KR101721094B1 - Compostion comprising blue honeysuckle extract for treatment of nausea - Google Patents

Abstract

The present invention relates to a composition for enhancing therapeutic effects on nausea and vomiting, containing ondansetron and a Lonicera caerulea var. edulis Turcz. ex Herder fruit extract. More specifically, by injecting ondansetron into a testing body and then the Lonicera caerulea var. edulis Turcz. ex Herder fruit extract, it is possible to remarkably increase therapeutic effects of ondansetron on nausea and vomiting, while reducing and preventing side effects due to various treatments of nausea and vomiting.

Description

[0001] The present invention relates to a composition for enhancing the nausea and vomiting therapeutic effects,

The present invention provides a composition for enhancing the therapeutic effect of nausea and vomiting comprising ondansetron and royal jelly extract.

Various chemotherapeutic agents are used for chemotherapy. Cisplatin, one of the various anticancer drugs, is a representative platinum-complexing anticancer drug containing platinum (II) in the center. It is one of the broad-spectrum chemotherapeutic agents used in various tumor treatments and has a relatively excellent anticancer effect. However, It has been shown that the synthesis of serotonin in endocrine cell cells and the activation of serotonin 5-HT3 receptors have resulted in various gastrointestinal toxicities, such as gastrointestinal tract damage, gastric emptying, It acts as a typical redox cycler to produce gastrointestinal mucosal cell function and DNA damage, and it is known to cause gastrointestinal dysfunction, especially movement disorder. Therefore, the control of gastrointestinal side effects by cisplatin has become a very important aspect to take advantage of the strong anticancer effect of this drug. In the development of various digestive system drugs including the antiepileptic drugs, cisplatin Various 5-HT3 receptors blockers, anti-inflammatory agents and antioxidants have been shown to be effective against these cisplatin-induced gastrointestinal disorders.

In this regard, Ondansetron is a representative oral serotonin 5-HT3 receptor antagonist, which is frequently used for the relief and treatment of various nausea and vomiting symptoms. However, it has relatively strong toxicity in preclinical phase, Prolongation problems have been raised, limiting their use [GlaxoSmithKline, 2006, 2010; McKechnie and Froese, 2010; Hafermann et al., 2011].

On the other hand, Blue honeysuckle is a species of Lonicera caerulea var. edulis has been used as a traditional medicinal plant in northern Russia, China and Japan. It is a relatively uncommon medicinal fruit in North America and Europe, and contains abundant ascorbic acid and phenolic components. Especially, it has excellent antioxidative effect It contains abundantly known anthocyanins, flavonoids and small phenolic acids.

Accordingly, in studying a method for alleviating the toxicity or side effects caused by the use of the ondansetron, the inventors of the present invention confirmed that the treatment of onion skin with the extract of Buttermilk can reduce side effects while increasing the therapeutic effect of ondansetron, Thereby completing the invention.

Accordingly, it is an object of the present invention to provide a composition for enhancing the treatment, improvement, and prevention of nausea and vomiting symptoms and reducing the side effects thereof by using the extract of royal jellyfish The purpose of that is to do.

However, the technical problem to be solved by the present invention is not limited to the above-mentioned problems, and other matters not mentioned can be clearly understood by those skilled in the art from the following description.

In order to achieve the above object, the present invention provides a composition for promoting nausea and vomiting therapeutic effect comprising ondansetron and royal jelly extract.

In one embodiment of the present invention, the nausea and vomiting are characterized by being caused by chemotherapy.

In another embodiment of the present invention, the composition is characterized by alleviating side effects due to nausea and vomiting treatment,

In another embodiment of the present invention, the ondansetron and the royal thai fruit extract are pre-mixed and formulated or separately formulated.

In another embodiment of the present invention, the ondansetron and the royal thai fruit extract are administered parenterally, orally, locoregionally, or percutaneously.

In another embodiment of the present invention, the administration of the royal jelly extract is started within 5 minutes to 4 hours after administration of the ondansetron.

The composition for enhancing the nausea and vomiting therapeutic effect composition containing the extract of the royal thyrotexin provided by the present invention as an active ingredient is administered in combination with ondansetron to improve the efficiency of treatment for nausea and vomiting and alleviate adverse side effects of ondansetron alone .

Fig. 1 is a diagram showing an experimental design of Example 1. Fig.
FIG. 2 is a graph showing the results of observation of Ondansetron concentration in blood of Example 2. FIG.
3 is a view showing an experimental design of the second embodiment.
Fig. 4 is a view showing the result of checking the change in body weight according to each group of Example 2. Fig.
FIG. 5 is a graph showing the results of observing changes in blood Ondansetron concentration in Example 2. FIG.
6 is a diagram showing an experimental design of the third embodiment.
Fig. 7 is a graph showing the results of confirming changes in body weight according to each group of Example 3. Fig.
8 is a diagram showing the results of confirming histopathological changes of the spleen in Example 3. Fig.
9 is a diagram showing the results of observation of histopathological changes of the heart in Example 3. Fig.
10 is a diagram showing the results of observation of histopathological changes of the lungs in Example 3. Fig.
11 is a view showing the result of observing histopathological changes of testicular tubules in Example 3. Fig.
FIG. 12 is a diagram showing the results of observation of histopathological changes in liver in Example 3. FIG.
13 is a diagram showing the results of observation of histopathological changes of the epididymal duct in Example 3. Fig.
14 is a diagram showing the results of observing histopathological changes of submandibular lymph nodes in Example 3. Fig.
Fig. 15 is a diagram showing the results of confirming histopathological changes on the body in Example 3. Fig.
16 is a view showing the result of confirming the histopathological changes in the pylorus in Example 3. Fig.
17 is a diagram showing the experimental design of the fourth embodiment.
18 is a view showing the result of checking the change in body weight according to each group of Example 4. Fig.
FIG. 19 is a graph showing the results of observing changes in pituitary tryptophan hydroxylase activity according to each group in Example 4. FIG.
20 is a graph showing the results of observing changes in the activity of monoamine oxidase in the pylorus according to each group in Example 4. FIG.
FIG. 21 is a graph showing the results of confirming the change in the gastrin content of the pylorus according to each group in Example 4. FIG.
FIG. 22 is a graph showing the results of confirming changes in the content of serotonin in the pylorus according to each group in Example 4. FIG.
FIG. 23 is a graph showing the results of histopathological changes observed on the stomach (in the figure, A = medium control group: rats administered peritoneally and rats / distilled water orally administered rats / B = Cisplatin control group: rats administered with cisplatin And rats (O1) / D = Cisplatin Oral rats / C = Cisplatin Rats and ondansetron 0.1 mg / ml Single oral doses rats and BH 40 mg / ml Single oral doses rats (BH400) / E = Cisplatin rats (O + BH400) / F = Cisplatin rats and ondansetron 0.1 mg / ml and BH 40 mg / ml rats and ondansetron 0.1 mg / ml and BH 20 mg / ml (O + BH100) / GI = gastrointestinal / LU = lumen / rats (O + BH200) / G = Cisplatin administered intraperitoneally and oral ondansetron 0.1 mg / ml and BH 10 mg / MU = mucosa / ML = muscle layers / SM = submucosa layer).
24 is a view showing the results of confirming the histopathological changes of the pylorus according to each group in Example 4. Fig.
Fig. 25 shows the histopathological changes of the colon including residual feces according to each group in Example 4. Fig.
FIG. 26 is a graph showing the results of confirming changes in mucosal immunoreactive cells of the pyloric stomach and colon according to each group in Example 4. FIG.

In order to develop a composition capable of increasing the tumor volume reduction effect while mitigating various adverse effects of administration of ondansetron for the treatment of nausea and vomiting induced by chemotherapy, , And it was confirmed that the extract of Fusarium oxysporum in natural products has excellent nausea and vomiting treatment effect and side effect reduction effect, thus completing the present invention.

Accordingly, it is an object of the present invention to provide a composition for enhancing the therapeutic effect of nausea and vomiting comprising ondansetron and royal juniper fruit extract.

That is, the nausea and vomiting may be caused by chemotherapy but are not limited thereto.

In one embodiment of the present invention, the ondansetron and the royal jelly extract may be previously mixed and formulated or separately formulated.

The administration period of the royal jellyfish extract may be performed within 5 minutes to 4 hours after administration of the ondansetron, preferably within 30 minutes to 3 hours, most preferably within 30 minutes to 2 hours and 30 minutes And may be administered 2 hours after the administration of ondansetron as in an embodiment of the present invention, but the present invention is not limited thereto.

As ondansetron are 5-HT3 receptor antagonist for representative serotonin oral administration used in the present invention, the formula of the ondansetron is a _{C 18 H 19 N 3O · 2} O, to the same as the formula (1).

[Chemical Formula 1]

The ondansetron and royal jelly extract may be administered parenterally, orally, locoregionally, or transdermally. The above-described nutrient extract is preferably orally administered, but may be suitably selected by those skilled in the art depending on the condition and the weight of the patient, the degree of disease, and the period of time.

In the present invention, an 'individual' refers to a subject in need of treatment for diseases, and more specifically, a human or non-human primate, mouse, rat, dog, cat, horse, And the like.

In addition, the present invention can provide a method for treating nausea and vomiting, which comprises a farina nut extract.

The pharmaceutical composition of the present invention may comprise a pharmaceutically acceptable carrier. The pharmaceutically acceptable carrier may include, but is not limited to, physiological saline, polyethylene glycol, ethanol, vegetable oil, and isopropyl myristate.

In one embodiment of the present invention, the preferred dosage of the pharmaceutical composition varies depending on the condition and the weight of the patient, the degree of disease, the drug form, the administration route, and the period, but can be appropriately selected by those skilled in the art. However, it is preferably administered at a daily dose of 0.001 to 300 mg / kg body weight, more preferably 0.01 to 200 mg / kg body weight.

The pharmaceutical composition of the present invention can be administered to mammals such as rats, mice, livestock, humans, and the like in various routes. The method of administration is not limited and can be administered, for example, orally, rectally, or by intravenous, intramuscular, subcutaneous, intrauterine, or intra-cerebroventricular injection.

The composition for enhancing the nausea and vomiting therapeutic effect comprising the extract of Staphylococcus aureus according to the present invention can increase the blood sugar enhancing effect and can alleviate various side effects caused by the administration of ondansetron alone have.

Hereinafter, preferred embodiments of the present invention will be described in order to facilitate understanding of the present invention. However, the following examples are provided only for the purpose of easier understanding of the present invention, and the present invention is not limited by the following examples.

[Example]

Example  One. Nausea and vomiting for chemotherapy  remedy Ondansetron (ZofranTM) and  blue honeysuckle (BH): Effect of Ondansetron on pharmacokinetics - Single oral administration

In Example 1, the effect of ondansetron on the drug power of ondansetron and BH according to the BH complex was evaluated using normal male rats, in order to study on the combination of ondansetron and BH, , And drug interactions. After oral administration of ondansetron (0.1 mg / ml) and BH (40, 20 or 10 mg / ml) in a single oral dose of 10 ml / kg, 30 minutes before administration, 30 minutes after administration, , Ondansetron (0.1 mg / ml), and ondansetron (0.1 mg / ml) were measured by measuring the concentration of ondansetron in the blood and calculating noncompartmental pharmacokinetics data (Cmax, Tmax, AUC, t _1/2 and MRT) And compared with the composition administration group.

The solution of Duckweed fruit used in this study was a solution of 200 g of 63brix solution in which Duckweed was dissolved in distilled water so that it had 25 brix. After the lyophilization, the concentration of the powder was adjusted by adding distilled water to each concentration.

1.1. Preparation of experimental animals and group separation

Four male SPF / VAF Outbred Crl: CD [SD] male rats (OrientBio, Seungnam, Korea) were obtained and after 18 days of purification, (349.85 ± 17.12 g, 310 to 387 g).

(A) Ondansetron (0.1 mg / ml) single composition administration group

(B) Ondansetron and BH (0.1 and 40 mg / ml) composite composition administration group

(C) Ondansetron and BH (0.1 and 20 mg / ml) composite composition administration group

(D) Ondansetron and BH (0.1 and 10 mg / ml) complex composition administration group

1.2. Preparation and administration of experimental material composition

In the ondansetron and BH complex composition administration group, 0.1 mg of ondansetron (CTX Lifesciences (P) Ltd., Gujarat, India) was dissolved and administered at a dose of 10 ml / mg / ml of the ondansetron composition, respectively, and BH was dissolved at a concentration of 40, 20, and 10 mg / ml, and then administered orally at a dose of 10 ml / kg (Table 1, Fig. 1).

Group Sex Oral Dose (mg / kg / day) Animal No. O + BH PK Mix formulation of ondansetron and BH Active Male Ondansetron 0.1 mg / ml single formula 10 m / kg A01 ~ A05 Active Male Ondansetron 0.1 mg / ml and BH 40 mg / ml mixed formula 10 ml / kg B01 to B05 Active Male Ondansetron 0.1 mg / ml and BH 20 mg / ml mixed formula 10 ml / kg C01 ~ C05 Active Male Ondansetron 0.1 mg / ml and BH 10 mg / ml mixed formula 10 ml / kg D01 ~ D05

1.3. Blood collection methods and Ondansetron  Method of analysis of blood concentration

Approximately 0.5 ml of whole blood from the orbital venous orbit was infused with 50 IU of heparin (Sigma-Aldrich, St. Louise, MO, USA) at 30 minutes, 30 minutes, 1, 2, 3, 4, 6, USA), and the plasma was separated by centrifugation at 13,000 rpm for 10 minutes immediately after collection. The separated plasma was stored at -150 ° C until LC-MS / MS analysis.

The concentration of ondansetron was measured by LC-MS / MS method using Carbamazepine (Sigma, St. Louise, MO, USA) as an internal standard. Chromatographic analysis was performed using Agilent 1100 Series HPLC (Agilent Technologies, Santa Clara, Calif., USA) and column effluent was analyzed using an API 2000 triple-quadruple mass spectrometer (Applied Biosystems, Foster City, CA, USA) . The conditions of each analysis are as follows:

HPLC  Condition

Column: Waters Symmetry TMC18 (2.1 x 50 mm, 3.5 m) (Waters Corp., Milford, MA, USA)

Column Oven: 30 ° C

Mobile phase: 50% distilled water (0.1% formic acid) / 50% acetonitrile

Flow rate: 0.30 ml / min

Injection Volume: 5.00 μl

LC-MS / MS

Ion source: Turbo Ion Spray (500 < 0 > C)

Polarity: Positive

Carbamazepine (IS) = m / z 237 > 194 (Retention time: 0.74 min), ondansetron = 294 > 170 (Retention time: 0.40 min)

Standard Curve: Analyst 1.4.1, Linear (1 / x ² , noIterate)

1.3. Check the result

The concentrations of ondansetron (ng / ml), pharmacokinetic index (ng / ml), blood pressure, blood pressure and blood pressure were measured at 30 minutes before, 30 minutes after, 1, 2, 3, 4, 6, 8 and 24 hours before administration of Ondansetron alone or three doses of Bd and ondansetron composite composition Cmax, Tmax, AUC, t _1/2, and MRT were compared using noncompartmental pharmacokinetics data analyzer program (PK solutions 2.0; Summit, Montrose, CO, USA).

