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. 2025 Sep 29;15(10):363. doi: 10.1007/s13205-025-04532-x

Sesamin and α-linolenic acid protection against hyperlipidemia-induced obesity in Drosophila melanogaster model

Abu Safana Biswas 1, Rajeshwari Selvaraj 2, Kamsagara Linganna Krishna 1,✉, Ganavi Bethanagere Ramesha 1, Bharat Jayaprakash Byalahunashi 1, Seema Mehdi 1, Suman Pathak 3
PMCID: PMC12480314  PMID: 41035527

Abstract

This study evaluates the anti-hyperlipidemic activity of phytoconstituents, such as sesamin and α-linolenic acid (ALA), in a high-fat diet (HFD)- induced Drosophila melanogaster obese model. Male wild-type Drosophila were fed an HFD (10% coconut oil) to induce obesity for 5 days. The obese flies were treated with the test compounds (sesamin, ALA) and rosuvastatin as a standard drug. Sesamin and ALA were tested for acute toxicity studies to calculate the doses for the efficacy study. Body weight, climbing, negative geotaxis, and cold stress were performed as behavioural assays, and lipid profiling (total cholesterol, triglycerides (TG, LDL, and HDL), glucose levels, and total protein content was determined as biochemical assays. Results demonstrated that sesamin and ALA have good potency in significantly improving behavioural parameters (P < 0.05). However, when compared, ALA showed better potency in enhancing these parameters. ALA has better potency in reducing TG and LDL levels, which are responsible for cardiovascular disease. Sesamin has better efficacy in improving the level of HDL, which an HFD reduces in flies, which is responsible for atherosclerosis. Sesamin reduced lipid profile in a dose-dependent manner, but required higher doses to achieve comparable lipid-lowering effects. In conclusion, ALA exhibited more substantial protective effects in reducing hyperlipidemia compared to control flies as an anti-hyperlipidemic agent in a dose-dependent manner.

Keywords: α-linolenic acid, Sesamin, Drosophila melanogaster, Anti-hyperlipidaemic activity, Rosuvastatin

Obesity is a primary global health concern and a prominent non-communicable disease that significantly increases the risk of cardiovascular disease (CVDs), type 2 diabetes, hypertension, and metabolic disorders. Once considered a condition affecting primarily affluent nations, obesity now affects diverse age groups worldwide, including children and adolescents, with rising prevalence in developing countries such as India (Sharma et al. 2024; Banerjee et al. 2021). It is linked to increased morbidity, mortality, and reduced life expectancy. Remarkably, excessive production of reactive oxygen species (ROS) plays a key role in the pathogenesis and progression of obesity-associated CVDs (Parsanathan et al. 2020).

According to recent projections, the global obesity rate is expected to rise dramatically in the coming years, with some reports estimating that over half the global population may be overweight or obese by 2030 (Luhar et al. 2020). In India, the prevalence of obesity shows significant regional variation, ranging from 11.5% to 16.5% in Jharkhand to 31.5% to 37% in Chandigarh, as highlighted by the National Family Health Survey (NFHS-IV). Kerala, Punjab, Delhi, and Sikkim report the highest obesity rates, whereas Jharkhand and Bihar exhibit comparatively lower rates (Madhu et al. 2022).

Sesamum indicum Linn., a member of the Pedaliaceae family, is one of the earliest domesticated species. Its seeds, rich in secondary metabolites such as steroids, terpenes, phenolic compounds, etc., are utilized for medicinal use. In addition to the seed, which is already utilized for medicinal uses, the other parts of the plant are also helpful for wound healing, analgesic activity, alopecia, constipation, and antifungal activity (Mili et al. 2021; Andargie et al. 2021; Zoclanclounon et al. 2023; Mushtaq et al. 2020). Sesame seed oil, extracted using a Soxhlet apparatus, was tested on Wistar Albino Rats and showed significant anti-dyslipidaemia activity (Khatun et al. 2021). In an in vivo study, Sesamin (50 mg/kg) inhibited high glucose-induced inflammation and oxidative stress in retinal microglia (Zhang et al. 2022). Sesame seeds are widely cultivated in tropical and subtropical regions of Africa, Asia, and the Americas (Adeyanju et al. 2022). Flaxseed, derived from Linum usitatissimum of the Linaceae family, is a functional food rich in dietary fibre, α-linolenic acid (ALA), proteins, and phytoestrogens (Villarreal et al. 2022). Flaxseeds are rich in PUFAs, lignans, omega-3 s, and fibre, with ALA as a key component. Flaxseed oil has anti-inflammatory and antioxidant properties that support its anti-hyperlipidaemic effects (Babar et al. 2021; Ismail et al. 2024).

