Study on Pharmacological Effects of Peikun Pills

Renming XIE, Jinfeng CHEN*, Wangting SONG, Lin YANG, Xinyue ZHANG, Lixia ZAN, Shanshan ZHAO, Mengqi YANG, Min CHENG

1. Academy of Traditional Chinese Medicine and Materia Medica of Shaanxi Province, Xi’an 710031, China; 2. School of Biological Sciences and Engineering, Shaanxi University of Technology, Hanzhong 723001, China; 3. College of Biology Pharmacy and Food Engineering, Shangluo University, Shangluo 726000, China; 4. College of Pharmacy, Shaanxi University of Chinese Medicine, Xianyang 712046, China

Abstract [Objectives] To explore the effects of Peikun Pill on blood pressure of healthy adult male rats. [Methods] The duodenum of anesthetized rats was given different doses of Peikun Pills, normal saline was used as a blank control group, and captopril was used as a positive western medicine control group. The systolic blood pressure, diastolic blood pressure, mean blood pressure and pulse pressure difference (systolic blood pressure-diastolic blood pressure), and heart rate were measured, myocardial tension time index was calculated, and statistical processing was performed. [Results] Peikun Pills have a significant pressure boosting effect, but have no effect on the myocardial tension time index. Captopril has the effects of lowering blood pressure, slowing down the heart rate and reducing the myocardial tension time index, and significantly increasing the pulse pressure difference. [Conclusions] Peikun Pills can increase blood pressure of rats. Peikun Pills are suitable for women with hypotension accompanied by anemia. However, patients with hypertension are prohibited to use.

Key words Peikun Pills, Rats, Blood pressure

1 Introduction

Peikun Pill is the secret prescription of Zaolutang, a former pharmacy and now a famous TCM hospital in Xi’an City. It has a history of more than 300 years, and a traditional Chinese medicine compound pill made by changing the dosage form of traditional Chinese medicine. Peikun Pills are modified by Sijunzi Decoction, Siwu Decoction, and Danggui Buxue Decoction. The whole prescription of Peikun Pills is composed of 21 medicines, making Peikun Pills have the functions of nourishing qi and blood, and nourishing liver and kidney. Peikun Pills are used for women with blood deficiency, indigestion, irregular menstruation, white vaginal discharge, cold pain in the lower abdomen, insufficiency of qi and blood, and prolonged infertility. The main components of Peikun Pills include 48 g of Astragali Radix Praeparata Cum Melle, 32 g of Citri Reticulatae Pericarpium, 8 g of Glycyrrhizae Radix Et Rhizoma Praeparata Cum Melle, 48 g of fried Atractylodis Macrocephalae Rhizoma, 16 g of Glehniae Radix, 32 g of Poria, 80 g of wine-processed Angelicae Sinensis Radix, 32 g of Ophiopogonis Radix, 16 g of Chuanxiong Rhizoma, 32 g of fried Ziziphi Spinosae Semen, 16 g of wine-processed Paeoniae Radix Alba, 9 g of Amomi Fructus, 32 g of carbonized Eucommiae Cortex, 20 g of Juglandis Semen, 40 g of salted Trigonellae Semen, 16 g of vinegar-fried Artemisiae Argyi Folium, 32 g of Longan Arillus, 32 g of Cornifructus (processed), 4 g of processed Polygalae Radix, 64 g of Rehmanniae Radix Praeparata, and 8 g of Schisandrae Chinensis Fructus (cooked)[1-2].

Zaolutang, formerly known as Deruntang, was built in the second year of Tianqi of the Ming Dynasty (1622). It has been 388 years, which is 47 years earlier than the Tongrentang founded in 1669. Zaolutang has always adhered to the business philosophy of "complying with the ancient concoctions, cheating neither old nor young". In terms of treating gynecological diseases, infertility, male diseases, intractable diseases and chronic diseases, it has excellent medical skills and noble medical ethics. It adheres to the principle that "strenuous efforts in making exquisite medicines can rarely be seen, but heaven knows our sincerity". Even the famous old Qin opera artists Tang Disu and Yan Zhensu relished, in the lyrics ofMr.BaiSeeingaDoctor, that "Rendan Baodan and Wujidan, Zaolutang’s Peikun Pills".

In order to re-develop the national intangible cultural heritage Chinese time-honored product Peikun Pills, we performed this experiment.