1.3.1. Blood ondansetron  Concentration observation

Ondansetron alone or three doses of BH and ondansetron combination compositions started to detect ondansetron in the blood from 30 minutes after each administration, and were continuously detected over 3 hours after administration, and ondansetron and BH 40 or 20 mg / , Ondansetron of 0.21 to 0.49 ng / ml was detected at 4 hours after administration. The ondansetron and BH 40 mg / ml compound administration group showed no significant change in ondansetron concentration compared to ondansetron alone group (p <0.01) The decrease in ondansetron concentration in the blood was observed in 30 minutes to 1 hour after administration of the ondansetron and BH 20 mg / ml composite composition, and the ondansetron concentration was significantly (p <0.01 or p <0.05) Reduction was observed in the ondansetron and BH 10 mg / ml composite composition administration groups for 30 minutes to 2 hours after administration. (Fig. 2).

In the Ondansetron and BH 40 mg / ml compound administration group, ondansetron concentration in the blood of 30 minutes after administration, 1, 2 and 3 hours after administration was 4.26, 23.48, 44.96 and 6.93% points, respectively Change.

In the Ondansetron and BH 20 mg / ml compound administration group, the ondansetron concentration in blood of 30 minutes after administration, 1, 2 and 3 hours after administration was -64.13, -63.57, -56.32 and - 36.58% of the points.

In the Ondansetron and BH 10 mg / ml compound administration group, blood ondansetron concentration of 30 minutes, 1, 2 and 3 hours after administration was -61.64, -59.72, -61.20 and - 53.56% of the points.

1.3.2. Of Tmax  change

Ondansetron had a Tmax of 0.50 ± 0.00 hr, which was the same as 0.50 ± 0.00 hr in the ondansetron alone group (Table 2) .

1.3.3. Cmax  change

The ondansetron Cmax was 4.41 ± 2.39 ng / ml, which was significantly lower than 4.23 ± 1.97 ng / ml in the ondansetron alone group in the administration of Ondansetron and BH 40 mg / ml. Ondansetron and BH 20 or 10 mg / ml concentrations of ondansetron showed 1.52 ± 0.53 and 1.62 ± 0.37 ng / ml of ondansetron, respectively, which were significantly higher than those of ondansetron alone (-64.13 and -61.64% (p < 0.01) (Table 2).

1.3.4. AUC  change

In the ondansetron group, the AUC _{0 -t} was 5.94 ± 3.02 hr · ng / ml, which was higher than the 4.56 ± 2.50 hr · ng / ml in the ondansetron alone group. The Ondansetron and BH 40 mg / The ondansetron AUC _{0 -t} levels were 1.78 ± 0.72 and 1.65 ± 0.43 hr · ng / ml, respectively, in ondansetron and BH 20 or 10 mg / (P <0.01 or p <0.05) reduction of -60.90 and -63.73% points, respectively. In addition, ondansetron and AUC0-inf of ondansetron alone were 6.44 ± 3.09 and 2.20 ± 0.84 hr · ng / ml, respectively, in ondansetron and BH 40 or 20 mg / ml, but ondansetron and BH 10 mg / ml showed a significant change in the AUC _{0 - inf} of 1.96 ± 0.48 hr · ng / ml in the ondansetron group, indicating that ondansetron (P <0.01) decrease compared to the single administration group at -61.48% point. (Table 2).

1.3.5. t _1/2 Change of

Ondansetron plasma t _1/2 was 0.99 ± 0.10, 1.13 ± 0.35, and 0.72 ± 0.12 hr, respectively, in the combination administration of Ondansetron and three doses of BH 40, 20 or 10 mg / (P <0.001), but the difference was not significant (p <0.05)

1.3.6. MRT _inf Change of

It is observed as Ondansetron three capacity of BH 40, 20 or 10 mg / ml concentration of the compound composition treated blood MRT _inf of ondansetron 1.59 ± 0.15, 1.72 ± 0.38 and 1.25 ± 0.12 hr, ondansetron 1.23 ± 0.36 hr of a single composition administered group Compared with those of the control group (Table 2).

Treatment
PK Parameters Ondansetron single formula Ondansetron mixed formulas with BH 0.1 mg / ml 40 mg / ml 20 mg / ml 10 mg / ml Tmax (hrs) 0.50 0.00 0.50 0.00 0.50 0.00 0.50 0.00 Cmax (ng / ml) 4.23 ± 1.97 4.41 ± 2.39 1.52 ± 0.53 ^a 1.62 ± 0.37 ^a AUC _0-t (hr 4.56 ± 2.50 5.94 + - 3.02 1.78 ± 0.72 ^b 1.65 + 0.43 ^a AUC _0-inf (hr 5.09 ± 2.74 6.44 ± 3.09 2.20 ± 0.84 1.96 + 0.48 ^a t _1/2 (hr) 0.72 + 0.28 0.99 ± 0.10 1.13 + 0.35 0.72 + - 0.12 MRT _inf (hr) 1.23 + - 0.36 1.59 ± 0.15 1.72 + - 0.38 1.25 + - 0.12

(AUC) of ondansetron, indicating that the concentration-dependency of the complex-formulated BH was not recognized, thus determining whether it can be used as an effective herbal oriental medical treatment for general cancer patients. In conclusion, the effect of ondansetron on the pharmacokinetics of BH complex composition should be evaluated after repeated administration of combination therapy for a certain period of time.

Example  2. Nausea and vomiting for chemotherapy  remedy Ondansetron (ZofranTM) and  Development of a complex formulation of blue honeysuckle (BH): Effects on ondansetron pharmacokinetics - Repeated oral administration for 7 days

In Example 2, the effect of ondansetron on the drug power of BH complex was investigated by repeated oral administration for 7 days, in order to study the development of a combination preparation of ondansetron and BH, Male rats were evaluated to observe clearer drug interactions. The combination of ondansetron (0.1 mg / ml) and BH (40, 20 or 10 mg / ml) was orally administered at a dose of 10 ml / kg for 7 days and then 30 minutes before administration, calculating a, 2, 3, 4, 6, 8 and 24 hours blood was then measuring the concentration of blood ondansetron, noncompartmental pharmacokinetics data (Cmax, Tmax, AUC, t _1/2 and MRT), ondansetron 0.1 mg / ml &lt; / RTI &gt; alone.

2.1. Isolation of experimental animals and groups

Male Sprague-Dawley rats (OrientBio, Seungnam, Korea) were obtained from 160 SPF / VAF Outbred Crl: CD [SD], and after 12 days of purification, the animals with constant weight were selected and classified into 4 groups (263.25 ± 11.36 g, 247 to 285 g). The administration method was repeatedly orally administered for 7 days, and sterilized distilled water was used as a medium (10 ml / kg).

Group separation (total 4 groups: 5 per group)

(A) Ondansetron (0.1 mg / ml) single composition administration group

(B) Ondansetron and BH (0.1 and 40 mg / ml) composite composition administration group

(C) Ondansetron and BH (0.1 and 20 mg / ml) composite composition administration group

(D) Ondansetron and BH (0.1 and 10 mg / ml) complex composition administration group

2.2. Experimental material composition preparation, administration, and blood collection method

In the Ondansetron alone group, 0.1 mg of ondansetron (CTX Lifesciences (P) Ltd., Gujarat, India) was dissolved per ml of sterilized distilled water and the solution was orally administered at a dose of 10 ml / kg for 7 days. , BH was dissolved in the ondansetron composition at a concentration of 0.1 mg / ml, respectively, at a concentration of 40, 20 and 10 mg / ml, and then orally administered at a dose of 10 ml / kg for 7 days (Table 3, Fig. 3).

Group Sex Seven day-repeated oral treatment Animal No. Mix formulation of ondansetron and BH Active Male Ondansetron 0.1 mg / ml single formula 10 m / kg A01 ~ A05 Active Male Ondansetron 0.1 mg / ml and BH 40 mg / ml mixed formula 10 ml / kg B01 to B05 Active Male Ondansetron 0.1 mg / ml and BH 20 mg / ml mixed formula 10 ml / kg C01 ~ C05 Active Male Ondansetron 0.1 mg / ml and BH 10 mg / ml mixed formula 10 ml / kg D01 ~ D05

Approximately 0.5 ml of whole blood from the orbital venous organs was infused with 50 IU of heparin (Sigma-Aldrich, St. Louis, MO) for 30 minutes before the last 7 doses, 30 minutes after dosing, 1, 2, 3, 4, 6, , MO, USA), and the plasma was separated by centrifugation at 13,000 rpm for 10 minutes immediately after collection. The separated plasma was stored at -150 ° C until LC-MS / MS analysis.

2.3. Check the result

The concentrations of ondansetron were measured by LC-MS / MS using the internal standard of Carbamazepine (Sigma, St. Louise, MO, USA) in the separated plasma for Ondansetron blood concentration analysis. Chromatographic analysis was performed using Agilent 1100 Series HPLC (Agilent Technologies, Santa Clara, Calif., USA) and column effluent was analyzed using an API 2000 triple-quadruple mass spectrometer (Applied Biosystems, Foster City, CA, USA) .

HPLC  Condition

_{^{Column: Waters Symmetry TM C 18 (}} 2.1 × 50mm, 3.5μm) (WatersCorp, Milford, MA, USA.)

Column Oven: 30 ° C

Mobile phase: 50% distilled water (0.1% formic acid) / 50% acetonitrile

Flow rate: 0.30 ml / min

Injection Volume: 5.00 μl

LC-MS / MS

Ion source: Turbo Ion Spray (500 &lt; 0 &gt; C)

Polarity: Positive

Carbamazepine (IS) = m / z 237 > 194 (Retention time: 0.74 min), ondansetron = 294 > 170 (Retention time: 0.40 min)

Standard Curve: Analyst 1.4.1, Linear (1 / x ² , noIterate)

Observations included changes in body weight for 7 days, 30 minutes before the last 7 doses of ondansetron alone or three doses of BH and ondansetron composite composition, 30 minutes after administration, 1, 2, 3, 4, 6, , analysis was performed using noncompartmental pharmacokinetics data analyzer program (PK solutions 2.0; Summit, Montrose, CO, USA) with ondansetron concentration (ng / ml), pharmacokinetic index Cmax, Tmax, AUC, t _1/2 and MRT Respectively.

2.3.1. Change in weight

In the case of Ondansetron and three doses of the BH 40, 20 or 10 mg / ml compound composition, significant changes in body weight and body weight were not observed during the experiment compared to the administration of ondansetron 0.1 mg / ml alone (Table 4 4).

Treatment

Periods Ondansetron single formula Ondansetron mixed formulas with BH 0.1 mg / ml 40 mg / ml 20 mg / ml 10 mg / ml Body weights At first administration [A] 238.80 ± 15.02 241.00 ± 8.40 237.80 ± 8.47 239.00 ± 13.34 At last 7th administration [B] 271.00 ± 19.69 278.00 + - 8.15 273.00 ± 15.60 270.00 ± 14.27 Body weight gains  Experimental periods [B] [A] 32.20 ± 6.98 37.00 + - 5.48 35.20 ± 7.53 31.00 ± 2.12

2.3.2. Blood ondansetron  Change in concentration

Ondansetron alone or in three doses of BH and ondansetron combination compositions started to detect ondansetron in the blood from 30 minutes after the last 7 doses of each, and continued to be detected over 3 hours after ondansetron and BH 20 or 10 mg / ml Only 0.10 to 0.15 ng / ml of ondansetron was detected in 4 hours after the last 7 doses, in some experimental animals in the composite composition administration group. In the Ondansetron and BH 40 or 20 mg / ml combination composition administration groups, significant changes in ondansetron concentration were not observed during the pre-collection time compared to the administration of ondansetron alone. In the ondansetron and BH 10 mg / (P < 0.05), a decrease in the ondansetron concentration in the blood was recognized (Fig. 5), compared to the administration of ondansetron alone.

In the Ondansetron and BH 40 mg / ml compound administration group, the ondansetron concentration in blood of 30 minutes after the last 7 doses and 1, 2 and 3 hours after administration was -39.97, -22.16, 67.91 And 119.81% point, respectively.

In the Ondansetron and BH 20 mg / ml compound administration group, the ondansetron concentration in the blood of 30 minutes after the last 7 doses and 1, 2 and 3 hours after administration was -32.55, -33.74, 84.12 And 133.65% point, respectively.

 In the Ondansetron and BH 10 mg / ml compound administration group, the ondansetron concentration in blood of 30 minutes after the last 7 doses and 1, 2 and 3 hours after administration was -51.35 and -52.70, respectively, compared with the administration of ondansetron 0.1 mg / 13.09 and 61.79%, respectively.

2.3.3. Of Tmax  change

Ondansetron showed a Tmax of 0.50 ± 0.00 hr in the ondansetron after the last 7 doses in the combination administration of Ondansetron and BH 40, 20 or 10 mg / ml at three doses, and was similar to 5.00 ± 0.00 hr in the ondansetron alone group (Table 5).

Treatment
PK Parameters Ondansetron single formula Ondansetron mixed formulas with BH 0.1 mg / ml 40 mg / ml 20 mg / ml 10 mg / ml Tmax (hrs) 0.50 0.00 0.50 0.00 0.50 0.00 0.50 0.00 Cmax (ng / ml) 2.87 ± 0.59 1.72 + - 0.67 1.94 + - 0.31 1.40 ± 0.29 ^a AUC _{0 -t} (hr 2.72 0.74 2.30 ± 0.78 2.31 0.41 1.50 + - 0.64 AUC _{0 - inf} (hr 2.85 ± 0.73 2.63 ± 1.01 2.61 + - 0.42 1.68 0.70 t _1/2 (hr) 0.44 ± 0.05 0.89 + 0.23 ^a 0.90 + 0.20 ^a 0.75 + 0.25 ^a MRT _inf (hr) 0.90 + 0.07 1.50 ± 0.26 ^a 1.46 ± 0.23 ^a 1.26 + - 0.37

2.2.4. Cmax  change

In the group with Ondansetron and BH 40 and 20 mg / ml, the plasma Cmax of ondansetron after the last 7 doses was 1.72 ± 0.67 and 1.94 ± 0.31 ng / ml, respectively, and 2.87 ± 0.59 ng / ml in the ondansetron alone group , But the ondansetron blood Cmax was 1.40 ± 0.29 ng / ml after 7 doses of ondansetron and BH 10 mg / ml combination composition, indicating that ondansetron alone (P <0.05) decrease compared to the control group (Table 5).