Flaxseed (Linum usitatissimum), from the Linaceae family, is another functional food source that is rich in α-linolenic acid (ALA), dietary fibre, protein, polyunsaturated fatty acids (PUFAs), lignans, and omega-3 fatty acids. It has been studied for its potent anti-inflammatory, antioxidant, and anti-hyperlipidaemic properties (Babar et al. 2021; Ismail et al. 2024; Villarreal et al. 2022).

In the present study, we used the fruit fly Drosophila melanogaster as a model organism to investigate the protective effects of sesamin and α-linolenic acid (ALA) against high-fat diet-induced obesity and lipid dysregulation. D. melanogaster has been widely used in biomedical research since the early 1900s due to its short life cycle (9–10 days at 25 °C), high fecundity, and well-conserved metabolic pathways (Eickelberg et al. 2022).

The study used male Drosophila melanogaster (wild-type Oregon K strain) procured from the Department of Studies in Zoology, University of Mysore, Mysuru, Karnataka. The compounds used for the experiment included sesamin, α-linolenic acid, and rosuvastatin. Sesamin was extracted from sesame oil using methanol, hexane, and Na2SO4, while ALA was isolated from flaxseed oil by employing a base solution and chloroform partitioning. Biochemical kits were utilized for analysing glucose (SPINLAB-180, Amposta, Spain), total protein (SPIN640 kits, Amposta, Spain), high-density lipid (HDL), low-density lipid (LDL), total cholesterol, and triglycerides (TGA) (Agappe, Kerala, India). A high-fat diet (HFD) was prepared by supplementing the control diet with 10% coconut oil to induce obesity in the flies. Equipment such as 96-well plates was used during the experiments, and a Nanodrop plate reader was employed to measure biochemical parameters.

Sesamin was extracted from 100 g of sesame oil using methanol, a polar solvent, for initial extraction. The oil was mixed with 500 ml of methanol and incubated at 60 °C in a water bath for 8 h to dissolve polar and mid-polar components. The upper methanol layer was separated and concentrated using a rotary evaporator. The resulting residue was then treated with 20 ml of non-polar solvent hexane to isolate non-polar components such as sesamin. The hexane phase was again concentrated using a rotary evaporator. The residue was filtered through anhydrous sodium sulphate (Na2SO4) to remove residual moisture. A few drops of fresh hexane were added to initiate crystallization, and the crystal powder was collected and stored for further use (Dar et al. 2015). The percentage of yield was found to be 0.2% w/w.

To extract α-linolenic acid (ALA), 10 g of flaxseed oil was transferred into a round-bottom flask. A basic hydrolysis method was used for extraction, in which five volumes of alcoholic potassium hydroxide solution (ethanolic KOH, 0.5 N) were added and stirred at 60 °C for 2 h to break down triglycerides into free fatty acids. The reaction mixture was then partitioned twice with chloroform, a nonpolar solvent, to extract the liberated fatty acids. The chloroform layers were collected and concentrated using a rotary evaporator. Approximately 2 g of oil-rich ALA fraction was obtained (Khattab et al. 2013).

Flies were maintained in a wheat cream agar medium supplemented with dry yeast powder in the Pharmacology Department, JSS College of Pharmacy, Mysuru. They were housed under controlled laboratory conditions, a 12:12 light–dark cycle at 25 ± 1.2 °C and relative humidity of 70–80%. Adult flies were transferred to fresh vials every 48 h to maintain optimal health.

An acute toxicity assay was conducted to determine the safe working doses of sesamin and ALA. Thirty flies per compound (n = 10 in triplicate) were randomly selected and exposed to different concentrations of each test compound mixed into the wheat cream agar medium. The concentrations for ALA were 0.226 mg/ml, 0.045 mg/ml, 0.0113 mg/ml, while sesamin was tested at 0.02 mg/ml, 0.04 mg/ml, 0.01 mg/ml. The lethal concentration 50% (LC50) values were calculated using probit analysis (Sentürk et al. 2020).

20 Flies per group were divided into nine groups: Normal, Control, Standard (rosuvastatin 10 mg/kg), sesamin at high (SHD), middle (SMD), and low (SLD) doses; ALA at high (AHD), middle (AMD), and low (ALD) doses. A high-fat or control diet was prepared with 10% coconut oil mixed with normal wheat cream media, and the high-fat diet was fed to Drosophila for 5 days (Asiimwe et al. 2023). Treatments were given for 28 days, with behavioural assays including body weight, climbing test, and negative geotaxis conducted before inducing a high-fat diet and on days 1, 7, 14, 21, and 28 (Spierer et al. 2021; Liao et al. 2021). Data were collected on days 0, 1, 7, 14, 21, and 28 to evaluate the effectiveness of test compounds against high-fat diet-induced obesity in flies. After a 2 h, 5 °C water bath, survival was assessed by counting recovered flies after 24 h (Singh et al. 2022).

