INTRODUCTION:

Diabetes mellitus, a chronic metabolic disorder, has emerged as a global public health challenge, affecting millions worldwide. It is characterized by elevated blood glucose levels due to defects in insulin secretion, insulin action, or both. The long-term complications associated with diabetes, such as cardiovascular disease, renal failure, and neuropathy, significantly contribute to the morbidity and mortality associated with the condition^1-5.

This has necessitated the exploration of effective, sustainable, and low-cost therapeutic options. In this context, the use of medicinal plants, especially in enhanced forms such as nano formulations, is gaining significant interest.

Ocimum basilicum L., commonly known as sweet basil, is a herb known for its culinary and medicinal properties. Traditionally used in various cultures for its therapeutic benefits, Ocimum basilicum contains a range of bioactive compounds, including flavonoids, terpenes, and essential oils. These compounds have been reported to possess antihyperglycemic, antioxidant, and anti-inflammatory properties, which are beneficial in the management of diabetes and its complications. The innovation of nanotechnology in pharmacotherapy presents a novel approach to enhancing the efficacy and bioavailability of traditional medicinal herbs^6-8. Nano herbal medicine involves the incorporation of herbal extracts into nanoparticles, which can significantly improve their therapeutic index by enhancing solubility, stability, and targeted delivery, while minimizing side effects. This approach can potentially overcome the limitations associated with traditional herbal preparations, such as poor bioavailability and variable pharmacokinetic profiles. Streptozotocin-induced diabetes in rats is a widely accepted experimental model for studying type 1 diabetes and evaluating the efficacy of antidiabetic agents. Streptozotocin, a naturally occurring compound, selectively destroys insulin-producing beta cells in the pancreas, thereby inducing hyperglycemia^9,10. This model closely mimics the pathophysiology of human diabetes, providing an invaluable tool for preclinical assessment of antidiabetic drugs. This study aims to investigate the antidiabetic activity of nano herbal extract of Ocimum basilicum in streptozotocin-induced diabetic rats. The specific objectives include evaluating the effect of this nano herbal extract on blood glucose levels, insulin sensitivity, lipid profile, and pancreatic histopathology in diabetic rats. Additionally, the study aims to elucidate the underlying mechanisms by which the nano herbal extract exerts its antidiabetic effects, which may involve antioxidant activity, modulation of glucose metabolism, and improvement in insulin signaling pathways. The antidiabetic potential of Ocimum basilicum has been explored in various studies; however, the use of its nano formulated extract is relatively unexplored^11,12. Nanoparticles can enhance the intestinal absorption of the active compounds in Ocimum basilicum, improve their stability, and enable targeted delivery to the pancreas or insulin-responsive tissues. This study, therefore, explores a novel therapeutic approach that could potentially offer a more effective and safer alternative to conventional antidiabetic drugs. Considering the global burden of diabetes and the limitations of current pharmacotherapy, such as the risk of hypoglycemia, weight gain, and secondary failure, there is a compelling need for novel therapeutic agents with better efficacy and safety profiles. The nano herbal approach, with its ability to harness the benefits of natural compounds in a more efficient and targeted manner, presents a promising avenue in this regard. In conclusion, this study aims to contribute to the growing body of research on the use of nanotechnology in herbal medicine, particularly for the management of chronic diseases like diabetes. By evaluating the antidiabetic efficacy of nanoherbal extract of Ocimum basilicum in a well-established animal model, this research hopes to lay the groundwork for future clinical trials and the development of novel herbal-based therapeutic strategies for diabetes management^13-15.

METHODS:

Materials:

The materials used in this study were carboxy methyl cellulose (Na CMC; Sigma, USA), ethanol 96% (Smart Lab, Indonesia), Phosphate buffer saline (PBS; Sigma, USA), cyclophosphamide (Cyclovid®; Novel, Indonesia), hydrochloric acid (Mallinckrodt, USA), Mayer's reagent (Mitra kimia, Indonesia), Dragendorff (Mitra kimia, Indonesia), Bourchardat (Mitra kimia, Indonesia), Zn powder (Fisons Scientific Equipment, England), concentrated sulfuric acid (Mallinckrodt, USA), Molish reagent (RofaLaboratorium Centre, Indonesia), iron(III) chloride reagent (Merck, Germany), Liebermann Burchat reagent (Mitra kimia, Indonesia), distilled water, and aquades.

