Evaluation of skin absorption of the Citrullus colocynthis in treatment of type II diabetic patients

Abstract

Background and purpose

Nowadays, among the herbal medicines utilized to treat diabetes, Citrullus colocynthis (CCT) is highly noticeable as it reduces blood glucose (BG) and stimulating insulin secretion. However, long-term oral consumption of this herbal medicine has often associated with digestive complications. In this study, skin absorption of CCT as a new therapeutic approach in the treatment of type II diabetic patients has been surveyed.

Materials and methods

40 patients with type II diabetic (aged 45–65) were selected. Participants were asked for placing their metatarsus daily in a decoction containing 2% CCT solution for 40–60 min each day and continuing that for 10 days. Blood and urine samples of patients collected at the beginning and the end of the study. The samples were examined for the BG levels, serum insulin content, lipid profiles, hepatic enzymes, urea, creatinine, and microalbuminuria, The quantitative insulin sensitivity check index (QUICKI), Homeostatic Model Assessment of Insulin Resistance (HOMA-IR), homeostasis model assessment of β-cell function (HOMA-β) and disposition index (DI) indicators were also calculated.

Results

Local treatment of CCT could significantly decrease BG levels, stimulate insulin secretion and improve the function of pancreatic beta cells. It also decreased serum urea levels comparing to pre-treatment levels (p < 0.05) but there was no significant change in creatinine levels, lipid profiles, hepatic enzymes, micro-albuminuria, and other insulin sensitivity indexes.

Conclusion

This study demonstrated that the CCT plant can also have systemic therapeutic effects on type II diabetic patients through dermal absorption.

Introduction

Diabetes mellitus is the most common endocrine disorder associated with impaired carbohydrate metabolism, which also disturbs fat and protein metabolism in chronic conditions. Diabetes mellitus affects around 200 million people worldwide [1, 2]. Due to microvascular (retinopathy, neuropathy, and nephropathy) and macrovascular (heart attack, stroke, and peripheral vascular disease) side effects, this disease has caused significant difficulties and mortality [3].

Recently, considerable studies are being conducted to find safe and effective drugs that can help to protect diabetic patients from other complications of diabetes. In this regard, the WHO has recommended the use of herbal medicines [4]. However, there is a long history of using plants to treat diabetes [2].

“Citrullus colocynthis” is a member of the Cucurbitaceae family that is known as “bitter apple” in the English language, “Hindal” in Arabic, and “hendewane abujahl” Persian [5]. CCT is a grapevine-like plant that is found mainly in Mediterranean Europe, Asia, Turkey, Nubia, Trieste, Egypt, Iran, Pakistan, Afghanistan, India and North Africa [6]. Researches have shown that the use of CCT extract leads to a reduction in glycosylated hemoglobin, The regulation of fasting blood glucose levels, and induction of insulin secretion [7]. It has antioxidant effects and daily intake of 300 mg/kg of CCT seed powder results in a significant reduction in cholesterol and triglyceride levels in hyperlipidemia patients [8]. However, some studies on laboratory animals demonstrated that daily intake of 100 mg/kg of this plant’s pulp extract can cause severe damages to the small intestine, kidney, and liver [5]. Concentrations over 400 mg/kg cause digestive and liver disorders in humans, and in several studies, this concentration level has been reported as toxic concentration [9].

Over the past two decades, the skin has become particularly important for local, regional and systemic use of medications. Human skin is an effective barrier that does allow external particles to come into contact with internal tissues. Consequently, the dermal absorption of the therapeutic agents is not efficient [10]. Currently, fatty acids are considered as permeation multipliers of subcutaneous and skin absorption [11]. Myristic, lauric, oleic, linoleic, and linolenic acids are fatty acids that increase skin permeation and usually use as an efficient absorption enhancer. Indeed, these formulations are suitable for the development of systemic absorption of local medicines [12].