Glucose, protein, HDL, LDL, cholesterol, and TGA levels were determined in Drosophila to assess the effects of a high-fat diet and test compounds. Serum was obtained from the fly blood by centrifugation on day 28 post-sacrifice. Blood glucose levels were measured using SPINLAB-180 kits, with serum stored in Tris–HCl buffer and absorbance recorded at 505 nm. Total protein levels were estimated using SPIN640 kits, with serum stored in phosphate buffer and absorbance at 540 nm. HDL and LDL levels were measured using specific kits, with serum preserved in a 2:1 chloroform–methanol solution and absorbance at 630 nm and 578 nm. Cholesterol levels were determined using cholesterol kits, with absorbance recorded at 505 nm. TGA levels were measured using TGA kits, following the same serum preservation method, with absorbance at 505 nm. Blank and standard absorbance values were used for final concentration calculations.

All data were expressed as Mean ± SEM. Statistical significance was assessed using two-way ANOVA in GraphPad Prism Software, followed by Tukey’s multiple comparisons, with relevance at P < 0.05 (Pluijmert et al. 2020). Significance levels were represented by superscripts (a-significant when compared to Normal, b-significant when compared to control, and c-significant when compared to standard).

Mortality was recorded after test sample exposure at 24, 48, 72, and 96 h, and LC50 values were found to be 16.819 µg/ml for ALA and 1.99 mg/ml for sesamin. The efficacy of the test samples was evaluated using sublethal doses of 1/10th, 1/20th, and 1/30th concentrations of 16.819 µg/ml for ALA and 1.99 mg/ml for sesamin.

Body weight, the climbing test, and the negative geotaxis assay were behavioural markers. The parameters were measured before starting the study, and the data were compared with high-fat diet-induced flies to determine whether the test samples were effective. Following the administration of test compounds on days 1, 7, 14, 21, and 28 of treatment, data were collected and compared to assess the effectiveness of the test materials. It was observed that the test compounds significantly reduced body weight, which had increased due to HFD, and improved locomotor activity.

A high-fat diet induced obesity in Drosophila, significantly increasing body weight compared to the normal flies. Measurements on treatment days 7, 14, 21, and 28 showed that high doses of sesamin and ALA reduced weight from day 7, while low doses of sesamin and ALA showed effects from days 21 and 14, respectively. High doses of both tested compounds showed similar effects to those of the standard drug (Fig. 1).

Fig. 1.

Fig. 1

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Shows the protective effect of sesamin and α-linolenic acid (ALA) against high-fat diet-induced hyperlipidaemia model of Drosophila melanogaster (Changes in Body weight). Values are expressed as Mean ± SEM (n = 20). The statistical significance was calculated by two-way ANOVA followed by Tukey’s multiple comparisons. a P < 0.05. Compared with the normal group, b P < 0.05 Compared with the Control group. c P < 0.05 Compared with the standard group

A high-fat diet induced obesity in Drosophila, reducing their climbing ability. Climbing assays on treatment days 7, 14, 21, and 28 showed that high doses of sesamin and ALA significantly improved mobility from day 14, while low doses showed effects from day 21. At high-dose test compounds showed no significant difference from the standard drug (Fig. 2).

Fig. 2.

Fig. 2

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Shows the protective effects of sesamin and α-linolenic acid (ALA) against high-fat diet-induced hyperlipidaemia model of Drosophila melanogaster (Climbing assay). Values are expressed as Mean ± SEM (n = 20). The statistical significance was calculated by two-way ANOVA followed by Tukey’s multiple comparisons. a P < 0.05. Compared with the normal group, b P < 0.05 Compared with the Control group. c P < 0.05 Compared with the standard group

Before starting treatment, a high-fat diet was fed to the Drosophila to induce obesity. After inducing obesity in the flies on the first day, negative geotaxis was performed and compared between normal and obese flies. This graph represents the comparison between the control group and treatment group, and it was observed that sesamin and ALA at high doses showed significant changes from the 14th day of treatment. In contrast, sesamin and ALA at low doses showed substantial changes at the 28th day. The test compounds with high doses have no significant difference from standard drugs (Fig. 3).

Fig. 3.