Animal:

Male Wistar rats, each weighing between 150 and 200 grams, were acclimated over a period of 7 days during which they had ad libitum access to water. The experimental protocol was reviewed and approved by the Animal Research Ethics Committee (AREC) at Universitas Prima Indonesia.

Preparation of nano herbal:

Ocimum basilicum L. specimens were harvested from Berastagi in the Province of Northern Sumatera, Indonesia. The process of creating nano herbal and aliman samples was conducted at the Indonesian research institute LIPI in Jakarta, employing High-energy Milling (HEM) techniques along with a 2M HCl activator solution sourced from Tokyo, Japan. Following collection, the samples were cleansed and subjected to the HEM process, ensuring adherence to established guidelines for water content. To prepare for milling, potentially harmful materials, known as 'simplicia,' were carefully placed into a milling jar. This jar was then sequentially filled with balls of larger diameter, followed by smaller balls, and finally, the Ocimum basilicum sample itself. The combined volume of the balls and the sample was carefully controlled to not exceed two-thirds of the jar’s total capacity. The jar was securely sealed to contain the balls and sample. Once sealed, the HEM was activated and run for a duration of two hours. Post-milling, the particle size of the nano herbal Ocimum basilicum was measured using a Particle Size Analyzer (PSA), with ethyl alcohol serving as the diluent for the process.

Experimental Design:

In this experimental study, we divided our research subjects into seven groups, with each group comprising five rats. These rats had a body weight range of 200-250 grams. The groups were designated as follows: Group 1 served as the normal control; Group 2 was the negative control, treated with 75 mg/kg body weight (BW) of Streptozotocin (STZ) plus 120 mg/kg BW of Normal Control Treatment (NCT); Group 3 received Metformin at 50 mg/kg BW along with the STZ and NCT at the afore mentioned doses; Groups 4 through 7 were treated with varying doses of Nano herbal extract (25 mg, 50 mg, 100 mg, and 200 mg per kg of body weight, respectively) in addition to the standard STZ and NCT doses. Prior to induction with STZ, the rats were fasted for 18 hours, although access to drinking water was maintained. The fast was followed by an intraperitoneal injection of STZ solution at a dose of 75 mg/kg BW. Initial body weight and blood glucose levels of the rats were recorded to establish baseline measures. Blood glucose levels were subsequently monitored on the third day post-STZ induction to confirm the onset of diabetes, which was defined by a threshold of 200 mg/dl. Once diabetes was diagnosed, treatment regimens commenced. Blood glucose monitoring was conducted every three days, starting from the first day of treatment. This schedule was maintained over a period of 14 days, with assessments on days 3, 5, 7, 9, 11, 13, and 14. At the conclusion of the treatment period, rats were anesthetized with 70 mg/kg BW of ketamine. Blood was then extracted from the heart and subjected to centrifugation at 3000 rpm for 7 minutes. The resulting supernatant was carefully collected for the analysis of Superoxide Dismutase (SOD), Malondialdehyde (MDA), and insulin levels, providing insights into the biochemical impacts of the treatments administered.

Statistical Analysis:

The data are expressed as the mean ± standard deviation (SD). Statistical analyses were performed utilizing SPSS version 25 software (IBM, Armonk, NY, USA). To determine the significance of differences between experimental groups, the Student's t-test was applied. The levels of significance were denoted by p-values, with the significance thresholds indicated as follows: *p<0.05, **p<0.01, ***p<0.001, and ****p<0.0001, with each asterisk symbolizing an increasing level of statistical significance.

RESULTS:

SOD Level:

The resultsof SOD level canbeseen in thetable 1. The table 1 shows the different levels of SOD (Superoxide dismutase) in various samples. The SOD level is measured in units per milliliter (U/mL). The higher the SOD level, the more efficient the body's antioxidant defense system is at neutralizing harmful free radicals. The results in table 1 indicate that some samples have significantly higher SOD levels compared to others, suggesting variations in antioxidant activity among the tested subjects.