There are several bioactive compounds of CCT fruit registered in the papers which are classified as glycosides, flavonoids, alkaloids, carbohydrates, fatty acids, and essential oils. Studies have shown that compounds including glycosides, flavonoids, and alkaloids play a major role in the anti-diabetic effects of this plant [2, 6, 13, 14]. On the other hand, the association of these compounds with fatty acids such as stearic, myristic, palmitic, oleic, linoleic, linolenic acids in fruit CCT [15], established the hypothesis that whether local utilization of this plant could have anti-diabetes impacts on type 2 diabetic patients. Interestingly, CCT is being used for a long time in traditional medicine for its anti-diabetic effects, but no research has been done to support this method for treating diabetes with modern medical parameters. Thus, the present study aims to explore the potential of topical consumption of CCT fruit products for the treatment of type 2 diabetes.

Materials and methods

The fresh CCT fruit was collected from southern Iran (Hormozgan) and was confirmed by the botanist at the Herbarium Center of Jundishapur University of Medical Sciences. After drying in the shade and temperature of <40 °C, it was prepared as powder by a mechanical grinder.

40 patients with type II diabetes (aged 45–65), including both genders, were randomly selected referred to the Diabetes Clinic of Golestan Hospital in Ahvaz. All of the patients who participate in this study were diagnosed with type II diabetes by the observer physician at the clinic. Patients selected with following conditions: 1) patients who had not used herbal therapy over the past two months 2) patients who had taken only one or two types of antidiabetic drugs 3) patients with fasting glucose level greater than 140 mg/dl 4) patients who were suffering from diabetes for 1–8 years and had normal blood pressure. The patients who had insulin treatments or complications such as cardiovascular disease, Infectious disease, pregnancy, lactation, and cancer were excluded from the study. Patients were interviewed by the researchers and informed about the goals and conditions of the study. An approval form was completed for each participant, under the supervision of the Jundishapur Medical University’s ethical Committee (code: IR.AJUMS.REC.1397.420).

At first, a 5 ml fasting venous blood sample and random urine sample were taken from all participants. The height and weight of individuals were also measured to determine the body mass index (BMI). Participants were asked to do the procedure for 10 days, daily after breakfast, decoct a pack of CCT powder (20 ± 2 g in 1 l of water) to prepare 2% w / v solution and then place their metatarsus in it for 40 to 60 min until they taste its bitterness in their mouths, as we had found by doing the pilot. It is recommended that if they take the antidiabetic drug, continue the drug as before. Finally, blood and urine samples of the patients were collected and all tests were repeated. Urine samples were analyzed for presence of microalbuminuria and venous blood samples, after centrifugation and immediately after serum separation, were evaluated for blood glucose, total cholesterol, triglyceride, LDL, HDL, aspartate transaminase (AST), alanine transaminase (ALT), alkaline phosphatase (ALP), urea and creatinine levels by an autoanalyzer using available commercial kits (Pars Azmoon, Iran) at the Jundishapur University of Medical Sciences Diabetes Research Center. ELISA method (Monobind Inc., USA, within assay precision 5.1%, sensitivity 0.75μIU/ml) was used to measure serum insulin content. Various calculations such as QUICKI, HOMA-IR, HOMA-β, DI were also performed using the following formulas:

HOMA-IR fasting blood glucose (FBS) (mg/dL) × insulin (μIU/mL)/405.

HOMA-β 20 × insulin (μIU/mL)/ (FBS (mMol/L) - 3.5).

QUICKI 1/ (log FBS (mg/dL) + log insulin (μIU/mL)) [16].

DI HOMA-β/HOMA-IR assayed [17].

Statistical analysis of the data obtained at the beginning and the end of the study was conducted using SPSS software, paired t-test. The data resulted from the study are expressed as Mean ± SEM and the significance level was considered as p < 0.05.