Fig. 3

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Shows the protective effects of sesamin and α-linolenic acid (ALA) against the high-fat diet-induced hyperlipidaemia model of Drosophila melanogaster (locomotor activity). Values are expressed as Mean ± SEM (n = 20). The statistical significance was calculated by two-way ANOVA followed by Tukey’s multiple comparisons. a P < 0.05. Compared with the normal group, b P < 0.05 Compared with the Control group. c P < 0.05 Compared with the standard group

Flies were submerged to observe survival rates, and cold stress was applied on day 28. The treatment group showed a higher survival rate than the obese group (Fig. 4).

Fig. 4.

Fig. 4

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Shows the protective effects of sesamin and α-linolenic acid (ALA) against the high-fat diet-induced hyperlipidaemia model of Drosophila melanogaster (Survival rate). Values are expressed as Mean ± SEM (n = 20). The statistical significance was calculated by two-way ANOVA followed by Tukey’s multiple comparisons. a P < 0.05. Compared with the normal group, b P < 0.05 Compared with the Control group. c P < 0.05 Compared with the standard group

The study assessed Drosophila’s glucose, lipid profile, cholesterol, protein, and triglyceride levels. Test groups had lower glucose, cholesterol, LDL, and triglyceride (TGA) levels and higher HDL levels than the obese group. High-dose test drugs were most effective in reducing glucose, cholesterol, LDL, and TGA levels, comparable to the standard drug. Sesamin at high doses raised HDL levels more effectively than ALA or lower doses. ALA significantly reduced LDL at all doses, while sesamin showed minimal effect at medium and low doses. Total protein content remained unchanged across groups (Table 1).

Table 1.

Protective effect of sesamin and ALA against high-fat diet-induced hyperlipidaemia in the Drosophila model

Group Glucose (µg/mg) Protein (µg/mg) HDL (µg/mg) LDL (µg/mg) Cholesterol (µg/mg) TGA (µg/mg)
Normal 05.29 ± 0.77 0.31 ± 0.05 26.10 ± 0.25 11.48 ± 0.25 66.67 ± 0.95 127.86 ± 0.68
Control 26.30 ± 2.15a 0.13 ± 0.04a 21.78 ± 0.13a 16.19 ± 0.21a 95.53 ± 4.45a 173.02 ± 0.06a
Standard 08.73 ± 0.83b 0.22 ± 0.04 25.47 ± 0.13b 10.95 ± 0.27b 73.22 ± 0.50b 127.66 ± 0.15b
SLD 19.20 ± 1.15b,c 0.17 ± 0.05 22.32 ± 0.13 14.88 ± 0.28c 86.34 ± 1.06b,c 172.24 ± 0.66c
SMD 16.20 ± 0.37b,c 0.21 ± 0.03 24.50 ± 0.25 13.45 ± 0.19 84.15 ± 0.79b,c 145.65 ± 0.87b,c
SHD 11.9 ± 0.22b 0.22 ± 0.06 25.83 ± 0.13b 10.33 ± 0.11b 76.50 ± 0.21b 141.54 ± 0.65b,c
ALD 21.2 ± 0.067b,c 0.16 ± 0.04 22.14 ± 0.22 14.14 ± 0.082b 87.21 ± 0.87b,c 165.98 ± 0.87b,c
AMD 09.24 ± 0.62b 0.23 ± 0.06 22.05 ± 0.13 12.01 ± 0.067b 81.53 ± 0.53b,c 163.04 ± 0.09b,c
AHD 07.18 ± 0.29b 0.29 ± 0.05 23.14 ± 0.13 11.11 ± 0.16b 77.38 ± 0.95b,c 147.99 ± 0.45b,c
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SLD Sesamin low dose, SMD Sesamin medium dose, SHD Sesamin high dose, ALD α-linolenic acid low dose, AMD α-linolenic acid medium dose, AHD α-linolenic acid high dose

Values are expressed as Mean ± SEM (n = 20). The statistical significance was calculated by two-way ANOVA followed by Tukey’s multiple comparisons

a P < 0.05. Compared with the normal group

b P < 0.05 Compared with the Control group

c P < 0.05 Compared with the standard group

The plant-derived alpha-linolenic acid (ALA), which was isolated from Linum usitatissimum, and sesamin, a lignan derived from Sesamum indicum, are both recognized for their strong anti-inflammatory and antioxidant qualities (Yuan et al. 2022; de Abreu et al. 2022; Kitipaspallop et al. 2023; Majdalawieh et al. 2022). A Drosophila melanogaster model of diet-induced obesity and hyperlipidaemia was used in the current work to assess their preventive effects.