2.2.5. Changes in AUC

In the group administered with Ondansetron and BH 40, 20 or 10 mg / ml, the blood AUC _{0- t} of ondansetron was 2.30 ± 0.78, 2.31 ± 0.41 and 1.50 ± 0.64 hr · ng / ml, -15.11 and -44.69% points compared with 2.72 ± 0.74 hr · ng / ml of the ondansetron alone group. In addition, ondansetron and BH 40, 20, or 10 mg / ml concentrations of the combination composition showed that the blood AUC _{0 - inf} of ondansetron was 2.63 ± 1.01, 2.61 ± 0.42, and 1.68 ± 0.70 hr · ng / ml, , and -7.62, -8.58, and -41.17% points, respectively, compared with 2.85 ± 0.73 hr · ng / ml in the ondansetron alone group (Table 5).

2.2.6. t _1/2 Change of

Ondansetron showed a t _1/2 of 0.89 ± 0.23, 0.90 ± 0.20, and 0.75 ± 0.29 hr in the ondansetron after the last 7 doses in the combination composition with Ondansetron and three doses of BH 40, 20 or 10 mg / (P <0.05) of the points of 102.48, 105.05 and 71.57%, respectively, compared with 0.44 ± 0.05 hr of the composition administration group (Table 5)

2.2.7. MRT _inf Change of

In the group with Ondansetron and BH 40 or 20 mg / ml, the MRT _inf of ondansetron was 1.50 ± 0.26 and 1.46 ± 0.23 hr after the last 7 doses, respectively, which was higher than that of 0.90 ± 0.07 hr in the ondansetron alone group and eoteuna indicate significant (p <0.05) increase in the 62.48% point, ondansetron and the BH 10 mg / ml concentration of the compound composition is administered ondansetron blood MRT _inf after the final 7th dose observed in 1.26hr, ondansetron alone composition administered group (40.36%), respectively (Table 5).

As a result of Example 2, it was observed that a single administration of ondansetron and BH 40 mg / ml composite composition did not significantly affect the overall bioavailability (AUC) of ondansetron. However, ondansetron and BH 20 or 10 mg / Unidirectional administration of ondansetron 0.1 mg / ml and BH 40, 20 or 10 mg / ml composite composition on day 7, unlike Example 1, which resulted in a reduction in overall oral bioavailability through ondansetron absorption inhibition Repeated oral administration did not significantly affect the overall bioavailability of ondansetron. Therefore, it is expected that the complex composition of BH and ondansetron will provide a new method for treating patients with generalized cancer by using additional pharmacokinetic experiments or toxicity reduction experiments.

Example  3. Nausea and vomiting treatment with ondansetron  blue honeysuckle (BH) Compound preparation  Development: According to BH complex composition ondansetron  Toxicity-reducing effect

3.1. Experimental animal  And group separation

(Average: 34.95 ± 1.36 g, 31.60 to 37.10 g) after 9 days of purification by male ICR mice (Male ICR mice) and SPF / VAF Outbred CrljOri: CD1 [ICR] male mice (OrientBio, Seungnam, Korea) 7 rats were selected and divided into 6 groups (Table 6, Fig. 6).

Total 6 groups (including medium control group); Seven grains per group (total 42 used)

(G3M) ondansetron 0.5 mg / ml and BH 40 mg / ml composite composition administration group, (G4M) medium control group, (G1M) ondansetron 0.5 mg / ml single composition administration group, (G2M) BH 40 mg / ml single composition, ondansetron 0.5 mg / ml and BH 20 mg / ml composite composition administration group, (G5M) ondansetron 0.5 mg / ml and BH 10 mg / ml composite composition administration group

Group Sex Dose (mg / kg) Animal No. Mouse repeated oral dose toxicity test Control Male Distilled water 10ml / kg M01 ~ M07 Reference Male Ondansetron single ( 5 mg / kg ) M08 ~ M14 Reference Male BH single ( 400 mg / kg) M15 to M21 Active Male Ondansetron and BH ( 5 and 400 mg / kg) M22 to M28 Active Male Ondansetron and BH ( 5 and 200 mg / kg) M29 to M35 Active Male Ondansetron and BH ( 5 and 100 mg / kg) M36 to M42

3.2. Purpose of experiment, method of administration and dosage

Oral administration ( oral gavage ; Ondansetron 0.5 mg / ml and BH 40 mg / ml alone were administered as sterile distilled water as a solvent and orally administered once daily for 28 days with a dose of 10 ml / kg, a general oral dose of rodent [Flecknell, 1996] Compositions containing Ondansetron 0.5 mg / ml and BH 40, 20 or 10 mg / ml were orally administered once daily for 28 days, respectively. In the medium control, only the sterile distilled water, medium, Day.

In the Ondansetron alone group, 0.5 mg of ondansetron (CTX Lifesciences (P) Ltd., Gujarat, India) was dissolved per ml of sterilized distilled water, and 40 mg of BH was dissolved per ml of sterilized distilled water. In addition, in the ondansetron and BH complex composition administration groups, BH was additionally dissolved at concentrations of 40, 20 and 10 mg / ml in the ondansetron composition at a concentration of 0.5 mg / ml (Table 6, FIG. 6).

In Examples 1 and 2, single and 7-day repeated oral administration of ondansetron 0.1 mg / ml and BH 40, 20 or 10 mg / ml composite compositions did not significantly affect the overall bioavailability of ondansetron. In this Example 2, the ondansetron toxicity reduction effect according to the complex composition of BH was evaluated using a male mouse as an integrated medical study of ondansetron and BH in general cancer patients. . Dose of Ondansetron is a mouse oral administration the known first median lethal dose (LD ₅₀₎ of 10-30 mg / kg [Hayashi et al , 1993; Khedhaier et al., 2003; GlaxoSmithKline, 2010], and the administration dose of 400, 200 or 100 mg / kg of BH was set on the basis of Examples 1 and 2.

The items of 4-week mortality, clinical symptoms, weight change, and autopsy findings were as follows.

28 days; (14 items; Table 7) and blood chemistry (20 items; Table 8) changes, histopathological changes (23 organ: brain, cerebellum and soft tissue, Heart, thymus, lung, testis, epididymis, kidney, adrenal gland, liver, pancreas, digestive tract esophagus, stomach, pylorus, duodenum, plant, ileum, cecum, colon and rectum, Changes in glutathione (GSH) contents, catalase (CAT) and superoxide dismutase (SOD).

Hematology Items Units Methods Abbreviations Full name 1. RBC Red blood cell count M / L Laser optical (Flow cytometry) 2. HGB Hemoglobin concentration g / dl Cyanmethemoglobin method 3. HCT Hematocrit % Calculated from Item 1 and 4 4. MCV Mean corpuscular volume fL Laser optical (Flow cytometry) 5. MCH Mean corpuscular hemoglobin pg Calculated from Item 1 and 2 6. MCHC Mean corpuscular hemoglobin concentration g / dL Calculated from Item 2 and 3 7. PLT Platelet count K / μL Laser optical (Flow cytometry) 8. RET Reticulocyte count ea / 1000 Laser optical with cytochemical reaction 9. WBC White blood cell count K / μL Laser optical with cytochemical reaction Differential counts of white blood cells 10. NEU%  Percentages of neutrophils % Perox optical with chemical reaction 11. LYM%  Percentages of lymphocytes % Perox optical with chemical reaction 12. MON%  Percentages of monocytes % Perox optical with chemical reaction 13. EOS%  Percentages of eosinophils % Perox optical with chemical reaction 14. BAS%  Percentages of basophils % Perox optical with chemical reaction

Hematology Items Units Methods Abbreviations Full name 1. AST Aspartate aminotransferase IU / L UV-Rate method 2. ALT Alanine aminotransferase IU / L UV-Rate method 3. ALP Alkaline phosphatase IU / L P-NPP method 4. BUN Blood urea nitrogen mg / dL Urease-UV method 5. CRE Creatinine mg / dL Jaffe method 6. GLU Glucose mg / dL Enzyme method 7. CHO Total cholesterol mg / dL Enzyme method 8. PRO Total protein g / dL Biuret method 9. CPK Creatine phosphokinase IU / L UV-Rate method 10. ALB Albumin g / dL BCG method 11. BIL Total bilirubin mg / dL Jendrassik-cleghorn method 12. Globulin Globulin g / dL BCG method 13. A / G Albumin / globulin ratio Ratio Calculated from Item 10 and 12 14. IP Inorganic phosphorus mg / dL UV method 15. Ca Calcium mg / dL OCPC method 16. TG Triglyceride mg / dL Enzyme method 17. LDH  Lactate dehydrogenase IU / L UV-Rate method 18. Na Sodium mmol / L Electrode method 19. K Potassium mmol / L Electrode method 20. Cl Chloride mmol / L Electrode method

3.3. Experiment result

3.3.1. Mortality and clinical symptoms

As a result of Example 3, the deaths associated with the administration of the test substance were not recognized during the 28-day experimental period, and a final autopsy was performed on all the experimental animals (7/7; 100%) in all experimental groups (Table 9).

As a result of Example 3, the clinical symptoms associated with the administration of the test substance were not observed during the 28-day experimental period (Table 10).

Periods
Groups ^{Days of treatment Periods (Day 0 a} ~ 27) At termination (28 days of administration) Total * Vehicle control  Distilled water 0 0 0/7 (0%) Ondansetron single formula 0.5 mg / ml 0 0 0/7 (0%) BH single formula  40 mg / ml 0 0 0/7 (0%) Ondansetron 0.5 mg / ml and BH mixed formula 40 mg / ml 0 0 0/7 (0%) 20 mg / ml 0 0 0/7 (0%) 10 mg / ml 0 0 0/7 (0%)

Signs
Groups Normal appearance Clinical signs Any abnormal signs Vehicle control  Distilled water 7/7 (100%) 0/7 (0%) Ondansetron single formula 0.5 mg / ml 7/7 (100%) 0/7 (0%) BH single formula  40 mg / ml 7/7 (100%) 0/7 (0%) Ondansetron 0.5 mg / ml and BH mixed formula 40 mg / ml 7/7 (100%) 0/7 (0%) 20 mg / ml 7/7 (100%) 0/7 (0%) 10 mg / ml 7/7 (100%) 0/7 (0%)

3.3.2. Changes in body weight and weight gain

Except for the increase in body weight gain during the day 14-27 days (p <0.01) compared to the administration of ondansetron 0.5 mg / ml alone, which was limited to Ondansetron 0.5 mg / ml and BH 10 mg / ml compound composition administration group, And the increase in body weight and weight gain as compared with the single composition administration group were not observed in all the composite composition administration groups. In the BH 40 mg / ml single composition administration group, significant weight gain and weight increase compared with the medium control group and ondansetron 0.5 mg / And changes in weight gain were not observed during the entire period of the experiment (Table 11; Fig. 7).

Intervals
Groups Intervals Day 0 * ~ Day 14 Day 14 ~ Day 27 Day 0 ~ Day 28 ** Vehicle control  Distilled water 7.21 ± 2.44 1.10 ± 1.35 2.56 ± 1.18 Ondansetron single formula 0.5 mg / ml 7.49 + 1.16 0.90 + 0.72 2.34 ± 1.50 BH single formula  40 mg / ml 7.97 ± 1.45 1.49 ± 0.58 3.36 ± 1.11 Ondansetron 0.5 mg / ml and BH mixed formula 40 mg / ml 6.09 ± 0.62 1.03 + - 0.86 2.53 ± 1.22 20 mg / ml 6.33 ± 1.32 1.24 0.95 2.26 ± 1.30 10 mg / ml 7.13 ± 1.02 1.96 + 0.84 ^a 3.50 + - 0.78

3.3.3. Change in long-term weight

In the administration group of Ondansetron 0.5 mg / ml alone, significant increases (p <0.01) in the absolute and relative weight of the heart and decrease in the spleen absolute and relative weights were observed compared to the medium control group. However, all BH 10, 20 or 40 mg / In the ondansetron 0.5 mg / ml composite composition administration group, a significant reduction in cardiac weight and an increase in spleen weight were observed, respectively, compared to the administration of ondansetron 0.5 mg / ml single composition. On the other hand, in the administration group of BH 40 mg / ml alone, there was no significant change in the relative weights and absolute long weights as compared with the medium control group. In the group administered with BH 20 mg / ml and ondansetron 0.5 mg / All ondansetron 0.5 mg / ml and all three types of ondansetron and BH complex composition administration groups showed significantly higher lung, thymus, kidney, adrenal (p <0.05) than control group, , Testis, liver, pancreas, brain, epididymal and subcutaneous lymph nodes were not observed (Table 12 and 13).

Organs
Groups Principal organs Lung Heart Thymus Kidney L Adrenal G L Spleen Vehicle control 0.185 + 0.016 0.162 + 0.011 0.038 ± 0.007 0.285 + 0.021 0.003 0.001 0.093 + 0.011 Ondansetron single formula 0.5 mg / ml 0.186 + 0.014 0.187 ± 0.003 ^a 0.036 0.011 0.283 + 0.032 0.003 0.001 0.068 ± 0.008 ^d BH single formula  40 mg / ml 0.187 + 0.017 0.164 0.011 ^b 0.035 0.008 0.287 + 0.024 0.003 0.001 0.092 0.007 ^f Ondansetron 0.5 mg / ml and BH mixed formula 40 mg / ml 0.183 + 0.012 0.174 ± 0.005 ^ab 0.041 ± 0.009 0.280 + 0.024 0.003 0.001 0.082 0.006 ^ef 20 mg / ml 0.184 + 0.011 0.176 + 0.008 ^ac 0.042 ± 0.008 0.277 + 0.027 0.004 0.002 0.082 ± 0.003 ^df 10 mg / ml 0.187 + 0.017 0.181 ± 0.005 ^a 0.044 0.013 0.289 + 0.033 0.003 0.001 0.081 ± 0.006 ^ef Groups Organs Testis L Liver Pancreas S Brain Epididymis L LN L Vehicle control 0.102 0.029 1.332 + 0.098 0.180 0.020 0.477 + 0.023 0.042 ± 0.007 0.004 0.002 Ondansetron single formula 0.5 mg / ml 0.110 ± 0.007 1.381 + 0.085 0.181 + 0.032 0.480 0.019 0.046 ± 0.004 0.003 0.001 BH single formula  40 mg / ml 0.106 ± 0.007 1.354 + 0.111 0.180 0.024 0.485 + 0.025 0.045 ± 0.005 0.004 0.002 Ondansetron 0.5 mg / ml and BH mixed formula 40 mg / ml 0.112 + 0.013 1.321 + 0.097 0.170 + 0.019 0.474 + 0.017 0.045 ± 0.004 0.003 0.001 20 mg / ml 0.109 0.023 1.365 + 0.090 0.192 + 0.049 0.479 + 0.016 0.048 ± 0.007 0.005 0.003 10 mg / ml 0.108 0.019 1.383 + 0.079 0.181 + 0.021 0.479 + 0.026 0.046 ± 0.004 0.005 0.003