2 Materials and methods

2.1 Materials

2.1.1Laboratory animals. Male and female KM mice with weight of 18-20 g were provided by the Laboratory Animal Center of School of Pharmacy of Xi’an Jiaotong University, certificate No.0012084, and license No.SCXX (Shaanxi) 2018-003. And 40 pieces of Male KM rats with weight of 230-350 g were provided by the Laboratory Animal Center of School of Pharmacy of Xi’an Jiaotong University, certificate No.0012084, and license No.SCXX (Shaanxi) 2012-003.

2.1.2Instruments and reagents. 1 mL syringe, mortar, surgical forceps, balance, surgical scissors, scalpel, YLS-16A small animal bone strength tester (equipment station of Shandong Academy of Medical Sciences); XS105 analytical balance (Mettler-Toledo Instruments Co., Ltd.); YSL-4C Rota-rod system (equipment station of Shandong Academy of Medical Sciences; RM6240BD multi-channel physiological signal acquisition and processing system (Chengdu Instrument Factory).

2.1.3Test drugs. Peikun Pills (Xi’an C.P. Pharmaceutical Co., Ltd., National Medicine Permit No.Z61021462); Ruibai (Recombinant Human Granulocyte Colony-stimulating Factor Injection, Qilu Pharmaceutical Co., Ltd., National Medicine Permit No.S19990050); Urethane (ethyl carbamate, produced by China Caoyang Second Reagent Factory, Shanghai Q/HG22-771-68); 0.9% sodium chloride injection (Sichuan Kelun Pharmaceutical Co., Ltd., National Medicine Permit No.H51021158); Perfect Major Supplementation Pill (Lanzhou Foci Pharmaceutical Co., Ltd., National Medicine Permit No.Z62020795); Captopril (Changzhou Pharmaceutical Factory Co., Ltd., National Medicine Permit No.H32023731); Heparin Sodium Injection (Shanghai No.1 Biochemical & Pharmaceutical Co., Ltd., National Medicine Permit No.H31022051); Cyclophosphamide for Injection (Jiangsu Suncadia Medicine Co., Ltd., National Medicine Permit No.H32020857).

2.2 Experiment method

2.2.2Effects on blood biochemical and hematological indicators. Fifty KM mice were randomly divided into a blank control group, a high-dose Peikun Pill group, a medium-dose Peikun Pill group, a low-dose Peikun Pill group, and a Perfect Major Supplementation Pill group, a total of 5 groups, each with 10 mice. The high, medium, and low-dose Peikun Pill groups were given Peikun Pills at the dose of 6.00, 3.00, and 1.50 g/kg/d by intragastric administration for 40 consecutive days; Perfect Major Supplementation Pill group mice were given by intragastric administration at the dose of 6.00 g/kg/d for 40 consecutive days; the blank control group mice were given an equal volume of water by intragastric administration. On the 40th day after the administration, the mice were anesthetized and the eyeballs were removed to obtain blood for hematology and biochemical indicator examination. Took the heart, liver, spleen, lung, kidney, thymus, adrenal gland, ovary, uterus, weighed and calculated the organ index.

2.2.3Effects on cyclophosphamide-induced leukopenia and the prevention and treatment of osteoporosis. Sixty KM mice were randomly divided into a blank control group, a model group, a positive group (Ruibai group), a high-dose Peikun Pill group, a medium-dose Peikun Pill group, a low-dose Peikun Pill group, a total of 6 groups, each with 10 mice. The high, medium, and low-dose Peikun Pill groups were given Peikun Pills at the dose of 6.00, 3.00, and 1.50 g/kg/d by intragastric administration for 16 consecutive days; Ruibai group mice were given recombinant human granulocyte stimulating factor injection (Ruibai) by subcutaneous injection at the dose of 10 μg/kg/d for 16 consecutive days; the model group and blank control group mice were given an equal volume of water by intragastric administration. After 10-12 d of administration, except the blank control group, all animals in the other groups were injected subcutaneously with cyclophosphamide at the dose of 100 mg/kg on the 10th day, and 80 mg/kg on the 11th and 12th days. On the 16th day after the administration, the mice were anesthetized and the eyeballs were removed to obtain blood for hematology examination. Continued to administer for 3 d, then mice were killed, took the heart, liver, spleen, lung, kidney, thymus, adrenal gland, ovary, uterus, weighed and calculated the organ index. Separated the mouse femur, weighed the femur, and measured the length, fracture force and crushing force of the femur.