Results

Participants of this study included 33.3% male and 66.7% female. The demographic data obtained from participants (Table 1) shows that the average age of subjects and type 2 diabetes suffering duration were 54.1 ± 1.9 and 5 ± 0.49 years, respectively. They also had a weighted average of 71.76 ± 2.3 Kg and a BMI of 25.6 ± 0.87 m²/Kg. Blood and urine samples were collected twice from all participants of the study. The first sample collection was performed one day before the beginning of the study and the second one was done at the end of the study, after local and daily use of CCT powder for ten days. The two groups of collected samples were evaluated for fasting blood glucose levels, insulin, QUICKI, HOMA-IR, HOMA-β and DI. The results of this evaluation are summarized in Table 2. The average level of fasting blood glucose in these patients was 255.11 ± 21 mg/dl. After local treatment with CCT, it was significantly decreased to 233.57 ± 19 mg/dl (p < 0.05). The average insulin level also was increased dramatically from 6.99 ± 1.1 to 11.80 ± 1.9 μIU/L. (p < 0.05). The evaluation of HOMA-β showed a significant increase in pancreatic beta-cell function, from a base value of 93.67 ± 18.8 to 150.23 ± 22.2 (p < 0.05). However, the QUICKI, HOMA-IR and DI indices did not express any significant changes comparing to pre-treatment values (p > 0.05).

Table 1.

Demographic characteristics of 40 type II diabetic patients participating in the study, which are demonstrated as average age, duration of disease, weight, and BMI

Demographic characteristics	Mean±SEM
Age(year)	54.1±1.9
Duration of disease(year)	5±0.49
Weight (Kg)	71.76±2.3
BMI (m2/Kg)	25.6±0.87

Table 2.

Comparison of blood glucose (mg/dl), fasting insulin level (μIU/L) and insulin sensitivity index average (QUICKI, HOMA-IR, HOMA-β, and DI) before and after local treatment with CCT in 40 patients with type II diabetes

	Before	After
Variable	Mean±SEM	Mean±SEM	P Value
BG (mg/dl)	255.11±21.5	233.57±19.2	0.023
Insulin(μIU/L)	6.99±1.1	11.80±1.9	0.031
QUICKI	0.35±0.02	0.3±0.007	0.095
HOMA-IR	3.91±0.61	6.39±1.1	0.054
HOMA-β	93.67±18.8	150.43±22.2	0.036
DI	26.71±5.9	28.09±4.3	0.683

The data resulted from the evaluation of lipid profiles, liver, and renal function indices are summarized in Table 3. Among the surveyed renal function indexes, serum urea average level altered from 25.92 ± 1.8 to 23.34 ± 1.7 mg/dl, which shows a significant decrease (p < 0.05). However, there were no considerable changes in serum creatinine and urine albumin levels. As shown in Table 3, there is no significant change in any of the lipid and liver profiles.

Table 3.

Comparison of average renal function indexes, lipid profile and hepatic enzymes levels in 40 patients with type II diabetes before and after local treatment with CCT

	Before	After
Variable	Mean±SEM	Mean±SEM	P Value
Urea(mg/dl)	25.92±1.8	23.34±1.7	0.027
Creatinine(mg/dl)	0.846±0.3	0.824±0.4	0.0552
Albumin(μg/ml)	200±33.1	181.8±26.4	0.604
Triglyceride(mg/dl)	203±34.7	227.33±34.2	0.086
Cholesterol(mg/dl)	147.95±8.1	154.23±8.2	0.190
HDL(mg/dl)	45±2	45±1.7	1.000
LDL(mg/dl)	68.90±5.8	68.85±5.4	0.992
AST(U/L)	21.57±2.3	21.52±1.8	0.979
ALT(U/L)	25.28±4.7	27.52±5.1	0.313
ALP(U/L)	250.23±15.3	256.42±14.6	0.155