Organs
Groups Principal organs Lung Heart Thymus Kidney L Adrenal G L Spleen Vehicle control 0.583 + 0.066 0.511 + 0.029 0.112 + 0.023 0.898 + 0.093 0.010 ± 0.004 0.292 0.027 Ondansetron single formula 0.5 mg / ml 0.585 ± 0.058 0.588 + 0.028 ^a 0.113 + 0.032 0.892 + 0.131 0.009 0.002 0.214 + 0.028 ^a BH single formula  40 mg / ml 0.572 + 0.044 0.501 0.030 ^c 0.107 0.020 0.879 + 0.080 0.009 0.003 0.281 0.015 ^c Ondansetron 0.5 mg / ml and BH mixed formula 40 mg / ml 0.572 + - 0.052 0.544 0.023 ^bc 0.128 + 0.025 0.874 + 0.087 0.010 ± 0.004 0.255 + 0.023 ^ac 20 mg / ml 0.577 + 0.038 0.552 + 0.025 ^ad 0.133 + 0.027 0.868 ± 0.087 0.012 ± 0.006 0.255 + 0.011 ^ac 10 mg / ml 0.565 + 0.040 0.548 + 0.026 ^bc 0.132 + 0.042 0.871 + - 0.105 0.009 0.003 0.246 0.018 ^ac Groups Organs Testis L Liver Pancreas S Brain Epididymis L LN L Vehicle control 0.322 + 0.091 4.185 ± 0.192 0.567 + 0.055 1.501 + 0.090 0.131 + 0.020 0.011 ± 0.007 Ondansetron single formula 0.5 mg / ml 0.343 + 0.016 4.337 + 0.337 0.563 + 0.073 1.509 + 0.124 0.144 + 0.016 0.009 0.004 BH single formula  40 mg / ml 0.325 + 0.027 4.145 ± 0.319 0.550 + 0.065 1.483 + 0.038 0.139 + 0.017 0.013 ± 0.007 Ondansetron 0.5 mg / ml and BH mixed formula 40 mg / ml 0.349 + 0.048 4.121 0.328 0.531 + - 0.056 1.477 + 0.062 0.141 + 0.014 0.008 ± 0.004 20 mg / ml 0.342 + 0.077 4.269 + 0.233 0.601 + - 0.158 1.497 + 0.029 0.150 0.024 ^b 0.015 ± 0.009 10 mg / ml 0.324 + 0.050 4.171 + 0.172 0.545 + 0.047 1.445 ± 0.091 0.139 + 0.009 0.014 ± 0.009

3.3.4. Hematological change

As a result of 14 haematological tests, a decrease in the WBC and lymphocyte ratio and an increase in the ratio of neutrophilic leukocytes associated with ondansetron 0.5 mg / ml alone group (p <0.01) / ml and BH 40, 20 or 10 mg / ml, respectively, were associated with a marked increase in the WBC and lymphocyte ratio and a corresponding decrease in the neutrophil leukocyte ratio compared with the administration of ondansetron 0.5 mg / ml alone. On the other hand, significant hematologic changes were not observed in the administration group of BH 40 mg / ml alone compared with the medium control group, and significant effects were observed in the ondansetron 0.5 mg / ml single composition and all three kinds of ondansetron and BH complex administration groups Changes in RBC, HGB, HCT, MCV, MCH, MCHC, PLT, RET, MONO%, EOS% and BAS% were not observed (Table 14).

Items
Groups Hematological Items: Red Blood Cells RBC HGB HCT MCV MCH MCHC PLT RET Vehicle control 8.47 ± 0.66 18.31 + - 1.52 46.80 ± 1.69 47.76 ± 2.60 19.50 ± 1.49 38.04 + - 2.90 907.57 ± 233.45 0.51 ± 0.20 Ondansetron single formula 0.5 mg / ml 8.51 + - 0.62 18.50 + - 0.58 46.89 ± 1.63 47.80 ± 1.74 19.64 ± 1.23 38.14 1.56 902.86 + 261.85 0.50 0.26 BH single formula  40 mg / ml  8.53 ± 0.53 18.27 ± 0.52 45.67 + - 4.20 47.56 1.84 19.81 ± 1.80 38.27 + 1.81 894.71 + 267.38 0.49 0.25 Ondansetron 0.5 mg / ml and BH mixed formula 40 mg / ml  8.50 ± 0.50 18.39 ± 0.69 46.14 ± 2.29 47.70 ± 1.91 19.69 + - 0.95 38.36 + 1.83 910.14 + - 221.63 0.49 + 0.13 20 mg / ml 8.34 ± 0.71 18.36 ± 0.97 46.84 ± 2.10 48.04 0.88 19.56 ± 1.66 38.01 + - 1.61 929.71 + - 168.17 0.56 ± 0.27 10 mg / ml  8.43 + - 0.57 18.39 ± 0.64 46.76 ± 1.81 47.69 ± 1.31 19.41 ± 1.25 38.07 ± 1.44 904.57 ± 160.99 0.53 + - 0.29 Items
Groups Hematological Items: White Blood Cells WBC NEU (%) LYM (%) MONO (%) EOS (%) BASO (%) Vehicle control 5.33 ± 1.00  8.56 ± 1.73 85.41 + 1.41 2.36 ± 0.71 0.83 ± 0.77 0.97 + 0.64 Ondansetron single formula 0.5 mg / ml 2.75 ± 0.45 ^a 18.59 ± 2.52 ^a 73.20 ± 2.06 ^a 2.27 + - 0.85 0.81 ± 0.19 1.13 + - 0.85 BH single formula  40 mg / ml 5.25 ± 1.48 ^b 8.90 ± 2.41 ^b 85.67 ± 2.40 ^b 2.47 ± 0.63 0.80 + - 0.23 0.84 0.74 Ondansetron 0.5 mg / ml and BH mixed formula 40 mg / ml 3.80 ± 0.62 ^ac 12.36 ± 1.26 ^ab 81.73 ± 1.66 ^ab 2.31 ± 0.55 0.89 + 0.64 0.96 0.32 20 mg / ml 3.50 ± 0.52 ^a 13.23 ± 0.50 ^ab 81.13 ± 1.63 ^ab 2.27 ± 0.39 0.77 + - 0.72 0.84 + 0.64 10 mg / ml 3.43 ± 0.51 ^a 13.69 ± 1.29 ^ab 80.60 ± 1.43 ^ab 2.31 0.47 0.83 0.26 0.89 ± 0.65

3.3.5. Blood biochemical change

Twenty blood biochemical tests showed no biochemical changes associated with ondansetron, BH, and ondansetron and BH combination administration (Table 15).

Items
Groups Serum Biochemical Items AST ALT ALP BUN CRE GLU CHO Vehicle control 66.29 ± 16.22 33.43 + - 6.55  99.43 + - 15.71  20.69 + - 2.33  0.57 + 0.14  96.71 ± 15.99  154.00 ± 22.20 Ondansetron single formula 0.5 mg / ml  66.86 ± 15.20  36.71 ± 11.12  92.14 + - 14.69  20.87 ± 3.70  0.53 + - 0.14  97.57 ± 10.75  154.43 + - 21.07 BH single formula  40 mg / ml  60.00 ± 15.44  32.29 + - 5.38 101.43 ± 25.30  20.60 ± 1.58  0.51 + - 0.16  97.14 ± 19.68  155.29 ± 18.43 Ondansetron 0.5 mg / ml and BH mixed formula 40 mg / ml  66.43 + - 13.70  35.29 ± 12.08  96.29 + 19.66  20.69 ± 1.52  0.59 + 0.13  95.00 ± 12.40  156.29 ± 23.26 20 mg / ml  65.14 ± 17.99  34.43 + - 5.71  88.14 +/- 12.77  19.91 + 1.69  0.47 + 0.25  98.14 + - 7.95  149.29 ± 19.24 10 mg / ml  67.86 ± 17.37  36.29 ± 10.63  96.14 + 24.84  20.76 ± 2.31  0.59 0.22  98.00 ± 17.59  157.14 ± 19.79 Groups Items PRO CPK BIL ALB Globulin A / G TG Vehicle control  5.31 ± 1.04  194.29 + - 41.47 0.13 ± 0.05  2.34 ± 0.36  2.97 ± 0.87  0.84 0.23 229.29 + - 39.19 Ondansetron single formula 0.5 mg / ml  5.43 ± 1.11  196.86 ± 21.75 0.13 ± 0.05  2.24 ± 0.53  3.19 ± 1.37  0.96 + 0.78 225.29 ± 27.96 BH single formula  40 mg / ml  5.20 ± 0.50  204.00 ± 17.17 0.14 ± 0.05  2.31 ± 0.39  2.89 ± 0.73  0.88 0.37 233.57 ± 30.21 Ondansetron 0.5 mg / ml and BH mixed formula 40 mg / ml  5.56 ± 1.19  199.71 + - 47.89 0.13 ± 0.05  2.30 ± 0.86  3.26 ± 1.31  0.84 0.43 227.57 + - 43.53 20 mg / ml  5.40 + - 0.72  181.57 ± 35.08 0.13 + 0.08  2.33 ± 0.31  3.07 ± 0.73  0.80 + - 0.25 229.29 ± 32.64 10 mg / ml  5.26 ± 0.99  193.29 ± 20.48 0.14 ± 0.05  2.34 ± 0.24  2.91 ± 1.05  0.89 0.31 221.57 ± 25.34 Groups Items LDH (x 10 ² ) Ca P Na K Cl Vehicle control 11.17 + - 4.17 10.86 0.95 11.56 ± 1.19  141.71 + - 2.14 10.57 ± 0.26  113.43 + - 2.70 Ondansetron single formula 0.5 mg / ml 10.46 + - 3.02 10.91 ± 0.91 11.63 + - 0.95  141.57 + - 2.44 10.63 ± 0.33  113.86 ± 1.35 BH single formula  40 mg / ml 10.28 ± 2.13 11.00 + 0.66 11.87 ± 0.87  141.57 1.51 10.59 + - 0.33  113.43 + - 2.37 Ondansetron 0.5 mg / ml and BH mixed formula 40 mg / ml 11.99 ± 3.43 10.89 ± 1.17 11.53 + - 1.36  141.29 ± 1.70 10.56 ± 0.11  113.29 ± 2.14 20 mg / ml  9.89 ± 1.14 10.99 ± 1.07 11.59 ± 1.27  141.29 ± 2.21 10.61 ± 0.20  113.57 ± 1.90 10 mg / ml 11.57 + - 3.61 10.81 ± 0.56 11.39 ± 0.83  141.29 ± 3.64 10.54 ± 0.26  113.57 ± 3.26

3.3.6. Autopsy findings

Ondansetron 0.5 mg / ml and BH 10, 20 or 40 mg / ml compound administration group showed significant cardiac hypertrophy and increased incidence of spleen atrophy compared to the medium control group. However, ondansetron Significant cardiac hypertrophy and decreased incidence of spleen atrophy were observed in each group compared to the 0.5 mg / ml single composition administration group. At the same time, pulmonary hyperemia of the minor [1+] or moderate [2+], thymic atrophy, splenic enlargement, testicular and epididymal atrophy, and sublingual lymph node enlargement were sporadically observed in all experimental groups including the medium control group (Table 16).

Groups

Organs / Findings Vehicle control Single from BH
Single 0.5 mg / ml and BH mixed formula 0.5 mg / ml 40 mg / ml 40 mg / ml 20 mg / ml 10 mg / ml Lung
Normal
Congestion
1+ 5/7
2/7
2/7 6/7
1/7
1/7 7/7
0/7
0/7 6/7
1/7
1/7 6/7
1/7
1/7 6/7
1/7
1/7 Heart
Normal
Hypertrophy
1+
2+ 7/7
0/7
0/7
0/7 0/7
7/7
1/7
6/7 7/7
0/7
0/7
0/7 3/7
4/7
4/7
0/7 2/7
5/7
5/7
0/7 2/7
5/7
3/7
2/7 Thymus
Normal
Atrophy
1+ 6/7
1/7
1/7 6/7
1/7
1/7 6/7
1/7
1/7 7/7
0/7
0/7 7/7
0/7
0/7 6/7
1/7
1/7 Spleen
Normal
Hypertrophy
1+
Atrophy
1+
2+ 4/7
2/7
2/7
1/7
1/7
0/7 0/7
0/7
0/7
0/7
5/7
2/7 5/7
1/7
1/7
1/7
1/7
0/7 4/7
0/7
0/7
3/7
3/7
0/7 3/7
0/7
0/7
4/7
4/7
0/7 3/7
0/7
0/7
4/7
4/7
0/7 Testis
Normal
Atrophy
2+ 6/7
1/7
1/7 7/7
0/7
0/7 7/7
0/7
0/7 7/7
0/7
0/7 7/7
0/7
0/7 7/7
0/7
0/7 Epididymis
Normal
Atrophy
2+ 6/7
1/7
1/7 7/7
0/7
0/7 7/7
0/7
0/7 7/7
0/7
0/7 7/7
0/7
0/7 7/7
0/7
0/7 Lymph node ^a)
Normal
Hypertrophy
1+ 5/7
2/7
2/7 6/7
1/7
1/7 6/7
1/7
1/7 7/7
0/7
0/7 6/7
1/7
1/7 6/7
1/7
1/7 Others
Normal 7/7 7/7 7/7 7/7 7/7 7/7

3.3.7. Histopathological observation

In the control group and the BH 40 mg / ml single composition administration group, mild spleen white watery lymphocyte reduction findings (Fig. 8) were observed only in 1 case (1/7; 14.29%) respectively. However, in the case of ondansetron 0.5 mg / (Fig. 9) and decreased spleen white water quality lymphocyte (Fig. 8), and increased frequency of observation, to various degrees (mild, moderate or severe [3+]) compared to the medium control group. In contrast, ondansetron and BH 10, 20 or 40 mg / ml compound administration group showed marked reductions in the degree and frequency of these cardiac muscle fiber local necrosis and spleen white water lymphocyte reduction findings compared to the administration of ondansetron 0.5 mg / ml alone It was acknowledged. (Fig. 10), splenic red water lymphocyte enlargement (Fig. 8), testicular tubular atrophy (Fig. 11), inflammatory cell infiltration with liver necrosis (Fig. 12), abnormal epididymal sperm (Fig. 13), diffuse lymphocytic lymphocyte enlargement (Fig. 14) or supraglottic cyst formation (Fig. 15) were sporadically observed in all experimental groups including the medium control group. In addition, only 1 case (1/7; 14.29%) of pyloric cystic formation (Fig. 16) was noted in the ondansetron and BH 40 mg / ml compound administration group (Table 17).