2.2.4Effects on strengthening and consolidating body resistance. (i) Loaded swimming experiment of mice: the purchased Kunming mice were divided into 5 groups according to body weight and sex. Control group: a low-dose Peikun Pill 1.5 g/kg, a medium-dose Peikun Pill 3.0 g/kg, a high-dose Peikun Pill 6.0 g/kg, and Perfect Major Supplementation Pill 6.0 g/kg. Mice were intragastrically administered one time daily for 10 d. After 10 d, a 10% weight-bearing lead sheet was tied to the hind limbs of the mice, then placed in a water tank with a length of 45 cm, a width of 30 cm, and a depth of 25 cm. The fatigue time was determined by burying the head of the mouse in the water 3 times, timed by a stopwatch, and statistical processing of swimming time was performed. (ii) Rotarod test in male mice: 50 pieces of Kunming mice purchased were evenly divided into 5 groups according to their body weight. Control group: a low-dose Peikun Pill 1.5 g/kg, a medium-dose Peikun Pill 3.0 g/kg, a high-dose Peikun Pill 6.0 g/kg, and Perfect Major Supplementation Pill 6.0 g/kg. Mice were intragastrically administered one time daily for 12 d. After 7 d of administration, each mouse was pre-adapted to the rotating rod, and each mouse was rotated for 5 min daily at a speed of 24 rpm. On the 12th day after the administration, a formal test was performed, and the time when the mouse rotating rod fell was counted, and taking the mouse not falling as a limit of 200 min.

3 Results and analysis

3.1 Effects on the blood of anesthetized ratsPeikun Pills have a significant effect on boosting the blood pressure, but have no effect on the myocardial tension time index; captopril has the effect of lowering the blood pressure, slowing the heart rate, increasing the pulse pressure difference and reducing the myocardial tension time index. Peikun Pills have effects of boosting the blood pressure of rats. These indicate that Peikun Pills are suitable for women with hypotension accompanied by anemia, but patients with hypertension are prohibited to use. The results are shown in Table 1-6.

Table 1 Effects of Peikun Pills on the systolic blood pressure of anesthetized rats mmHg)

Table 2 Effects of Peikun Pills on the diastolic blood pressure of anesthetized rats mmHg)

Table 3 Effects of Peikun Pills on the mean blood pressure of anesthetized rats mmHg)

Table 4 Effects of Peikun Pills on the pulse pressure difference of anesthetized rats mmHg)

Table 5 Effects of Peikun Pills on the heart rate of anesthetized rats times/min)

Table 6 Effects of Peikun Pills on the myocardial tension time index of anesthetized rats

3.2 Effects on blood biochemical and hematological indicatorsPeikun Pills have no significant effects on the growth and development, organ index, liver and kidney function, glucose and lipid metabolism, serum calcium and phosphorus content, alkaline phosphatase and red blood cells in female mice. The low and medium doses of Peikun Pills can significantly reduce the total number of white blood cells in the blood of female mice. Perfect Major Supplementation Pill also can significantly reduce the total number of white blood cells in the blood of female mice. Peikun Pills have a significant effect on the total platelet count and plateletcrit (PCT) in the blood of mice. Therefore, Peikun Pills are safe for long-term administration. Long-term administration can significantly reduce the total number of white blood cells and significantly increase the total platelet count and plateletcrit. It is suggested that Peikun Pills can relieve the thrombocytopenia caused by radiotherapy and chemotherapy in tumor patients, and may have a therapeutic effect on the uterine bleeding of women. The results are shown in Table 7-15.

Table 7 Effects of Peikun Pills on the weight gain of female mice n=10, g)

Table 8 Effects of Peikun Pills on the organ index of female mice n=10, mg/10 g weight)

Table 9 Effects of Peikun Pills on the thymus, adrenal gland, ovary, uterus index of female mice n=10, mg/10 g weight)

Table 10 Effects of Peikun Pills on the liver and kidney functions of female mice n=10)

Table 11 Effects of Peikun Pills on the protein content of female mice n=10)

Table 12 Effects of Peikun Pills on the sugar, lipid, alkaline phosphatase, calcium and phosphorus content in female mice n=10)

Table 13 Effects of Peikun Pills on the white blood cells in blood of female mice n=10)

Table 14 Effects of Peikun Pills on the red blood cells in blood of female mice n=10)

Table 15 Effects of Peikun Pills on the blood platelets in female mice n=10)