According to acute toxicity studies, ALA and sesamin had LC₅₀ values of 16.819 µg/ml and 1.99 mg/ml, respectively, which indicated comparatively low toxicity. To evaluate their therapeutic potential, sublethal dosages of LC₅₀ (1/10th, 1/20th, and 1/30th) were employed. The anti-obesity and metabolic-modulating effects of both compounds were assessed by flies’ behavioral and biochemical parameters such as body weight, climbing ability, negative geotaxis, cold stress, and lipid profile.

Body weight, climbing ability, and negative geotaxis were selected as the primary behavioural indicators. Confirming the successful induction of obesity, baseline data on day 0 showed that the control (high-fat diet) group had a significantly higher body weight than the normal group (Nayak et al. 2021). Interestingly, both ALA and sesamin showed dose-dependent decreases in body weight starting on day 21, indicating a reversal of fat storage. Obese flies showed markedly reduced climbing activity (Fig. 2), a marker of locomotor dysfunction frequently linked to metabolic load. ALA and sesamin treatment enhanced climbing behaviour; the highest dosages showed notable improvements from day 14, indicating muscular and metabolic recovery. Additionally, both drugs, particularly at higher dosages, improved the poor locomotion in the negative geotaxis assay compared to the control group (Fig. 3), confirming their function in maintaining muscular coordination. The control group’s survival rates were much lower under cold stress conditions (Fig. 4). On the other hand, both test compounds increased stress resilience, which suggests that they have cytoprotective effects due to their antioxidant qualities.

According to biochemical analysis, the control group exhibited a typical hyperlipidaemic and hyperglycaemic state with considerably higher levels of glucose, cholesterol, triglycerides (TG), and LDL and lower levels of HDL (González et al. 2023; Prasad et al. 2022). Sesamin and ALA treatment vividly reversed these patterns. Superior antihyperglycemic efficacy was demonstrated by the constant superiority of ALA over sesamin in lowering blood glucose levels, especially at lower dosages. In the treatment groups, the levels of both LDL and cholesterol were significantly reduced, whereas ALA showed a higher effect. Both sesamin and ALA showed cardioprotective potential, as increased LDL is a major contributor to atherosclerosis and cardiovascular disorders (Mattiuzzi et al. 2020; Domanski et al. 2020). Sesamin and ALA restored HDL levels, which are usually decreased in hyperlipidaemia, with sesamin having a marginally higher effect. This is important because HDL prevents cardiovascular events and mediates reverse cholesterol transfer (Casula et al. 2021; Sekhar et al. 2020). Both compounds successfully reduced triglyceride levels. ALA demonstrated better effectiveness, especially at low dosages, indicating a strong lipid-lowering effect (Packard et al. 2022). Overall, the treatments did not affect protein metabolism, as seen by the lack of substantial differences in protein levels between groups.

These results imply that ALA and sesamin have diverse therapeutic potential in addressing metabolic dysregulation from a high-fat diet. Their potential as natural treatments for obesity-related issues is highlighted by their effectiveness in lowering body weight, enhancing behavioural performance, and reestablishing glucose and lipid balance. A strong contender for additional research in models of metabolic disorders, ALA in particular, showed marginally better efficacy across the board.

We investigated the protective effects of sesamin and ALA against obesity in Drosophila fed with HFD. The sesamin and ALA demonstrated dose-dependent protective effects through behavioural and biochemical parameters employed for the study. When the data obtained from various parameters were compared with a high-fat diet-fed Drosophila, it was observed that these test phytochemicals have the potential to reduce the hyperlipidaemic condition, which is responsible for cardiac disorders such as atherosclerosis, angina pectoris, heart stroke, etc. By comparing the data of sesamin and ALA, it was observed that ALA has more potency than sesamin at low doses (8.1% vs 12.7% for LDL and 0.5% vs 4.1% for TG) to reduce the lipid profile. As a result of this finding, ALA may be explored to develop a novel therapy for cardiac diseases, whereas sesamin may be studied at higher doses for CVD.

Acknowledgements

The authors thank the Department of Pharmacology and the Principal, JSS College of Pharmacy, JSS Academy of Higher Education & Research, Mysuru, for their support and the necessary infrastructure to perform the research. We also thank the Department of Studies in Zoology at the University of Mysore, Mysuru, for providing Drosophila melanogaster flies.

Funding

This research received no external funding.

Data availability

Data will be made available on request.

Declarations

Conflict of interest

All authors declare no conflict of interest.

Ethical approval

No fish or rodent models are used in this article, and ethical approval is not applicable.

Informed consent

No human subjects are in this article, and informed consent is not applicable.

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