Groups Vehicle control From him
Single BH
Single
40 mg / ml 0.5 mg / ml and BH mixed formula 0.5 mg / ml 40 mg / ml 20 mg / ml 10 mg / ml Lung
Normal
CG ^† 1+ 5/7
2/7 6/7
1/7 7/7
0/7 6/7
1/7 6/7
1/7 6/7
1/7 Heart
Normal
fFN ^† 1+
2+
3+ 7/7
0/7
0/7
0/7 0/7
1/7
5/7
1/7 7/7
0/7
0/7
0/7 3/7
4/7
0/7
0/7 1/7
5/7
1/7
0/7 0/7
3/7
4/7
0/7 Spleen
Normal
rHP ^† 1+
wDE ^† 1+
2+ 4/7
2/7
1/7
0/7 0/7
0/7
4/7
3/7 5/7
1/7
1/7
0/7 3/7
0/7
4/7
0/7 3/7
0/7
4/7
0/7 2/7
0/7
5/7
0/7 Testis
Normal
TA ^† 2+ 6/7
1/7 7/7
0/7 7/7
0/7 7/7
0/7 7/7
0/7 7/7
0/7 Liver
Normal
IF-FN ^† 1+ 6/7
1/7 7/7
0/7 7/7
0/7 7/7
0/7 7/7
0/7 7/7
0/7 Epididymis
Normal
AS-OS ^† 2+ 6/7
1/7 7/7
0/7 7/7
0/7 7/7
0/7 7/7
0/7 7/7
0/7 Lymph node ^a)
Normal
dHP ^† 1+
2+
5/7
1/7
1/7
6/7
1/7
0/7
6/7
1/7
0/7
7/7
0/7
0/7
6/7
1/7
0/7
6/7
1/7
0/7 Fundus
Normal
fCY ^† 1+ 6/7
1/7 6/7
1/7 5/7
2/7 6/7
1/7 6/7
1/7 7/7
0/7 Pylorus
Normal
fCY ^† 1+ 7/7
0/7 7/7
0/7 7/7
0/7 6/7
1/7 7/7
0/7 7/7
0/7 Others
Normal 7/7 7/7 7/7 7/7 7/7 7/7

^† CG = congestions; fFN = focal myocardial necrosis; rHP = hyperplasia of red pulp lymphoid cells; wDE = white pulp lymphoid cell; TA = focal semeniferous tubular atrophy; IF = focal inflammatory cell infiltrations; FN = focal cellular necrosis; AS-OS = Oligospermatozoa with abnormal sperm exists; dHP = diffused hyperplasia of lymphoid cells; fCY = focal cyst formation in the mucosa

3.3.8. Changes in heart lipid peroxidation and antioxidant defense systems

In the administration of Ondansetron 0.5 mg / ml alone, the increase in cardiac lipid hepatotoxicity and the reduction of endogenous antioxidants or related enzymes, GSH, SOD and CAT, were significantly (p <0.01) , 20 or 40 mg / ml composite composition administration group, the increase of GSH content or SOD and CAT activity was recognized to be dependent on the complexed BH concentration, respectively, as compared with the administration of ondansetron 0.5 mg / ml alone composition. On the other hand, significant changes in cardiac lipid peroxidation and antioxidant defense system were not observed in the group administered with BH 40 mg / ml alone (Table 18).

Cardiac lipid peroxidation was 147.39% in the ondansetron 0.5 mg / ml single administration group compared with the vehicle control group. Ondansetron and BH 10, 20 or 40 mg / mg / ml compared to the control group, -62.00, -45.11, -31.94 and -20.50%, respectively.

The concentration of GSH in the cardiac tissue was -57.75% as compared with the control group in the administration of ondansetron 0.5 mg / ml alone, and the administration of BH 40 mg / ml alone, ondansetron and BH 10, 20 or 40 mg / Compared with those treated with ondansetron 0.5 mg / ml alone, 170.86, 77.10, 62.26 and 37.53%, respectively.

The CAT activity in cardiac tissue was -70.20% compared with the control group in the administration of ondansetron 0.5 mg / ml alone, and the administration of BH 40 mg / ml alone, ondansetron and BH 10, 20 or 40 mg / Showed a change of 297.37, 102.63, 69.74 and 48.68%, respectively, compared to the administration of ondansetron 0.5 mg / ml alone.

The SOD activity in cardiac tissue was -67.52% compared with the control group in the administration of ondansetron 0.5 mg / ml alone, and the administration of BH 40 mg / ml alone, ondansetron and BH 10, 20 or 40 mg / Compared to those treated with ondansetron 0.5 mg / ml alone, 267.97, 129.41, 99.35 and 43.79%, respectively.

Items
(Unit)
Groups LPO Antioxidative Defense Systems MDA levels
(nM / g protein) Glutathione
(μM / mg tissue) CAT
(U / g protein) SOD
(U / g protein) Vehicle control  Distilled water 1.50 + - 0.32 314.43 + - 83.26  0.36 + 0.12 6.73 ± 1.36 Ondansetron single formula 0.5 mg / ml 3.72 ± 0.66 ^c 132.86 ± 36.81 ^a 0.11 0.04 ^c 2.19 ± 0.57 ^c BH single formula  40 mg / ml 1.41 ± 0.43 ^e 359.86 ± 58.31 ^b 0.43 ± 0.09 ^e 8.04 ± 2.79 ^e Ondansetron 0.5 mg / ml and BH mixed formula 40 mg / ml 2.04 ± 0.29 ^de 235.29 ± 29.14 ^ab 0.22 ± 0.05 ^de 5.01 ± 1.08 ^de 20 mg / ml 2.53 ± 0.33 ^ce 215.57 ± 22.30 ^ab 0.18 ± 0.03 ^ce 4.36 ± 0.87 ^ce 10 mg / ml 2.96 + 0.16 ^ce 182.71 ± 31.29 ^a 0.16 0.02 ^cf 3.14 ± 0.50 ^cf

^a p <0.01 as compared with vehicle control by LSD test

^b p <0.01 as compared with ondansetron single formula by LSD test

^c p <0.01 and ^d p <0.05 compared with vehicle control by MW test

^e p <0.01 and ^f p <0.05 as compared with ondansetron single formula by MW test

In Example 3, to evaluate the ondansetron toxicity reduction effect according to the complex composition of BH, male mice were used as an integrated medical study of ondansetron and BH in patients with general cancer patients.

According to Korea Food & Drug Administration Notification No. 2015-082 [KFDA, 2015], it is recommended that a dose of 20 ml or less per kg of body weight is recommended for mouse repeat oral toxicity test. Generally, If the material is well dissolved in the medium, administer a dose of 10 ml / kg. In this Example 3, the ondansetron dose was 10-30 mg / kg (Hayashi et al., 1993; Khedhaier et al., 2003; GlaxoSmithKline, 2010], and the complex composition concentration of 40, 20 or 10 mg / ml of BH was set on the basis of Examples 1 and 2. In Example 3, an appropriate dose of BH (40, 20 or 10 mg / ml) or ondansetron (0.5 mg / ml) was dissolved in sterilized distilled water in the form of a single or complex composition, / kg dose [Flecknell, 1996] once a day for 28 days.

As a result of Example 3, it was confirmed that the ondansetron 0.5 mg / ml or BH 40 mg / ml single composition, the deaths and the clinical symptoms associated with administration of ondansetron 0.5 mg / ml and BH 40, 20 or 10 mg / However, in the administration of ondansetron 0.5 mg / ml alone, the increase in cardiac weight, decrease in spleen weight, decrease in WBC and LYM% and increase in NEU% were recognized. On visual and histopathological examination, There was an increase in lipid peroxidation in cardiac tissue and a decrease in GSH, SOD and CAT content or activity of antioxidant defense substances, respectively. The ondansetron-induced cardiotoxicity and immunodeficiency findings were significantly inhibited by the complex composition of BH 10, 20 or 40 mg / ml. Compared with the administration of ondansetron 0.5 mg / ml alone, significant reductions in cardiac lipid peroxidation and antioxidant defense system Respectively. Therefore, as a result of Examples 1 and 2 of the present inventors, it was observed that the complex composition of BH did not affect the bioavailability of ondansetron, and the administration of the ondansetron and BH complex compositions did not affect ondansetron bioavailability , Already well-known immunomodulation [Jin et al., 2006; Zdarilova et al., 2010] and antioxidant [Chen et al., 2014], it was concluded that the effects of ondansetron and BH on general cancer patients The composite composition is expected to provide a novel therapeutic method which is very useful for both oriental medicine and treatment. On the other hand, significant clinical symptoms, hematologic, hematologic, biochemical, gross autopsy and histopathological changes were not observed in the BH 40 mg / ml single composition administration group compared to the medium control group.

The mice of the vehicle control group, ondansetron 0.5 mg / ml and BH 40 mg / ml alone composition administration group, all three types of ondansetron 0.5 mg / ml and BH 40, 20 or 10 mg / The age-normal mouse weight gain category [Plata and Murphy, 1972; (P <0.001). In addition, the ondansetron 0.5 mg / ml and BH 10 mg / ml administration groups showed a significant increase in weight gain (p < 0.01). The increase in body weight and weight gain was not observed in all compound administration groups, compared with vehicle control and single composition administration groups, except for the increase in body weight during 14 to 27 days. In the administration group of BH 40 mg / ml alone, And ondansetron 0.5 mg / ml alone, significant changes in body weight and body weight were not observed during the pre-experiment period. On the other hand, the increase in body weight of Day 14-27 days (p <0.01) compared to the administration of ondansetron 0.5 mg / ml alone and the approved dose of ondansetron 0.5 mg / ml and BH 10 mg / It is considered that it is difficult to regard it as a change associated with administration of the composite composition.

Recently, cardiac toxicity problems have been raised, including QT prolongation problems as part of serious cardiac side effects by ondansetron, and their use has been limited [Charbit et al., 2008; McKechnie and Froese, 2010; Hafermann et al., 2011]. Also in the results of Example 3, 28 days repeated oral administration of ondansetron 0.5 mg / ml single composition showed significant increase in absolute and relative weight of the heart and hypertrophy accompanied by localized necrosis of heart muscle fibers as compared with the medium control group , Decrease in the content of endogenous antioxidant GSH [Odabasoglu et al., 2006] and decrease in activity of antioxidative enzymes SOD and CAT [Cheeseman and Slater, 1993], along with an increase in harmful lipid peroxidation in cardiac tissues [Comporti, 1985] . Ondansetron-induced cardiac toxicity was significantly inhibited by ondansetron 0.5 mg / ml and BH 40, 20 and 10 mg / ml combination administration, and especially ondansetron 0.5 mg / ml alone Significant reductions in cardiac lipid peroxidation, an increase in GSH content, and an increase in SOD and CAT activity were found to be dependent on the BH concentration in the ondansetron 0.5 mg / ml and BH 40, 20 and 10 mg / ml combination compositions, respectively. Therefore, it is concluded that the BH complex composition of 10 mg / ml or more significantly mitigates the cardiac toxicity of ondansetron by activation of the antioxidant defense system of the heart.

Serotonin (5-HT) has been shown to play a pivotal role in immunomodulating activity [Mossner and Lesch, 1998], at least four of which are known to be immunomodulatory: 1) T cells and natural killer cells 2) production of chemotactic factors, 3) regulation of delayed hypersensitivity, and 4) control of congenital immune response by macrophages. In addition, at least four 5-HT1A, 5-HT1B / 1D, 5-HT2 and 5-HT3 receptors have been found to be present in immunocompetent cells [ Grimaldi and Fillion, 2000], 5-HT3 receptors are involved in the degranulation of mast cells and are involved in the regulation of immune responses [Castex et al., 1994]. Therefore, ondansetron, a 5-HT3 antagonist, has also been shown to block immunostimulation of 5-HT through blockade of 5-HT3 receptors and to induce immunosuppression [Martin et al., 2000; Dubayle et al., 2005]. In particular, 5-HT3 receptors are also present in neurons in most central nervous system, except the diencephalon [Morales et al., 1998], and mast cells that regulate the immune response by being influenced by 5-HT, Many are present in tissues and blood vessels [Dubayle et al., 2005]. The histopathological examination revealed that the spleen atrophy observed in the administration group of ondansetron 0.5 mg / ml alone in Example 3 was caused by the decrease in the splenic white water quality lymphocytes, and the decrease in spleen absolute and relative weight was also recognized. Hematologic tests also showed a significant decrease in LYM% and a relative increase in NEU% associated with a significant decrease in WBC in the ondansetron 0.5 mg / ml single administration group. These results imply typical lymphocyte immunosuppression findings [Sodikoff, 1995]. On the other hand, lymphocyte-reducing immunosuppression by 5 mg / kg ondansetron was significantly inhibited by the BH 40, 20 and 10 mg / ml complex composition, and the BH complex composition of 10 mg / It is considered that the findings are also significantly reduced. Changes in RBC, HGB, HCT, MCV, MCH, MCHC, PLT, RET, MONO%, EOS% and BAS% were not observed in all experimental groups and 20 blood biochemical tests , Blood biochemical changes associated with administration of ondansetron, BH and ondansetron and BH combination compositions were not observed and blood biochemical changes in the range of normal mice of the same age [Wolford et al., 1986] .

Mild or moderate pulmonary congestion, splenic red cell lymphoma enlargement, testicular lymphadenopathy, hyperplastic lymphadenopathy, glandular atrophy, spleen enlargement, testicular and epididymal atrophy, sublingual lymph node enlargement and histopathologic examination Spermatocyte infiltration, inflammatory cell infiltration with liver necrosis, spermatogenesis accompanied by abnormal spermatogenesis in the epididymal duct, diffuse lymphocytic enlargement of the lymph node, or localized cystic formation of the supratentorial duct were observed sporadically in all experimental groups including the medium control group , It is thought to be an accidental lesion, not a toxic symptom due to the administration of the test substance, and these symptoms are rarely observed in normal mice [Choi et al., 2014; Kim et al., 2015]. In addition, only one case (1/7; 14.29%) of Ondansetron and BH 40 mg / ml combination composition was observed, and the incidence of recognized pyloric cystic formation was too low to be regarded as meaningful change related to administration of the composite composition .

In Example 3, it was observed that the complex composition of BH 40, 20 or 10 mg / ml significantly suppressed ondansetron-induced cardiac toxicity and immune disorders through the immunomodulatory and antioxidative effects of BH itself. Therefore, the BH complex composition of 10 mg / ml or more was judged to significantly reduce ondansetron-induced cardiac toxicity and immune disorders through the activity of the immunological activity and antioxidant defense system without affecting the bioavailability of ondansetron, It is anticipated that it will provide a very useful new amount of herbal medicine for cancer patients.

Example  4. Nausea and vomiting for chemotherapy  remedy Ondansetron  ( ZofranTM ) And blue honeysuckle (BH): Evaluation of synergistic effects of cisplatin-induced gastrointestinal disorders on rats

In Example 4, the effect of the complex composition of BH on the pharmacological effect of ondansetron was investigated by using cisplatin-induced gastrointestinal motility disorder rats as a part of herbal medicine integrated medical research on the side effects of chemotherapy for general cancer patients, Respectively. In Example 4, ondansetron 0.1 mg / ml and BH 40 mg / ml alone composition, ondansetron 0.1 mg / ml, and BH 40, 20 were administered intraperitoneally to cisplatin 2 mg / Or 10 mg / ml combination composition was orally administered for 14 days and the effect on gastrointestinal motility, gastrointestinal mucosal injury and physiology of endocrine cells of the digestive tract were observed together with antioxidant effect. Experimental results in Example 4 were compared with the oral administration group of ondansetron 0.1 mg / ml alone. All candidate substances were dissolved in sterilized distilled water, and the dose of 10 ml / kg, which is the general oral dose of the rat, Orally for 14 days (Table 19, Fig. 17).

