Diabet is an important of metabolic disorders with an increasing incidence throughout the world (1). It is broadly classified into three types such as type 1, 2 and gestational diabetes thet they have different etiology and clinical presentation (2). Diabetes mellitus is one of the chronic and metabolic disease with hyperglycemia persistentence. In type 2 diabetes mellitus, the response to insulin is diminished as an insulin resistance (3). When insulin is ineffective, it is initially countered by an increase in its production to maintain glucose homeostasis, but over time, insulin content decreases, and lead diabetes mellitus type 2 (Nmila et al., 2000). Diabetes mellitus is most commonly observed in 45 years old and older person (4). Patients with type 2diabetes mellitus in Iran have been used the Persian traditional medicine highly. In simmilary with other traditional medical approaches, a special type applies of diagnostic model for patients’ individualization, known as a differentiation syndrome in the literature were applied. Based on a concept which is called Mizaj the syndromes in Persian traditional medicine are defined also (5). According to that, each person has a two spectra of warm and cold and wet and dry qualities of Mizaj and any treatments in Persian traditional medicine must have be selected based on this theory. Hence, different treatments may be aimed for the same diseases such as type 2 diabetes mellitus based on the person^,s Mizaj. The Colocynth (Citrullus colocynthis Schrad L) is  a fruit from the family Cucurbitaceae. It is also a known as a medicinal friut which used for some purposes in medicine (6). The glycosides, flavonoids, alkaloids, fatty acids and some essential oils are some of bioactive constituents in colocynth (7). Colocynth has been  used for treatment of diabetes and some parasitic infections (8). Saponine is known as the main component of involved with the blood glucose lowering effects of colocynth (9) and it is the hypoglycemic properties of colocynth (10). The colocynth oil of may have significant decreasion in the blood glucose levels of  experimental diabetic rats (11). The result of (12) study showed that  application of colocynth fruit has a benficial acts on reduction in the serum level of glycated hemoglobin and fasting blood glucose in the type 2 diabetes patients. The blood glucose lowering action of colocynth has mentioned by (13) in perivious studies. According to our knowledge there were no more clinical studies were done on colocynth fruit effectiveness or side effects among patients  with diabetes mellitus type 2 yet. Recently the traditional Persian medicine syndrome differentiation model was used to identify the patients who will benefit more from colocynth extract supplementation but there was not reported in previous studies. Additionally there were no previous founding  are available on Persian traditional medicine syndrome differentiation basis on women with diabetes mellitus type 2, Hence the present study was performed to found the beneficial acts of colocynth extarct intervention for women with diabetes mellitus type 2 basis on Persian syndrome differentiation model traditional medicine.

To run the test a randomized, 3-blinded, clinical trial controlled satudy and the computergenerated block randomization lists used a one by one allocation ratio and block size of four were used.  All the diabetes mellitus type 2 women were invited to the Iranian Medical univeristy from May to November 2022. All women signed an informed consent form after the research method was explained and were selected by eligibility criteria. The women demographic and clinical features were recorded and according to the International Physical Activity Questionnaire and Food Frequency questionnaire their physical activities and dietary intakes were recorded and registered and their blood samples were takenin basis. The women, aged 20-40 years old, and diagnosed with diabetes mellitus type 2 by an endocrinologist and a fasting blood glucose level of 130 to 240 mg/dl. They also used only oral hypoglycemic agents for their diabetes, and they did not have any uncontrolled diseases. The women who have any allergic reactions to colocynth ethanolic extract supplementation were consuming less than 75 percentage of the prescribed capsules.They completed the ten item of Mizaj questionnaires aqccording to the (14) described method. Researcher were asked the questions and the results were recorded in the pre-designed forms. The first and domains of the questions were related to the warmness and wetness dimensions of Mizaj. The total scores of ≤14,15-18, and ≥19 indicated as a cold, moderate, and warm Mizaj, and those of ≤ 3, 4, and ≥ 5 were considered as a wet, moderate, and dry Mizaj conditions. The analysis ofpurchased colocynth was carried out using a GC/MASS. The Clarus 580 chromatography apparatus equipped with a capillary column  and coupled to a mass spectrometer  was used in this assay. The GC/MASS reproted that it contains seventeen compounds were broadly identified and divided into five classes viz., alcohols, ketones, epoxy compounds, hydrocarbons and an acid. The colocynth ethanoic extract was filled into the capsules with 250 mg  of capacity. The placebo capsules as a control tretamnets were filled by 250 mg of heated starch. The supplemental capsules were administered to the women after fasting overnight in the morning and before going to bed at night for three months. The supplemented capsules were delivered to the women, and follow-up  were done by phone call every two weeks, whether they were taking the drugs regularly and if they were experiencing side effects until first month. After that, only women who used more than 75 percentage of the colocynth ethanoic extract supplemented capsules were treated using the same procedure for another two months. Upon completing the three months of treatment, the women^,s biochemical blood markers were re assessed.