3.3 Effect on the cyclophosphamide-induced reduction of white blood cells and the prevention and treatment of osteoporosisThe levels of white blood cells and platelets in the peripheral blood of the mice in each group were lower than those in the blank control group, and the modeling was successful. The alkaline white blood cell level of the Peikun Pill groups increased by 30% compared with the model group, the spleen index increased by 38.75% compared with the model group, the lung index increased by 16% compared with the model group, the thymus index increased by 38.46% compared with the model group, the bone length increased by 6.95% compared with the model group, the bone weight increased by 24.36% compared with the model group, and the bone crushing force increased by 27.13% compared with the model group. Peikun Pills have a certain antagonistic effect on the cyclophosphamide induced reduction of alkaline white blood cells, and have a certain protective effect on the immune organs of mice, the spleen, lung, and thymus, and have a significant prevention and treatment effects on osteoporosis in mice. The results are shown in Table 16-20 and Fig.1-4.

Table 16 Effects of Peikun Pills on the weight of mice n=10, g)

Table 17 Effects of Peikun Pills on the white blood cells of mice n=10)

Table 18 Effects of Peikun Pills on the red blood cells and platelets of mice n=10)

Table 19 Effects of Peikun Pills on the bone level of mice n=10)

Table 20 Effects of Peikun Pills on the organ index n=10)

Note: Compared with the control group, ##P<0.01; com-pared with the model group, *P<0.05.Fig.1 Effects of Peikun Pills on the white blood cells of miceNote: Compared with the control group, ##P<0.01; compared with the model group, **P<0.01.Fig.2 Changes of organ index of mice in each group

Note: Compared with the control group, ##P<0.01; compared with the model group, *P<0.05, **P<0.01.Fig.3 Changes of bone length and bone weight of mice in each groupNote: Compared with the model group, **P<0.01.Fig.4 Changes in bone biomechanics of mice in each group

3.4 Effects of strengthening and consolidating body resistanceThe administration of high, medium and small doses of Peikun Pills for 10 d significantly prolonged the loaded swimming time of mice. The detailed results are listed in Table 21 and Table 22. The loaded swimming time of male mice administered with small, medium and high doses of Peikun Pills prolonged 34.08%, 74.32%, and 155.73% respectively compared with the control group. The loaded swimming time of female mice administered with small, medium and high doses of Peikun Pills prolonged 146.02%, 230.26%, and 321.39% respectively compared with the control group, indicating that Peikun Pills have stronger effects on female mice. The administration of high, medium and small doses of Peikun Pills for 12 d significantly prolonged the rotating rod fatigue time of mice. The rotating rod fatigue time of mice administered with small, medium and high doses of Peikun Pills prolonged 38.43%, 87.22%, and 99.32% respectively compared with the control group. The Perfect Major Supplementation Pill also has effects of prolonging the rotating rod fatigue time, it prolonged the rotating rod fatigue time for 69.23% compared with the control group. The mouse loaded swimming experiment and rotating rod fatigue experiment of Peikun Pills prove that Peikun Pills have significant anti-fatigue effect, indicating that Peikun Pills have strong effects of strengthening and consolidating body resistance. Peikun Pills may have good effects on strengthening physical strength for weak or sick women. The results are shown in Table 21-23.

Table 21 Effects of Peikun Pills on the loaded swimming time of male mice n=10)

Table 22 Effects of Peikun Pills on the loaded swimming time of female mice n=10)

Table 23 Effects of Peikun Pills on the rotating rod fatigue time of male mice n=10)

4 Conclusions

Peikun Pills have effects of boosting the systolic blood pressure, diastolic blood pressure and mean blood pressure of anesthetized rats, suggesting that Peikun Pills can be used for hypotensive patients with anemia. Besides, Peikun Pills are safe for long-term administration. Long-term administration can significantly reduce the total number of white blood cells and significantly increase the total platelet count and plateletcrit, suggesting that Peikun Pills can relieve the thrombocytopenia caused by radiotherapy and chemotherapy in tumor patients, and may have a therapeutic effect on the uterine bleeding of women. Peikun Pills have no effect on the cyclophosphamide induced reduction of white blood cells in mice, but have a significant effect of resisting osteoporosis. Peikun Pills can significantly prolong the loaded swimming time and rotating rod fatigue time of mice, suggesting that Peikun Pills have strong effects of strengthening and consolidating body resistance.