4.1. Experimental Method

4.1.1. Experimental animals and Group separation

100 healthy male VAF / SPF outbred Crl: CD [SD] male rats were obtained and refined for 7 days. After 3 weeks of cisplatin administration, the body weight (normal control: mean 379.63 ± 13.98 g, 361-407 g; cisplatin Treated group: average 354.31 ± 7.89 g, 341 ~ 368 g) were used for the experiment. All experimental animals were fasted for 18 hours at the start of cisplatin treatment, on the day of drug administration, and on the final autopsy day (negative water was also freely supplied during this period), and individuals were identified with picric acid.

Experimental animals : SPF / VAF outbred Crl: CD [SD] rats (6 week old male, OrientBio, Seungnam, Korea) [ANNEX I, II].

Group separation ( total of 7 groups: 8 per group)

(1) Intact vehicle control: Oral administration of physiological saline peritoneal fluid and sterile distilled water Normal medium control

(2) Cisplatin control: Cisplatin Oral administration of sterilized distilled water

(3) O1: Oral administration of Cisplatin peritoneal and ondansetron 0.1 mg / ml alone composition

(4) BH400: Oral administration of Cisplatin peritoneum and BH 40 mg / ml alone composition

(5) O + BH 400: Oral administration of Cisplatin peritoneal and ondansetron 0.1 mg / ml and BH 40 mg / ml composite composition

(6) O + BH 400: Oral administration of Cisplatin peritoneal and ondansetron 0.1 mg / ml and BH 20 mg / ml composite composition

(7) O + BH 400: Oral administration of Cisplatin peritoneal and ondansetron 0.1 mg / ml and BH 10 mg / ml composite composition

Group Inducer treatment Test substance treatment (mg / kg / day) Animal No. Effects on cisplatin-induced GI disorders in rats Control Saline 1 ml / kg Distilled water 10 ml / kg R01 ~ R08 Control Cisplatin 2 mg / kg Distilled water 10 ml / kg R09 to R16 Reference Cisplatin 2 mg / kg Ondansetron ( 0.1 mg / ml) single formula R17 to R24 Reference Cisplatin 2 mg / kg BH ( 40 mg / ml) single formula R25 to R32 Active Cisplatin 2 mg / kg Ondansetron and BH ( 0.1 and 40 mg / ml, mixed) R33 to R40 Active Cisplatin 2 mg / kg Ondansetron and BH ( 0.1 and 20 mg / ml, mixed) R41 to R48 Active Cisplatin 2 mg / kg Ondansetron and BH ( 0.1 and 10 mg / ml, mixed) R49 to R56

4.1.2. Induction of gastrointestinal motility disorder

Vera et al. [2007] and Song et al. Cis-diaminedichloroplatinum II (cisplatin; Sigma-Aldrich, St. Louis, Mo., USA) was dissolved in physiological saline at a dose of 1 ml / In the case of normal control group, the same dose of physiological saline was administered instead of cisplatin by intraperitoneal administration in the same manner.

4.1.3. drug injection

Ondansetron 0.1 mg / ml and BH 40 mg / ml single composition, ondansetron 0.1 mg / ml and BH 40, 20 or 10 mg / ml combination composition were orally administered once a day for 14 days from 3 weeks after the start of cisplatin administration, In the cisplatin medium control, sterile distilled water, medium instead of the candidate substance, was orally administered at a dose of 10 ml / kg for the same period.

In the Ondansetron alone group, 0.1 mg of ondansetron (CTX Lifesciences (P) Ltd., Gujarat, India) was dissolved per ml of sterilized distilled water and 40 mg of BH per ml of sterilized distilled water was dissolved in the BH alone composition. In addition, in the ondansetron and BH complex composition administration groups, BH was additionally dissolved at concentrations of 40, 20 and 10 mg / ml in the ondansetron composition at a concentration of 0.1 mg / ml. All BH and ondansetron alone compositions, the ondansetron and BH complex compositions, were dissolved in sterile distilled water and dosed at a dose of 10 ml per kilogram of body weight (10 ml / kg) of the animal, the usual repeat oral dose of the rat [Flecknell, 1996] Orally injected using a 5 ml syringe. Based on the results of Examples 1 to 3 used in the fourth embodiment.

4.1.4. Observations

Changes in body weight, fecal index (fraction, fecal weight, and fecal water content), colon histopathologic changes including residual fecal pellet, fecal surface To evaluate the effect of mucus production on the mucous membrane thickness and mucous membrane per mm ² and the mean mucosal thickness on the constipation and to evaluate the effects on the mucosal damage of the digestive tract and on the physiology of endocrine cells in the digestive tract, , Changes in pylorus and colon mucosal thickness and damaged area ratio, changes in gastrin and serotonin immune response cells in pyloric gut mucosa, changes in serotonin immune response cells in colonic mucosa, serotonin contents, serotonin metabolism related enzymes - tryptophan hydroxylase and monoamine oxidase activity were observed, and the antioxidative effect was observed Glutathione (GSH), superoxide dismutase (SOD) and catalase (CAT) contents were measured and the levels of malondialdehyde (MDA) were measured to observe lipid peroxidation 19 and 20, Fig. 17).

Antisera or detection kits Code Source Dilution Primary antisera * Anti-gastrin polyclonal antibody 20055 Immunostar, Hudson, WI, USA 1: 2,000 Anti-serotonin polyclonal antibody AB938 Chemicon, Billerica, MA, USA 1: 1,000 Detection kits Vectastain Elite ABC Kit PK-6200 Vector Lab. Inc., CA, USA 1:50 Peroxidae substrate kit SK-4100 Vector Lab. Inc., CA, USA 1:50

4.2. Experiment result

4.2.1. Changes in body weight and weight gain

Significant weight loss (p <0.01) was observed in the cisplatin control group compared to the normal control group after 14 days of cisplatin administration, that is, 1 week before the administration of cisplatin As a result, the body weight gain during the 5-week experimental period and the 2-week administration period was significantly (p <0.01) significantly lower than that of the normal medium control group. Ondansetron (0.1 mg / ml) and BH (40 mg / ml) alone were significantly (p <0.01) higher than the cisplatin control group in all three concentrations of BH 40,20 and 10 mg / ml and ondansetron 0.1 mg / Body weight gain was observed from the 7th day after the start of the drug administration, and the body weight gain during the experimental period and the drug administration period also showed a significant increase (p <0.01) compared with the cisplatin control. In particular, ondansetron 0.1 mg / ml and BH 40, 20 and 10 mg / ml were significantly increased (p <0.01 or p <0.05) compared to the administration of ondansetron 0.1 mg / ml alone Or 13 days after the administration of the ondansetron 0.1 mg / ml, and the amount of gain during the experimental period and the drug administration period was also significantly increased (p <0.01) compared to the administration of the ondansetron 0.1 mg / ml single composition (Table 21, Fig. 18).

Ondansetron 0.1 mg / ml and BH 40 mg / ml alone administration group, ondansetron 0.1 mg / ml and BH 40 and 20, respectively, showed a change of -120.21% compared with the normal medium control group in the cisplatin control group Or 10 mg / ml of the composite composition showed a change of 203.61, 281.87, 385.35, 342.74 and 315.78%, respectively, compared to the cisplatin control group.

Ondansetron 0.1 mg / ml and BH 40 mg / ml were administered alone, and ondansetron 0.1 mg / ml and BH 40, respectively, in the cisplatin control group compared to the normal medium control group. , And 20 or 10 mg / ml, respectively, compared with the cisplatin control group, 26.73, 36.90, 50.32, 44.26 and 41.56%, respectively.

Groups Body weights (g) Body weight gains (g) At first cisplatin treatment [A] At 4th cisplatin treatment [B] Sacrifice [C] Total experiment periods [C-A] Test article dosing periods [C-B] Controls Intact 194.00 ± 8.96 345.00 ± 8.65 416.13 ± 16.99 222.13 + - 12.39 71.13 + - 11.03 Cisplatin 194.63 + - 7.71 324.50 ± 6.61 ^a 310.13 + - 8.18 ^a 115.50 ± 8.45 ^a - 14.38 + - 6.52 ^a Single formula Ondansetron 194.13 + - 9.17 325.75 ± 6.11 ^a 340.50 ± 9.35 ^ab 146.38337.639.41 ^ab 14.75 ± 9.48 ^ab BH 194.75 ± 3.99 326.88 ± 5.94 ^a 352.88 ± 7.61 ^abd 158.13 ± 7.92 ^abd 26.00 ± 5.68 ^abc Ondansetron 0.1 mg / ml and BH mixed formulation 40 mg / ml 194.25 + - 8.21 327.00 ± 12.46 ^a 367.88 ± 15.19 ^abc 173.63 ± 9.05 ^abc 40.88 + - 6.85 ^abc 20 mg / ml 192.25 + - 7.32 324.13 + - 7.30 ^a 358.88 ± 10.02 ^abc 166.63 ± 9.97 ^abc 34.75 ± 8.22 ^abc 10 mg / ml 192.00 8.02 324.63 + - 7.31 ^a 355.50 ± 10.43 ^abd 163.50 8.07 ^abc 30.88 ± 7.90 ^abc

^a p <0.01 compared with intact control by LSD test

^b p <0.01 compared with cisplatin control by LSD test

^c p <0.01 and ^d p <0.05 as compared with ondansetron single formula formula rats by LSD test

4.2.2. Changes in Carbon Shift Rate (Gastrointestinal Movement) in the Small Intestine

4.2.2.1. Change in the length of the small intestine

As a result of measuring the total length of the small intestine after fasting for 24 hours after the final administration, significant changes in the length of the small intestine compared to the normal medium control group were not observed in the experimental group causing gastrointestinal motility disturbance caused by all cisplatin. Ondansetron 0.1 mg / ml and BH 40 mg / ml alone, and all three doses of BH and ondansetron 0.1 mg / ml compound were not significantly different from the cisplatin control group. Ondansetron 0.1 mg / ml And BH 40, 20 and 10 mg / ml composite composition administration groups, no significant changes in the small intestine length were observed compared to the administration of ondansetron 0.1 mg / ml alone (Table 22).

Ondansetron 0.1 mg / ml and BH 40 mg / ml were administered alone, ondansetron 0.1 mg / ml and BH 40, 20 or 10 mg / ml, respectively, in the cisplatin control group compared to the normal medium control group. ml compound composition showed a change of -0.20, -0.39, 0.10, -0.29 and 0.10%, respectively, compared to the cisplatin control group.

Groups Gastrointestinal motilities (during 30min) Total small intestine length (cm) Length of charcoal meal transferred (cm) GI charcoal transit ratio (%) Controls Intact  127.50 + - 4.54  103.63 + - 10.25  81.34 ± 8.22 Cisplatin  127.25 + - 2.66 40.50 + - 13.76 ^a 31.83 ± 10.78 ^a Single formula treated Ondansetron  127.00 ± 2.83 65.38 ± 12.14 ^ac 51.41 + 9.07 ^ac BH  126.75 + - 5.04 64.75 ± 12.09 ^ac 51.14 ± 9.68 ^ac Ondansetron 0.1 mg / ml and BH mixed formulation 40 mg / ml  127.38 + - 4.84 92.13 ± 12.08 ^bcd 72.41 ± 9.85 ^cd 20 mg / ml  126.88 + - 5.03 87.88 ± 8.39 ^acd 69.36 ± 7.09 ^bcd 10 mg / ml  127.38 + - 2.50 81.13 ± 9.03 ^acd 63.70 + - 7.07 ^acd

4.2.2.2. Tombstone  Change of travel distance

(P <0.01) in the cisplatin control group compared to the normal control group. However, in the administration group of ondansetron 0.1 mg / ml and BH 40 mg / ml alone, in all BH and ondansetron combination administration groups, cisplatin (P <0.01) in the small intestine was significantly higher than that in the control group (p <0.01). In particular, (Table 22), respectively.

Ondansetron 0.1 mg / ml and BH 40 mg / ml alone administration group, ondansetron 0.1 mg / ml and BH 40, 20 or 10, respectively, showed a change of -60.92% compared to the normal medium control group in the cisplatin control group. mg / ml of the composite composition showed a change of 61.42, 59.88, 127.47, 116.98 and 100.31%, respectively, compared with the cisplatin control group.

4.2.2.3. The change of tongue in the small intestine

In the cisplatin control group, the ratio of the carbon shift distance to the total length of the small intestine, ie, the tongue in the cisplatin control group, was decreased, but the ondansetron 0.1 mg / ml single composition administration (P <0.01), respectively. In the case of ondansetron 0.1 mg / ml and BH 40, 20 or 10 mg / ml compound administration group, (p <0.01), respectively, as compared with the administration of ondansetron 0.1 mg / ml alone (Table 22).

Ondansetron 0.1 mg / ml and BH 40 mg / ml alone administration group, ondansetron 0.1 mg / ml and BH 40, 20 or 10, respectively, showed a change of -60.87% compared to the normal medium control group in the cisplatin control group. mg / ml compound composition showed a change of 61.54, 60.69, 127.57, 117.94 and 100.16%, respectively, compared with the cisplatin control group.

4.2.3. Changes in fecal indices

4.2.3.1. Variation of the variable

In the cisplatin-treated group, there was a significant decrease (p <0.01) in the number of the excreted fraction from 33 to 34 days and 24 hours after the initiation of cisplatin administration, (P <0.01), respectively. In the case of ondansetron 0.1 mg / ml and BH 40, 20 or 10 mg / ml compound administration group, ondansetron (P < 0.01 or p < 0.05), respectively, as compared with the 0.1 mg / ml single composition administration group (Table 23).

In the cisplatin-treated group, the total number of feces released during the period from 33 to 34 days and 24 hours after the initiation of cisplatin administration was -80.50% as compared with the normal medium control group. Ondansetron 0.1 mg / ml and BH 40 mg / The concentrations of ondansetron 0.1 mg / ml and BH 40, 20 or 10 mg / ml were 113.95, 105.81, 343.02, 296.51 and 247.67%, respectively, compared to the cisplatin control group.

