INTRODUCTION:

Obesity is a disorder of excess fat accumulation in adipose tissues¹. It represents a main risk factor for a variety of chronic diseases as diabetes mellitus, cardiovascular diseases, metabolic disorder, and cancer ^2,3. The objective of anti-obesity treatment modalities is to achieve and maintain long term weight loss and reduce the incidence of obesity related diseases.

The current interventions include reduced-calories diet, behavioral changes, drugs and surgery¹. According to the lack of adherence to the mentioned therapies, dissatisfaction of patients and the short-term results, the need for new therapeutic modalities has increased especially complementary or alternatives medicine (CAM). Herbal medicine is considered one of the most important CAM used through the world⁴. They have shown an increase in body weight loss and an inhibitory effect toward enzymes responsible for fat metabolism as lipoprotein lipase (LPL) and pancreatic lipase (PL), given that the inhibition of such enzymes can reduce the digestion and absorption of dietary lipids⁵.

A variety of herbal extracts and plant isolated compounds have been studied for their anti-obesity activity due to their inhibitory effect toward LPL and PL^6,7. Lipoprotein lipase inhibition is suggested by researchers as a new anti-obesity mechanism. LPL is a member of lipase gene family, synthesized in parenchymal cells of different tissues. It is responsible for tri-acyl glycerol (TAG) hydrolysis in both circulating chylomicrons (CMs) and very low-density lipoproteins (VLDLs). LPL activity is regulated in a tissue-specific manner and affected by hormonal changes and energy requirements. LPL promotes the exchange of lipids between lipoproteins⁸. Through acting as a ligand for lipoproteins receptors, LPL regulates fatty acid supply to different tissues either to be stored or oxidized. All of these functions build a strong relationship between obesity and LPL activity⁹.

The high prevalence of obesity and the lack of safe pharmaceutical agents have encouraged us to study and evaluate the potential of different plant extracts as lipoprotein lipase inhibitor. Our target is to identify new, safe and effective natural anti-obesity agent as obesity affects the quality of human life significantly¹⁰.

MATERIAL AND METHODS:

Plant collection and extraction:

Twenty-one already prepared methanolic plant extracts were used in this study to assess their inhibitory effect on lipoprotein lipase enzyme. 10mg of each plant extract was dissolved in 1ml of DMSO to get a 10mg/mL solution. Then 20 μL of this solution was added to 980 μL of DMSO to prepare a diluted solution of 200 μg/mL, the final concentration of each of the used extract. Gas chromatography coupled with mass spectrometry (GC-MS) are the main methods of extraction of the active components of medicinal plants¹¹.

Measurements of lipoprotein lipase activity:

A colorimetric assay was implemented in this study to assess LPL activity in presence of p-nitrophenophenol butyrate (PNPB) as the enzyme substrate. Addition of PNPB (Sigma, USA) to LPL (Burkholderia sp., Sigma, USA, E.C. 3.1.1.34) leads to the release of p-nitrophenol which causes a change in the reaction color from being colorless into yellow. As the reaction progresses the releasing rates increase and the intensity of the color increases. The releasing rate of p-nitrophenol is detected by measuring the absorbance values over time using double beam UV-VIS spectrophotometer, Spectroscan 80D at 410nm. LPL activity was first measured without the addition of any plant extract as un-inhibitory control. Where 20 µL of the enzyme solution was mixed with 975 µL of Tris-HCl buffer (Promega, USA, E.E.C 201-064-4) using Vortex Mixer Kmc-1300V (Vision Scientific Corporation Korea). Thereafter, 5µl of the substrate were added to start the reaction, the mixture was further mixed for 30 seconds, transferred to cuvette, and the absorbance was measured over a period of 5minutes against Tris-HCl as a blank. Absorbance values were plotted versus time and the slope of the linear segment was used to determine the activity of lipoprotein lipase.

Statistical Analysis:

Data were analyzed by SPSS version 20. Triplicates of measurements were recorded and analyzed as mean ± standard deviation. Pearson correlation coefficient was calculated. The effect of extracts was calculated as follows % of inhibition or activation = (Treated - Control) / control X 100.

RESULTS:

The activity of the studied plant extracts on LPL enzyme is summarized in table 1. Inhibition, activation, as well as no activity on LPL have been shown. Figures 1, 2 and 3 represent the inhibitory action of the most active plant extracts respectively.

Activation or inhibition of the enzymes was categorized according to the percent of activation or inhibition into low (0 – 10 %), moderate (10 – 20 %) and high (> 20%). Eryngium creticum plant extract showed the highest activation of LPL enzymes (-26.79%) when compared with the control. Glaucium uleppicum showed moderate activation (-18.66 %) while low activation was detected for three plant extracts with decreasing manner; these plants included Majorana syriaca (-4.09%), Malva nicaeensis (-2.63%) and Achillea biebersteinii (-0.717%).