Discussion

Many experimental and clinical studies have supported the beneficial effects of CCT in reducing fasting blood glucose levels and stimulating serum insulin secretion [18]. What makes this study notable is the presentation of a new therapeutic approach to develop the desired systemic effect using a high potential skin absorption. The present study truly showed that local consumption of CCT can significantly decrease fasting blood glucose levels and stimulate insulin secretion in type 2 diabetic patients. On the other hand, 1.6x improvement of beta-cell function in the HOMA-β homeostatic model can be considered as the strong point of this new therapeutic approach, which is unprecedented in its kind. Previous studies have shown that CCT antidiabetic impacts occur through the preservation and regeneration of pancreatic Beta cells in animal models, and CCT, such as glibenclamide, can stimulate insulin secretion [2, 13]. According to data resulted from this study, it can be claimed that the association of fatty acids with anti-diabetic compounds of CCT has allowed skin absorption and efficacy of this plant. In this survey, no significant change in the QUICKI, HOMA-IR, and DI indices was observed compared to the pre-treatment values. However, insulin sensitivity indexes which are estimated based on glucose and insulin levels should be used carefully when compared in populations with different genders and races, since in some races, there is no correlation between fasting insulin levels and HOMA-IR with glucose disposal rate (GDR) [19]. Regarding the fact that the present study has been conducted among both genders and Arab and Fars races in Ahwaz, Khuzestan province, the correlation between insulin sensitivity indexes, fasting glucose and insulin levels requires further studies. Investigating the results of renal function indices showed that local and daily consumption of 2% CCT solution for 10 days could significantly decrease urea levels in patients with type II diabetes. Although, there is no considerable change in serum creatinine and urine albumin levels compared to pre-treatment ones. Histological studies performed by other researchers of experimental fields demonstrated that oral consumption of the CCT fruit extract has nephroprotective effects and significantly reduces the levels of urea, creatinine, and albumin. These impacts occur by affecting the function of kidney tissue [20]. Comparison of liver enzymes including AST, ALT, and ALP before and after treatment with CCT showed no significant change in any of the liver enzymes levels. Earlier, Hosseini and colleagues reported in a study on type II diabetic patients that prescribing oral use of 300 mg CCT extract over 2 months did not dramatically change the activity of AST, ALT, and ALP enzymes. so, it can be said that the CCT extract did not cause liver damage [7]. Although, the dose-dependent effects of water and alcoholic CCT extracts on liver toxicity have been reported in experimental studies, and oral consumption of concentrations greater than 400 mg/kg leads to an increase in liver enzymes in adult rats [21–23]. In contrast, some other studies have reported that CCT has hepatoprotective effects which probably is accomplished through increased antioxidant defense systems [24]. In spite of many studies emphasizing the decreasing effect of watery and alcoholic CCT extracts on the serum cholesterol, triglycerides, HDL, and LDL levels in diabetic animal models [25], clinical studies showed controversial results. In one study, Youshan Li and colleagues demonstrated that daily consumption of 1 g capsules containing CCT powder for one month did not significantly alter cholesterol, triglyceride, HDL and LDL levels in type II diabetic patients [26]. Interestingly, we showed the same results in which, lipid profiles did not dramatically change after local use of CCT. However, the study by the Rahbar and his colleagues on nondiabetic hyperlipidemia patients suggested that daily intake of 300 mg of CCT powder for 6 weeks caused a significant decrease in triglyceride and cholesterol levels [27]. However, hypertriglyceridemia and reduced HDL are the most common lipid disorders in type II diabetic patients that remain even after proper glycemic control [28]. Therefore, it seems that the therapeutic effect of CCT on reducing serum glucose levels may not be associated with a reduction of serum lipids in diabetic patients whose mechanism needs further investigation. One of the main problems of this study was to supply the plant because of using 200 g for each patient, and this limitation led to reduce the number of patients and the duration of the treatment. HbA1c evaluation needs three months to give a valuable result, so this protocol wasn’t able to show changes in HbA1c. Finally, it would be better to use the control group as a comparison with the treatment group and the results would improve by increasing the number of patients and duration of treatment.

Conclusion

According to the results of the current study on 40 patients with type II diabetes, local consumption of the Citrullus colocynthis powder for 10 days, was able to develop systemic therapeutic effects of this plant including reduction in fasting blood glucose levels, stimulation of serum insulin secretion, improvement of pancreatic beta-cell function and reduction in serum urea levels. Although it requires further studies to understand the effects of topical use of this plant on other organs and to determine the precise dose in this type of treatment, this new therapeutic approach is a promising way to cure diabetes.

Compliance with ethical standards

Conflict of interest

The authors certify that they have no affiliations with or involvement in any organization or entity with any financial interest or non-financial interest in the subject matter or materials discussed in this manuscript.