Groups Fecal parameters Fecal pellet numbers Wet-weights (g) Dry-weights (g) Water contents (%) Controls Intact 55.13 + - 11.08 26.71 + - 3.74 4.33 ± 0.89 83.69 ± 3.13 Cisplatin 10.75 ± 3.11 ^d 5.98 ± 1.81 ^a 4.25 ± 0.65 25.25 ± 15.57 ^d Single formula treated Ondansetron 23.00 ± 6.35 ^df 12.38 ± 2.68 ^ab 4.33 ± 0.79 63.69 + - 8.65 ^df BH 22.13 + - 6.83 ^df 12.10 ± 2.63 ^ab 4.50 ± 1.16 61.70 ± 10.61 ^df Ondansetron 0.1 mg / ml and BH mixed formulation 40 mg / ml 47.63 ± 14.48 ^efg 26.27 ± 3.38 ^bc 4.40 ± 0.92 82.67 ± 5.43 ^fg 20 mg / ml 42.63 ± 11.10 ^eFg 21.1083.441.41 ^abc 4.42 + 0.72 78.85 ± 4.31 ^efg 10 mg / ml 37.38 ± 10.38 ^eh 19.72 ± 2.46 ^abc 4.38 ± 0.77 77.38 ± 5.36 ^efg

4.2.3.2. Changes in wet weight

Compared with the cisplatin control group, all of the experimental groups, including BH 40 mg / ml and ondansetron 0.1 mg / ml, showed a significant decrease (p <0.01) in the cisplatin control group compared with the normal medium control group (Ondansetron 0.1 mg / ml) and BH 40, 20, and 10 mg / ml were significantly (p <0.01) higher than those of the ondansetron 0.1 mg / <0.01). The increase of the wet weight was recognized as dependent on the BH concentration in the complex (Table 23).

In the cisplatin control group, the mean wet weight of the feces discharged for 33 to 34 days and 24 hours after the start of cisplatin administration was -77.59% as compared with the normal medium control group. Ondansetron 0.1 mg / ml and BH 40 mg / The concentrations of ondansetron 0.1 mg / ml and BH 40, 20 or 10 mg / ml were 106.83, 102.15, 338.96, 252.64 and 229.48%, respectively, in the combination administration group compared to the cisplatin control group.

4.2.3.3. Changes in fecal dry weight

The mean dry weights of fecal excretions for 33 to 34 days and 24 hours after initiation of cisplatin administration were found to be significantly higher than those of the normal medium control group in all cisplatin-induced gastrointestinal motility disorders The change in mean fecal dry weight was significantly greater in the mg / ml group compared to the cisplatin control group in the administration of the ondansetron 0.1 mg / ml and BH 40 mg / ml single composition, all three concentrations of BH and ondansetron 0.1 mg / And no significant change in mean fecal dry weight was observed in the group administered with 40, 20 or 10 mg / ml of BH and 0.1 mg / ml of ondansetron, respectively, as compared with the administration of ondansetron 0.1 mg / ml alone ).

In the cisplatin control group, the average dry weight of the feces released for 33-34 days and 24 hours after the initiation of cisplatin administration was -1.68% as compared with the normal medium control group. Ondansetron 0.1 mg / ml and BH 40 mg / In the group administered with ondansetron 0.1 mg / ml and BH 40, 20 or 10 mg / ml, the change was 1.79, 5.76, 3.53, 4.03 and 2.97%, respectively, compared with the cisplatin control group.

4.2.3.4. Feces  Change in moisture content

The mean fecal water content of the cisplatin control group was significantly lower than that of the normal control group (p <0.01), but the BH 20 mg / ml Ondansetron 0.1 mg / ml was significantly increased (p <0.01) compared to the cisplatin control group, and ondansetron 0.1 mg / ml and BH 40, 20 and 10 mg / ml composite composition, the increase in the average fecal water content was significantly (p <0.01) higher than that of the administration of ondansetron 0.1 mg / ml alone (Table 23)

In the cisplatin control group, the mean moisture content of the feces released during the period from 33 to 34 days and 24 hours after the initiation of cisplatin administration was -69.83% as compared with the normal medium control group. Ondansetron 0.1 mg / ml and BH 40 mg / The concentrations of ondansetron 0.1 mg / ml and BH 40, 20 or 10 mg / ml were 152.23, 144.33, 227.40, 212.26 and 206.44%, respectively, compared to the cisplatin control group.

4.2.4. Changes in antioxidant defense systems in the stomach

4.2.4.1. Above the gates MDA  Change in content

(P <0.01) in the cisplatin control group compared to the normal medium control group, but the MDA content was increased in all the tissues including the BH 10 mg / ml and ondansetron 0.1 mg / ml combination composition administration groups (P <0.01), respectively. In all of the ondansetron and BH complex administration groups, the MDA content was significantly (p <0.01) lower than that of the cisplatin control group, The reduction of lipid peroxidation in the pharyngeal tissues was recognized dependent on the complexed BH concentration (Table 24).

In the cisplatin control group, MDA content in the pharyngeal tissue of the final sacrifice day was 463.75% compared to the normal medium control group. Ondansetron 0.1 mg / ml and BH 40 mg / ml alone administration group, ondansetron 0.1 mg / ml and BH 40 , 20 or 10 mg / ml of the composite composition showed -29.54, -48.15, -71.95, -62.49 and -55.89%, respectively, as compared with the cisplatin control group.

Groups Fundus antioxidant defense systems Malondialdehyde
(nM / g tissue) Glutathione
(μM / mg tissue) Catalase
(nM / mim / mg tissue) Superoxide dismutase
(nM / mim / mg tissue) Controls Intact 3.75 ± 1.41 4.72 0.95 131.50 ± 18.19 216.00 ± 20.07 Cisplatin 21.11 ± 4.51 ^e 1.03 + 0.29 ^e 34.53 + - 11.80 ^a 61.88 ± 16.44 ^a Single formula treated Ondansetron 14.88 ± 2.03 ^ef 1.42 ± 0.19 ^eG 52.13 + - 10.78 ^ac 89.63 ± 11.11 ^ab BH 10.95 ± 0.87 ^EFh 1.92 ± 0.21 ^efh 64.63 ± 11.38 ^ab 103.88 ± 15.08 ^ab Ondansetron 0.1 mg / ml and BH mixed formulation 40 mg / ml 5.92 ± 1.03 ^eh 3.66 ± 0.84 ^fh 109.75 ± 12.49 ^abd 183.25 ± 30.07 ^abd 20 mg / ml 7.92 ± 2.10 ^eh 3.11 ± 0.91 ^eh 91.88 ± 12.23 ^abd 159.75 ± 21.36 ^usd 10 mg / ml 9.31 ± 2.59 ^eh 2.44 ± 0.68 ^efh 81.38 ± 18.39 ^abd 137.13 ± 19.23 ^abd

^a p <0.01 compared with intact control by LSD test

^b p < 0.01 and ^c p < 0.05 ascomparedwithcisplatincontrolbyLSDtest

^d p <0.01 as compared with ondansetron single formula rats by LSD test

^e p <0.01 as compared with intact control by MW test

^f p < 0.01 and ^g p < 0.05 ascomparedwithcisplatincontrolbyMWtest

^h p <0.01 compared with ondansetron single formula rats by MW test

4.2.4.2. Above the gates GSH  Change in content

(P <0.01), whereas GSH content of endogenous antioxidant was decreased in the cisplatin control group compared to the normal medium control group. However, all of the candidates administered with BH 10 mg / ml and ondansetron 0.1 mg / (P <0.01 or p <0.05), respectively. In the case of ondansetron 0.1 mg / ml and BH 40, 20 or 10 mg / ml, the concentration of ondansetron The increase in GSH content in the gingival tissues was significantly (p <0.01) dependent on the concentration of BH concentration (Table 24).

In the cisplatin control group, the GSH content in the pharyngeal tissue of the final sacrifice day was -78.23% compared to the normal medium control group. Ondansetron 0.1 mg / ml and BH 40 mg / ml alone administration group, ondansetron 0.1 mg / 40, 20, or 10 mg / ml, respectively, compared with the cisplatin control group, the change was 38.37, 86.60, 256.27, 203.17, and 137.76%, respectively.

4.2.4.3. Changes in CAT activity in the supraspinatus

In the cisplatin control group, significant decrease in CAT activity, which is an endogenous antioxidant enzyme, was observed in the cisplatin control group (p <0.01). However, in all candidates treated with ondansetron 0.1 mg / ml alone group compared to the cisplatin control group (Ondansetron 0.1 mg / ml) and BH 40, 20, or 10 mg / ml, respectively, were significantly increased (p <0.01 or p <0.05) The increase in CAT activity in the gingiva tissues was significantly (p <0.01) higher than that of the composition-administered group (Table 24).

In the cisplatin control group, the CAT activity in the pharyngeal tissue of the final sacrifice day was -73.74% compared to the normal medium control group. Ondansetron 0.1 mg / ml and BH 40 mg / ml alone administration group, ondansetron 0.1 mg / ml and BH 40, 20, or 10 mg / ml, respectively, compared with the cisplatin control group, the change was 50.94, 87.14, 217.81, 166.04, and 135.64%, respectively.

In the cisplatin control group, the CAT activity in the pharyngeal tissue of the final sacrifice day was -73.74% compared to the normal medium control group. Ondansetron 0.1 mg / ml and BH 40 mg / ml alone administration group, ondansetron 0.1 mg / ml and BH 40, 20, or 10 mg / ml, respectively, compared with the cisplatin control group, the change was 50.94, 87.14, 217.81, 166.04, and 135.64%, respectively.

4.2.4.4. Changes in SOD activity on the pulmonary trunk

In the cisplatin control group, SOD activity, which is endogenous antioxidant enzyme, was significantly decreased in the cisplatin control group (p <0.01) The increase of SOD activity in the tissues of the group treated with ondansetron 0.1 mg / ml and BH 40, 20 or 10 mg / ml was significantly (p <0.01) higher than that of ondansetron alone BH concentration-dependently (Table 24).

In the cisplatin control group, SOD activity in the pharyngeal tissue of the final sacrifice day was -71.35% compared to the normal medium control group. Ondansetron 0.1 mg / ml and BH 40 mg / ml alone administration group, ondansetron 0.1 mg / 40, 20, or 10 mg / ml, respectively, compared with the cisplatin control group, 44.85, 67.88, 196.16, 158.18, and 121.62%, respectively.

4.2.5. Changes of Tryptophan hydroxylase and monoamine oxidase activity

4.2.5.1. Changes in tryptophan hydroxylase activity in pylorus

In the cisplatin control group, the tryptophan hydroxylase activity, which is a serotonin synthetase, was increased in the pyloric tissue of the control group compared to the normal control group. However, all the candidates treated with the ondansetron 0.1 mg / (Ondansetron 0.1 mg / ml) and BH 40, 20 or 10 mg / ml, respectively, in the pyloric stomach tissues, respectively. The decrease in tryptophan hydroxylase activity in the pyloric tissues was significantly (p <0.01) dependent on BH concentration. On the other hand, a change in tryptophan hydroxylase activity similar to that of the cisplatin control was observed in the administration of ondansetron 0.1 mg / ml alone (FIG. 19).

In the cisplatin control group, the tryptophan hydroxylase activity in the pyloric tissue of the final sacrifice day was 163.79% compared to the normal medium control group. Ondansetron 0.1 mg / ml and BH 40 mg / ml alone administration group, ondansetron 0.1 mg / ml and BH 40, 20, or 10 mg / ml of the composite composition showed a change of -0.27, -35.91, -55.57, -46.35, and -37.30%, respectively, compared with the cisplatin control group.

4.2.5.2. Changes in monoamine oxidase activity in the pylorus

On the other hand, the mononamine oxidase activity in the cisplatin control group was significantly decreased (p <0.01) compared to the normal control group. Ondansetron 0.1 mg / ml (P <0.01) monoamine oxidase activity was significantly higher in all candidate compound administration groups than in the cisplatin control group, especially in all composite composition administration groups, compared to the administration of ondansetron 0.1 mg / ml alone p <0.01). The increase of the monoamine oxidase activity in the pyloric stomach tissues was recognized dependent on the complexed BH concentration (Fig. 20).

In the cisplatin control group, the monoamine oxidase activity in the pyloric tissue of the final sacrifice day was -67.60% compared to the normal medium control group. Ondansetron 0.1 mg / ml and BH 40 mg / ml alone administration group, ondansetron 0.1 mg / In the BH 40, 20 or 10 mg / ml compound administration group, the changes were 2.87, 65.07, 136.84, 106.70 and 67.94%, respectively, compared with the cisplatin control group.

4.2.6. Changes in gastrin and serotonin contents in pyloric tissue

4.2.6.1. Changes in gastrin content in the pylorus

There was a significant decrease in gastrin content in the pyloric stomach tissues of the cisplatin control group compared with the normal control group (p <0.01). However, in all candidates treated with ondansetron 0.1 mg / ml alone group, cisplatin control (Ondansetron 0.1 mg / ml) and BH 40, 20, or 10 mg / ml compound administration group showed significant increase of gastrin content in the pyloric tissue (p <0.01) (P <0.01). The increase of gastrin content in the pyloric tissues was dependent on the concentration of BH. On the other hand, in the administration group of ondansetron 0.1 mg / ml alone, a change in the gastrin content in the pyloric tissue similar to the cisplatin control was observed (FIG. 21).

In the cisplatin control group, gastrin content in the pyloric tissue of the final sacrifice day was -80.10% compared to the normal medium control group. Ondansetron 0.1 mg / ml and BH 40 mg / ml alone administration group, ondansetron 0.1 mg / ml and BH 40, 20, or 10 mg / ml, respectively, compared with the cisplatin control group, the changes were 5.56, 93.99, 293.77, 211.68, and 163.07%, respectively.

4.2.6.2. Changes in serotonin content in the pylorus

Serotonin levels were increased in the pyloric tissues of the cisplatin control group compared to the normal control group (p <0.01). However, all candidates except for the ondansetron 0.1 mg / ml single composition administration group Serum concentrations of ondansetron (0.1 mg / ml) and BH 40, 20 or 10 mg / ml (0.1 mg / ml) ondansetron were significantly lower than those of the cisplatin control group (p <0.01) than that of the single composition administration group, the decrease in the serotonin content in the pyloric tissue was found to be dependent on the complexed BH concentration. On the other hand, in the administration group of ondansetron 0.1 mg / ml alone, a change in the content of serotonin in the pyloric tissue similar to that of the cisplatin control was observed (FIG. 22).

In the cisplatin control group, the serotonin content in the pyloric tissue of the final sacrifice day was 219.73% compared with the normal medium control group. Ondansetron 0.1 mg / ml and BH 40 mg / ml alone group administration group, ondansetron 0.1 mg / ml and BH 40 , 20, or 10 mg / ml, respectively, were -3.05, -25.26, -57.08, -44.25 and -32.12%, respectively, as compared with the cisplatin control group.