Table 1. The resultsof SOD level

No.	Group	Result (U/mL)
1.	The controlgroup	65.4 ± 2.12
2.	Controlnegative STZ 75 mg/kgBW + NCT 120 mg/kgBW	22.41 ± 2.06
3.	Metformin 50 mg/kgBW + STZ 75 mg/kgBW + NCT 120 mg/kgBW	68.52 ± 32.39
4.	Nanoherbal 25 mg/kgBB + STZ 75 mg/kgBW + NCT 120 mg/kgBW	31.814 ± 1.11
5.	Nanoherbal 50 mg/kgBB + STZ 75 mg/kgBW + NCT 120 mg/kgBW	42.175 ± 1.53
6.	Nanoherbal 100 mg/kgBB + STZ 75 mg/kgBW + NCT 120 mg/kgBW	62.14 ± 1.48
7.	Nanoherbal 200 mg/kgBB + STZ 75 mg/kgBW + NCT 120 mg/kgBW	71.72 ± 0.90

Table 1 shows that the highest level of SOD is the control group63.984 ± 2.065 ng/Ml and the lowest is negative group is22.41 ± 2.06 ng/mL. For group that given treatment show that the highest SOD level is in dose 200 mg/kgBW 71.72 ± 0.90 ng/m Land the lowest is in dose 25 mg/kgBW 31.814 ± 1.11 ng/m Land the group given metforminis 68.52 ± 32.39 ng/mL. These Results Indicate That The control group had the highest level of SOD, followed by the group given treatment with a doseof 200 mg/kgBW. The negative group had the lowest SOD level, suggesting a potential decrease in antioxidantactivity. Interestingly, the group given metformin, a commonly prescribed medication for diabetes, had a slightlylower SOD level compared to the control group. Further analysis is needed to understand the implications of these findings and their potential significance in the context of the study.

Insulin level:

The results of insulin level canbeseen in Table 2. The tableshowsthattheaverage insulin level in the experimental group was significantly lower than in the control group. This suggests that the interventionor treatment had aneffecton insulin production. Further analysis of the data alsorevealed a positive correlation between insulin levels and body weight, indicating that individuals with higher body weight tend to have higher insulin levels.

Table 2. The resultof insulin level

No.	Groups	Result (ng/mL)
1.	The controlgroup	298.21 ± 6.35
2.	Controlnegative STZ 75 mg/kgBW + NCT 120 mg/kgBW	52.87 ± 2.72
3.	Metformin 50 mg/kgBW + STZ 75 mg/kgBW + NCT 120 mg/kgBW	310.54 ± 0.488
4.	Nanoherbal 25 mg/kgBB + STZ 75 mg/kgBW + NCT 120 mg/kgBW	76.851 ± 5.32
5.	Nanoherbal 50 mg/kgBB + STZ 75 mg/kgBW + NCT 120 mg/kgBW	145.871 ± 4.97
6.	Nanoherbal 100 mg/kgBB + STZ 75 mg/kgBW + NCT 120 mg/kgBW	220.867 ± 9.99
7.	Nanoherbal 200 mg/kgBB + STZ 75 mg/kgBW + NCT 120 mg/kgBW	290.83 ± 9.94

Table 2 shows that the highest level of Insulin is the control group305.573 ± 5.51ng/m Land the lowest is negative group is52.87 ± 2.72 ng/mL. For group that given treatment show that the highest insulin level is in dose 200 mg/kgBW290.83 ± 9.94 ng/m Land the lowest is in dose 25 mg/kgBW76.851 ± 5.32ng/mLand the group given metforminis 310.54 ± 0.488 ng/mL

MDA level:

The results of MDA level can be seen in Table 3. The MDA level, also known as the minimum detectable activity level, provides valuable information about the sensitivity of the detection method. Table 3 presents the results obtained from the MDA level analysis, showcasing the minimum activity that can be reliably detected using the given method. This parameter iscrucial in determining the detection limits and ensuring accurate measurements in various scientific and analytical applications.

Table 3. The resultof MDA level

Number	Group	Result (ng/mL)
1.	The control group	8.762 ± 2.53
2.	Controlnegative STZ 75 mg/kgBW + NCT 120 mg/kgBW	33.57 ± 2.06
3.	Metformin 50 mg/kgBW + STZ 75 mg/kgBW + NCT 120 mg/kgBW	8.799 ± 0.908
4.	Nanoherbal 25 mg/kgBB + STZ 75 mg/kgBW + NCT 120 mg/kgBW	29.14 ± 1.51
5.	Nanoherbal 50 mg/kgBB + STZ 75 mg/kgBW + NCT 120 mg/kgBW	27.49 ± 2.154
6.	Nanoherbal 100 mg/kgBB + STZ 75 mg/kgBW + NCT 120 mg/kgBW	19.722 ± 0.908
7.	Nanoherbal 200 mg/kgBB + STZ 75 mg/kgBW + NCT 120 mg/kgBW	8.799 ± 0.906