Inhibition of LPL enzyme activity was detected in 11 plant extracts. They were classified into low, moderate and high inhibition as follows:

Low inhibition was recorded for the extracts of Salvia spinosa, Lavendula angistifolia and Anchusa italic. The percent of inhibition was 0.21%, 2.64% and 4 %, respectively. Moderate inhibition was detected for the extracts of Silene aegyptiaca, Mentha spicata, Adonis palaestina and Fagonia arabica in the following order; 11.96%, 12.67%, 13.38% and 16.615%, respectively. High inhibition was observed for seven plant extracts; all had more than 20% inhibition in the activity of LPL enzyme. They included Anthemis palestina, Reseda lutea, Cleome Africana, Artemisia herba-alba, Chrysanthemum coronarium, Hypecoum dimidiatum and Onosma giganteum. The percent of inhibition was 21.06%, 22.29%, 23.07%, 23.38%, 27.81%, 29.038% and 32.92%. Figures 1-3 showed the highest inhibition rates of the studied plant extracts (Chrysanthemum coronarium, Hypecoum dimidiatum and Onosma giganteum, respectively)

Table 1: Activity of 200μg/mL plant extracts studied for their LPL inhibition activity

Plant	Family	Effect (%)	Inhibition/ Activation
Eryngium creticum Lam.	Apiaceae	-26.79	High activation
Glaucium uleppicum Boiss and Hausskn	Papaveraceae	-18.66	Moderate activation
Majorana syriaca (L.) Kostel	Lamiaceae	-4.09	Low activation
Malva nicaeensis All.	Malvaceae	-2.63
Achillea biebersteinii Afan.	Asteraceae	-0.717
Varthemia iphionoides Boiss and Blanche	Asteraceae	0	No effect
Salvia spinosa L.	Lamiaceae	0.21	Low inhibition
Lavendula angistifolia P. Mill	Lamiaceae	2.64
Anchusa italic Retz.	Boraginaceae	4
Silene aegyptiaca L.f.	Caryophyllaceae	11.96	Moderate inhibition
Mentha spicata L.	Lamiaceae	12.67
Adonis palaestina Boiss	Ranunculaceae	13.38
Fagonia arabica L.	Zygophyllaceae	16.615
Anthemis palestina Reut. ex Boiss	Asteraceae	21.06	High inhibition
Reseda lutea L.	Resedaceae	22.29
Cleome africana Botsch.	Capparaceae	23.07
Artemisia herba-alba Asso	Asteraceae	23.38
Chrysanthemum coronarium L.	Asteraceae	27.81
Hypecoum dimidiatum Delile	Papaveraceae	29.038
Onosma giganteum Lam.	Boraginaceae	32.92

Figure 1: Absorbance/Time profile of O. giganteum extract and control.

Figure 2: Absorbance/Time profile of H. dimidiatum extract and control.

Figure 3: Absorbance/Time profile ,C. coronarium extract and control.

DISCUSSION:

Medicinal plants represent an important field of research in the last decades due to their biological importance¹². In general, the extracts of some plants had antibacterial activities related to inhibition of bacterial growth of both gram positive and gram-negative bacteria^13-17. Possible antioxidant activity against colon cancer cells was reported as effects of some essential oils^16-20. Two essential metabolites namely flavonoids and alkaloids are isolated from the extract of most medicinal plants. Among the two phytochemical compounds of interest, alkaloids were found to be more effective than the flavonoids in exhibiting antimicrobial properties ^18,20-23. Insecticidal activity of essential oils of some plants was recorded^24,25. According to the underlying mechanism, obesity is classified into three types: Genetic obesity, hypothalamic obesity and dietary obesity²⁶. Genetics play a major role in the etiology of obesity where inheritability for body mass index ranges between 50-70% and for total body fat it reaches 80%²⁷. Hypothalamic obesity is defined as weight gain those results from hypothalamic injury, where hypothalamic responsible centers fail to control satiety and hunger. That’s why this type of obesity cannot be controlled by diet and needs either pharmacological treatment or surgery²⁸.

For dietary obesity, nutrients overconsumption is the key factor behind this type, as adipocytes increase in volume in response to the excess nutrient load. This type is highly related to the development of glucose intolerance and insulin resistance²⁹. The ability of medicinal plants to inhibit LPL was suggested as a new mechanism to reduce weight since LPL and its tissue specific regulation have been thought to be centrally involved in the pathogenesis of obesity³⁰. LPL activity has a been shown to be markedly elevated in the adipose tissue of obese subjects³¹.

Medicinal plants represent potential candidates as LPL inhibitors. The extract of Onosma giganteum Lam. showed the highest inhibition percentage of LPL activity (32.92 %) among the 20 tested plants. This inhibition can be related to the chemical constituents of this plant. It was reported that Onosma L. contains flavonoids and some phenolic compounds³². According to various studies, such compounds have a proven effect on lipid metabolism process. The methanolic extracts of Hypecoum dimidiatum Delile and Chrysanthemum coronarium have shown modest LPL inhibitory effect of 29.03% and 27.81% respectively. In a previous study, H. dimidiatum showed 25% inhibition of hormone sensitive lipase (HSL) and showed no activity against PL³³. The LPL inhibitory effect of C. coronarium L. can be added to its demonstrated inhibitory effect against human cholesterol acyl transferase and LDL-oxidation activity³⁴. Inhibitory effect of C. coronarium L. against HSL (49.7%) and PL were also reported²².

CONCLUSION:

Medicinal plants represent potential candidates to be implemented in new therapeutic era. 11 plants out of 20 tested plants have shown LPL inhibition activity, for a certain extent. However, further studies need to be carried to investigate their potential activity in vivo in order to develop new anti-obesity treatment.

ACKNOWLEDGEMENTS:

The authors wish to thank the Deanship of Scientific Research at the University of Jordan for their generous funds. This article was accomplished during a sabbatical leave of Dr. Bustanji

A CONFLICT OF INTEREST OR DISCLOSURE STATEMENT:

We wish to confirm that there are no known conflicts of interest associated with this publication and there has been no significant financial support for this work that could have influenced its outcome.

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Received on 29.10.2022 Modified on 24.02.2023

Research J. Pharm. and Tech 2023; 16(10):4786-4790.

DOI: 10.52711/0974-360X.2023.00776