of Swietenia mahagoni seed extract in ethyl acetate induced breast cancer cell T47D. InAIP Conference Proceedings. 2020;2243(1).
2. Kumari K, Mathew BC, Augusti KT. Antidiabetic and hypolipidemic effects of S-methyl cysteine sulfoxide isolated from Allium cepa Linn. Indian J Biochem Biophys. 1995;32(1):49-54.

   [Google Scholar] [PubMed]

3. Lamb FB. On further defining mahogany. Econ Bot. 1963;17(3):217-232.

   [Crossref] [Google Scholar]

4. Poole CF. Thin layer chromatography: Challenges and opportunities. J Chromatogr A. 2003;1000(2):963-984.

   [Crossref] [Google Scholar] [PubMed]

5. Feigl F. Identification of individual organic compound. Spot tests in organic analysis. 4^th edition. Elsevier, London. 1956:237-245.

   [Google Scholar]

6. Jiang L, Numonov S, Bobakulov K, Qureshi MN, Zhao H, Aisa HA. Phytochemical profiling and evaluation of pharmacological activities of Hypericum scabrum L. Molecules. 2015;20(6):11257-11271.

   [Crossref] [Google Scholar] [PubMed]

7. Singh A, Lal UR, Mukhtar HM, Singh PS, Shah G, Dhawan RK. Phytochemical profile of sugarcane and its potential health aspects. Pharmacogn Rev. 2015;9(17):45.

   [Crossref] [Google Scholar] [PubMed]

8. Batiha GE, Tene ST, Teibo JO, Shaheen HM, Oluwatoba OS, Teibo TK, et al. The phytochemical profiling, pharmacological activities, and safety of Malva sylvestris: A review. Naunyn Schmiedebergs Arch Pharmacol. 2023;396(3):421-440.

   [Crossref] [Google Scholar] [PubMed]