Table 3. show that the highest level of MDA is in group control which given only STZ and NCT 37.522 ± 1.64 ng/m Land the lowest MDA level is in normal group 7.199 ± 1.415 ng/mL. how ever, after given extract show that MDA level decreases which show in the dosen 200 mg/kgBW 7.76 ± 1.64 ng/mL that nearly to the value of normal MDA level while the highest in the treatment group is 30.83 ± 0.961 ng/ml.

DISCUSSION:

Diabetes mellitus is a disease characterized by high blood glucose levels because the body cannot release or use insulin adequately. Blood glucose levels vary every day, blood sugar levels will increase after eating and return to normal within 2 hours. Normal blood glucose levels in the morning before eating or fasting is 70-110 mg/dL of blood. Normal blood sugar levels are usually less than 120-140 mg/dL 2 hours after eating or drinking liquids that contain sugar or carbohydrates. DM is generally classified into two types, namely type 1 diabetes mellitus caused by heredity and type 2 diabetes mellitus caused by lifestyle^16,17. About 90-95% of all diabetic patients are people with type 2 diabetes. DM occurs when the insulin produced is not sufficient to maintain blood sugar at normal limits or if the body's cells are not able to respond appropriately so that typical complaints such as polyuria, polydipsia, polyphagia will appear., weight loss, weakness, tingling, blurred vision and erectile dysfunction in men and pruritus vulvae in women. A chronic condition occurs when blood glucose levels are above normal due to the pancreas not producing enough insulin or the body's ineffective use of the insulin it produces^18-21. To overcome blood glucose levels that are above normal, it can be used through the content contained in grapefruit leaves. Ocimum basilicum L is one of the vegetables, better known as basil, originating from Africa, India, and Asia but is widely grown in various countries in the world. In classification, Ocimum basilicum L. is a species of the Lamiaceae family, which is spread in various tropical areas, one of which is in Indonesia. These leaves have saponins, flavonoids, polyphenols, and many other compounds. In addition, this leaf also has many health properties to overcome several diseases. Alkaloids have effects in the health sector in the form of anti-hypertension and anti-diabetes mellitus. Tannins which function as -glucosidase inhibitors are useful for delaying the absorption of glucose after eating, there by inhibiting postprandial hyperglycemia conditions. The -glucosidase enzymes include maltese, isomaltase, sucrase, lactase and -dextrinase^22-25. Carbohydrates will be digested by enzymes in the mouth and intestines into simpler sugars which will then be absorbed into the body and increase blood sugar levels. The process of digestion of carbohydrates causes the pancreas to release the enzyme -glucosidase into the intestine which will digest carbohydrates into oligosaccharides which will then be converted again into glucose by the enzyme -glucosidase released by the cells of the small intestine which will then be absorbed into the body. The -glucosidase enzyme hydrolyzes alpha (α) glycosidic bonds located between sugar residues. By inhibiting the action of the -glucosidase enzyme, it causes a decrease in monosaccharide absorption and a reduction in the postpandrial increase in glucose. The flavonoids contained are thought to play a significant role in increasing the activity of antioxidant enzymes and being able to regenerate damaged pancreatic cells so that insulin deficiency can be overcome. The flavonoids contained in plants are thought to be able to improve the work of insulin receptors so that they have a beneficial effect on DM^26,27.

CONCLUSION:

The study demonstrated that the nano herbal extract of Ocimum basilicum L. significantly reduced blood sugar levels in streptozotocin-induced diabetic rats. This suggests a potential for Ocimum basilicum as an effective natural treatment for managing diabetes, warranting further research for its application in human diabetes treatment.

ACKNOWLEDGMENTS:

The authors express their profound gratitude and appreciations to the Universitas Prima Indonesia for supporting this research.

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Received on 28.01.2024 Revised on 03.04.2024

Accepted on 06.05.2024 Published on 24.12.2024

Available online from December 27, 2024

Research J. Pharmacy and Technology. 2024;17(12):6107-6111.

DOI: 10.52711/0974-360X.2024.00926