Blood biochemical measurements

The fasting plasma glucose content were measured by taking 5 ml of participant blood after twelve hours of fasting by glucose peroxidase method using an Biochemistry Company made in Iran assay kits and were anaylazed by Echo Plus auto-analyzer device made in Italy.The glycated hemoglobin assay was done after its primary separation via the ion-exchange chromatographic method. Additionally, the insulin level and resistance were determined using ELISA kits (Pars Azmoon Company made in Iran and homeostasis model assessment of insulin resistance were determined.

Statically analysis

The sample size of participant women was estimated for a one-sided testing at a 0.05 significance level. Demographics and clinical features of them were represented as mean ± SD or percentage where appropriate. Chi-square test and one way-ANOVA method were used in the statistical analyses using the statistical package for the social sciences, version 18.0 (SPSS Inc., Chicago, IL, USA). The (P≤0.05) was considered as a significant difference between means

Accrding to the result of demographic and clinical characteristics baseline in women with type 2 diabetes mellitus shown in table 2, no significant difference have found amoung the women demographic and clinical features. Result showed that there were not diffrences bitween Mizaj distribution in treatments (table 3).

Table 2. Demographic and clinical characteristics baseline in women with type 2 diabetes mellitus

Table 3. Distribution of Mizaj for women with type 2 diabetes mellitus

The higher improvement of the mean glycemic indices such as fasting blood glucose and glycated hemoglobin, insulin levels, and insulin resistance  were exisits in women with the colocynth extract application in comparision with placebo group (table 4). Revealed data showed that better improvement in women with cold and wet syndrome and it is more than the hot and dry syndrome (tables 4 and 5). The differences were statistically in women with wet compared to dry syndrome groups (P≤0.05). However, in the women with glycemic outcomes a significant improvement regardless of the coldness and warmness of Mizaj were found (P≤0.05). Additionally,  no adverse effects has reported by women in the colocynth extract applicated group. The allergic reaction to the control (placebo) by some patient of dry Mizaj syndrome, leading to the discontinuation of the intervention were reported. Data od this study showed that these allergic symptoms disappeared with no need for further intervention after 5 days.

Table 4. The reulst of glycemic outcomes of women with type 2 diabetes mellitus basis on the wetness and dryness of Mizaj

*Fsb: Fasting blood glucose,Gh: Glycosylated hemoglobin ,Ins: Insulin and Ins.R: Insulin resistance.

Table 5. The reulst of glycemic outcomes of women with type 2 diabetes mellitus basis on coldness and warmness of Mizaj

*Fsb: Fasting blood glucose,Gh:Glycosylated hemoglobin ,Ins: Insulin and Ins.R: Insulin resistance.

The result of current study have shown the daily dose of colocynth extarct in three months has to improved fasting plasma glucose, glycated hemoglobin and insulin resistance respectivley. The results were in accordance with studies that reported the aqueous extract of colocynth seed has hypoglycemic acts in animals with diabetes and it is associated by saponins and glycosidic components on it (15) investigated that the antihyperglycaemic effect of colocynth was related to its saponosides, that may effect by several metabolic pathways and metabolism of insulin. They also showed that the  saponosides could influenced on glucose homeostasis directly and indirectly. Additionaly (16) mentioned that the colocynth intervention had a good effects on improving the glycemic charecteristics without adverse effects in patients with type 2 diabet. (17) showed that the colocynth hypoglycemic effect mechanisms may be  through decreasing the absorption of ingested sugars in body that could be possible if the enzymes responsible for degradation of carbohydrates complex were inhibited. (18) showed that a  decreasion in glycosylated hemoglobin and fasting blood glucose level in patients who used citrullus colocynthis after two months compared to control group. In their study the serum triglyceride and cholesterol has been not changed in type 2 diabetic patients after prescription of citrullus colocynthis. There are to many mechanisms on hypotriglyceridemic and hypocholesteremic effects of colocynth in  animal and humans but many of them has not been found yet. No adays few clinical sduties have done on animal models on the hypolipidemic effect of colocynth to enhancement of catabolism or removal of lipoproteins their liver and inhibition of liver lipid hydrolytic enzymes synthesis (19). The current study outcomes showed that colocynth intervention were more enhanced in women in wet syndrome in comparision with the women  who were in the dry syndrome. The current study data were in line with the Persian syndrome differentiation syndrome of Mizaj (20). The improvements glycemic outcomes were in line in terms of reducing fasting blood sugar, glycated hemoglobin and increasing insulin secretion in (21) studies. According to the current study results the usage of colocynth seed supplementation in diabetic women with a wet Mizaj may have more beneficial acts on improvement of the glycemic parameters.

In conclusion we could demonestrate that the potential of Persian traditional medicine hypothesis to differentiate women responding to colocynth seed extarct supplements intervention. We may have also showed that the women with wet syndrome based on Persian traditional medicine syndrome benefit more from colocynth extraxt internvention compared to the others. Further studies are needed to investigate the better benficial effects or toxicity of colocynth ethanolic extract action in diabetes mellitus type 2 women.

Acknowledgements

The authors are many thankful of Iranian University Midwifery and Nursing students and stuffs.

Conflict of interests

Authours have no conflict of interests associated with the current study.

Financial supports

Authours reported that there has not been financial support for the current study.

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