4.2.7. Histopathological change

4.2.7.1. Histopathological changes on the gyrus

In the cisplatin control group, focal erosive damages of the surface mucosa with the atrophy of the entire gastric mucosa resulted in a decrease in the thickness of the gastric mucosa (p <0.01) compared to the normal medium control group, (P <0.01) increased significantly (p <0.05), but there was no statistically significant difference (p <0.01) between total surface pyloric mucosa length and surface erosive pyloric mucosa length. On the other hand, in all candidates treated with BH 40 mg / ml and ondansetron 0.1 mg / ml, the atrophy and local erosion of cisplatin-induced mucosa were significantly decreased, In addition, the ondansetron 0.1 mg / ml and BH 40, 20 or 10 mg / ml combination compositions showed a significant increase in gastric mucosal thickness and decreased mucosal injury rate. (p < 0.01 or p &lt; 0.05), respectively. The increase in the thickness of the gastric mucosa and the reduction of the gastric mucosal injury rate were recognized as dependent on the complexed BH concentration (Table 25, Fig. 23).

Ondansetron 0.1 mg / ml and BH 40 mg / ml were administered alone, ondansetron 0.1 mg / ml and BH 40, 20 or 10, respectively, in the cisplatin control group. mg / ml of the composite composition showed 32.63, 43.85, 74.27, 64.97 and 53.26%, respectively, as compared with the cisplatin control group.

Ondansetron 0.1 mg / ml and BH 40 mg / ml were administered alone, ondansetron 0.1 mg / ml and BH 40, 20 or 10, respectively, in the cisplatin control group compared with the normal medium control group. mg / ml composite composition, cisplatin

-34.97, -32.34, -71.53, -60.51 and -56.86%, respectively, as compared with the control group.

Groups Fundus Pylorus Total mucosa thickness (μm) Degenerative mucosa percentage (% / mucosa) Total mucosa thickness (μm) Degenerative mucosa percentage (% / mucosa) Controls Intact 982.32 + - 111.14 4.18 ± 2.08 330.20 ± 96.96 4.82 ± 2.87 Cisplatin 455.47 +/- 109.12 ^a 58.71 ± 11.89 ^a 134.58 ± 36.93 ^e 75.00 ± 10.77 ^a Single formula treated Ondansetron 604.08 + - 59.49 ^ab 38.18 ± 12.26 ^ab 188.81 ± 19.02 ^eg 50.77 ± 11.80 ^ab BH 655.18 ± 53.79 ^ab 39.72 ± 7.61 ^ab 197.38 ± 21.93 ^eg 57.19 ± 13.50 ^ab Ondansetron 0.1 mg / ml and BH mixed formulation 40 mg / ml 793.75 ± 99.19 ^abc 16.71 ± 3.94 ^abc 269.86 + - 31.04 ^gh 26.17 ± 10.72 ^abc 20 mg / ml 751.38 ± 60.32 ^abc 23.19 ± 5.96 ^abc 237.05 ± 37.01 ^fgh 31.69 ± 11.63 ^abc 10 mg / ml 698.08 ± 66.07 ^abd 25.33 + 4.85 ^abc 215.30 ± 14.37 ^egi 35.29 ± 10.76 ^abc

4.2.7.2. Histopathological changes in the pylorus

In the cisplatin control group, there was a significant decrease in the pyloric gastric mucosal thickness (p <0.01) compared to the normal medium control group due to the atrophy of the entire pyloric gastric mucosa and the local erosion of the surface mucosa, (P < 0.01) < / RTI &gt; On the other hand, in all experimental groups including 20 mg / ml of BH and 0.1 mg / ml of ondansetron, the cystatin-induced pyloric stomach mucosa atrophy and local erosion were markedly decreased, and the pylorus was significantly (p <0.01) In addition, ondansetron 0.1 mg / ml and BH 40, 20 or 10 mg / ml were more effective than ondansetron 0.1 mg / ml alone (p &lt; 0.01 or p &lt; 0.05), respectively. The increase in the thickness of the pyloric stomach gum and the decrease in the pyloric stomach mucosal injury rate were respectively recognized as dependent on the complexed BH concentration (Table 25, Fig.

Ondansetron 0.1 mg / ml and BH 40 mg / ml alone group, ondansetron 0.1 mg / ml and BH 40, 20 or 10, respectively, showed a change of -59.24% compared to the normal medium control group in the cisplatin control group. mg / ml composite composition showed a change of 40.29, 46.67, 100.53, 76.14 and 59.98%, respectively, compared with the cisplatin control group.

In the isplatin control group, the rate of damage to the gastric mucosa was 1456.50% compared to that of the normal control group. Ondansetron 0.1 mg / ml and BH 40 mg / ml alone administration group, ondansetron 0.1 mg / ml and BH 40, 20 or 10 mg / ml compound composition showed a change of -32.31, -23.75, -65.11, -57.75 and -52.95%, respectively, compared to the cisplatin control group.

4.2.7.3. Histopathological changes of the colon, including residual feces

In the cisplatin control group, there was significant decrease (p <0.01) in the number of mucin-producing cells per unit area (mm ² ) and residual fecal surface mucus thickness as well as atrophy of the whole crystalline mucosa including residual feces. There was a decrease in the thickness of the colon mucosa, the number of mucus producing cells in the mucosa, and the thickness of the residual fecal surface mucus, including the residual feces. On the other hand, in all experimental groups including BH 10 mg / ml and ondansetron 0.1 mg / ml compound composition, the decrease of cisplatin-induced colonic mucosal atrophy, number of mucus-producing cells in the mucosa and number of residual fecal surface mucus thickness was significantly suppressed, (P <0.01), respectively. The concentrations of ondansetron (0.1 mg / ml) and BH 40, 20 or 10 mg / ml were higher than those of the control group (P <0.01 or p <0.05), respectively, in comparison with the administration of ondansetron 0.1 mg / ml alone (P <0.01 or p <0.05), and the increase in the number of mucus-producing cells and the residual fecal surface mucus thickness 26, Fig. 25).

In the cisplatin control group, the surface mucus thickness of the remaining feces in the colon was -95.48% compared to the normal medium control group. Ondansetron 0.1 mg / ml and BH 40 mg / ml alone administration group, ondansetron 0.1 mg / ml and BH 40, 20, or 10 mg / ml, respectively, compared to the cisplatin control group, 409.66, 442.34, 655.97, 595.50, and 525.44%, respectively.

In the cisplatin control group, the number of mucin-producing cells per unit area of the colon mucosa was -84.90% as compared with that of the normal medium control group. Ondansetron 0.1 mg / ml and BH 40 mg / ml alone administration group, ondansetron 0.1 mg / ml and BH 40 , And 20 or 10 mg / ml, respectively, compared with the cisplatin control group, 68.91, 206.30, 540.90, 500.14 and 335.85%, respectively.

In the cisplatin control group, the mean mucosal thickness of the colon, including residual feces, was -57.85% compared to the normal medium control group. Ondansetron 0.1 mg / ml and BH 40 mg / ml alone administration group, ondansetron 0.1 mg / ml and BH 40, 20 or 10 mg / ml, respectively, compared to the cisplatin control group, which was 40.52, 58.44, 155.41, 133.98 and 92.20%, respectively.

Groups Histomorphometry (at sacrifice) Fecal pellet surface mucous thicknesses (μm) Mucous producing cell numbers (cells / mm ² ) Colon mucosa thicknesses (μm) Controls Intact  94.82 ± 25.58  591.252120.75  228.04 + - 40.17 Cisplatin 4.28 ± 1.52 ^a 89.25 + - 22.65 ^a 96.11 ± 16.06 ^a Single formula treated Ondansetron 21.82 ± 5.26 ^ab 150.75 ± 15.01 ^ab 135.05 ± 13.95 ^ab BH 23.22 ± 3.40 ^ab 273.38 ± 23.73 ^abc 152.27 ± 30.43 ^ab Ondansetron 0.1 mg / ml and BH mixed formulation 40 mg / ml 32.37 ± 5.39 ^abc 572.00 ± 86.06 ^bc 245.47 ± 26.21 ^bc 20 mg / ml 29.78 ± 5.41 ^abd 353.63 + - 116.18 ^bc 224.87 ± 32.38 ^bc 10 mg / ml 26.78 ± 3.15 ^usd 389.00 ± 88.60 ^abc 184.71 + - 34.90 ^bc

4.2.8. Immunohistochemical change

4.2.8.1. Changes of Gastrin Immunoreactive Cells in the Pyloric Gastric Mucosa

In the cisplatin control group, the number of gastrin-immunoreactive cells per unit area (mm ² ) of the pyloric gut mucosa was significantly (p <0.01) lower than that of the normal medium control group, but in all experimental groups except ondansetron 0.1 mg / Ondansetron 0.1 mg / ml and BH 40, 20, or 10 mg / ml, respectively, showed a significant increase in gastrin immunoreactive cells in the pyloric gastric mucosa compared to the cisplatin control group (p <0.01) (p <0.01), the number of gastrin-immunoreactive cells in the pyloric glands was significantly higher than that of the control group. On the other hand, the number of gastrin immunoreactive cells in the pyloric gastric mucosa similar to that of the cisplatin control group was recognized in the administration group of ondansetron 0.1 mg / ml alone (Table 27, Fig. 26).

In the cisplatin control group, the number of gastrin-immunoreactive cells per unit area of the pyloric stomach was -91.14% compared to the normal control group. Ondansetron 0.1 mg / ml and BH 40 mg / ml alone administration group, ondansetron 0.1 mg / 40, 20, or 10 mg / ml, respectively, compared with the cisplatin control group, the change was 2.86, 267.14, 681.43, 504.29, and 442.86%, respectively.

Groups Numbers of IR cells (cells / mm ^{2 of} mucosa) Gastrin-IR cells in the pylorus mucosa Serotonin-IR cells in the pylorus mucosa Serotonin-IR cells in the colon mucosa Controls Intact  98.75 ± 20.15  8.50 ± 2.45  10.25 + - 3.28 Cisplatin 8.75 ± 2.12 ^a 58.25 + 14.66 ^a 71.13 ± 12.10 ^a Single formula treated Ondansetron 9.00 ± 3.34 ^a 55.88 +/- 13.55 ^a 69.00 ± 14.56 ^a BH 32.13 ± 11.54 ^aces 37.75 ± 10.62 ^adf 50.25 + 10.40 ^acf Ondansetron 0.1 mg / ml and BH mixed formulation 40 mg / ml 68.38 ± 20.48 ^bce 12.63 + - 2.50 ^bce 13.50 ± 3.42 ^ce 20 mg / ml 52.88 ± 12.63 ^ace 27.13 ± 3.27 ^ace 21.88 ± 4.55 ^aces 10 mg / ml 47.50 ± 12.48 ^aces 35.50 +/- 10.93 ^acce 33.50 ± 6.89 ^aces

4.2.8.2. Pylorus  Mucosa serotonin  Changes in immune response cells

In the cisplatin control group, the number of serotonin immunoreactive cells per unit area (mm ² ) of the pyloric stomach mucosa was significantly increased (p <0.01) compared to the normal medium control group. However, all candidates except ondansetron 0.1 mg / The number of serotonin immunoreactive cells in the pyloric stomach mucosa was significantly decreased (p <0.01 or p <0.05) compared with the cisplatin control group. Especially, ondansetron 0.1 mg / ml and BH 40, 20 or 10 mg / (P <0.01), compared with the administration of ondansetron 0.1 mg / ml alone, the number of serotonin immunoreactive cells in the pyloric stomach mucosa was found to be dependent on the complex BH concentration. On the other hand, the number of serotonin immunoreactive cells in the pyloric gastric mucosa similar to that of the cisplatin control group was recognized in the administration group of ondansetron 0.1 mg / ml alone (Table 27, Fig. 26).

In the cisplatin control group, the number of serotonin-immunoreactive cells per unit area of the pyloric stomach was 585.29% higher than that of the normal control group. Ondansetron 0.1 mg / ml and BH 40 mg / , 20 or 10 mg / ml, respectively, compared to the cisplatin control group, the changes were -4.08, -35.19, -78.33, -53.43 and -39.06%, respectively.

4.2.8.3. Changes in Immunoreactive Cells of Colon Mucosa Serotonin

In the cisplatin-treated group, serotonin-immunoreactive cells per unit area (mm ² ) of the colon mucosa were significantly increased (p <0.01) compared to the normal medium control group, but in all experimental groups except ondansetron 0.1 mg / Serum concentrations of ondansetron (0.1 mg / ml) and ondansetron (0.1 mg / ml) were significantly lower in the group receiving the ondansetron 0.1 mg / ml and BH 40, 20 or 10 mg / ml combination compositions than in the cisplatin control group The number of serotonin immunoreactive cells in the colon mucosa was significantly (p <0.01) dependent on BH concentration. On the other hand, the number of serotonin immunoreactive cells in the colon mucosa similar to that of the cisplatin control group was recognized in the group administered with ondansetron 0.1 mg / ml alone (Table 27, Fig. 26).

In the cisplatin control group, the number of serotonin immunoreactive cells per unit area of the colon mucosa was 593.90% higher than that of the normal control group. Ondansetron 0.1 mg / ml and BH 40 mg / ml alone administration group, ondansetron 0.1 mg / ml and BH 40, 20, and 10 mg / ml, respectively, compared with the cisplatin control group, the changes were -2.99, -29.35, -81.02, -69.24, and -52.90%, respectively.

In Example 4, the study on the reduction of adverse effects of chemotherapy for general cancer patients was conducted. CIMI-15-02-02 O + BH], the effect of BH complex composition on the pharmacological effect of ondansetron was evaluated using cisplatin-induced gastrointestinal motility disorder rats.

In this study, ondansetron doses were selected as 1 mg / kg (0.1 mg / ml) [Malik et al., 2007] which is known to have a definite effect on previous cisplatin-induced digestive tract disorders. / kg (concentration of 40 mg / ml), concentrations of BH of 40, 20, and 10 mg / ml were investigated for the pharmacokinetic studies of the ondansetron and BH complex compositions of the present inventors [CIMI-14-02-11- 02 O-BH PK, 2015; Cisplatin 2 mg / kg was given once a week for 5 weeks, and ondansetron 0.1 mg / kg was administered from 4 doses of cisplatin. ml and BH 40 mg / ml single composition, ondansetron 0.1 mg / ml, and BH 40, 20 or 10 mg / ml were orally administered for 14 days to assess the carbohydrate mobility in the small intestine and evaluate the effect on gastrointestinal motility , Changes in body weight, fecal index (fraction, fecal weight, and fecal water content), colon histopathological changes including residual fecal pellet, fecal surface mucosal thickness, mucin production per mm2, and mean mucosal thickness In order to evaluate the effects on gastrointestinal mucosal injury and the physiology of endocrine cells in the digestive tract, changes in the thickness of stomach, pyloric and colon mucosa and the area of damaged area, gastrin and serotonin immunity in pyloric gut half Changes in gastrin and serotonin contents in the pyloric stroma, changes in serotonin metabolism related enzymes - tryptophan hydroxylase and monoamine oxidase activity were observed, respectively. In order to observe the antioxidant effect, GSH, SOD, and CAT content on the gates were measured and MDA content was measured to observe the effect on lipid peroxidation. Experimental results were compared with the effect of ondansetron on the complex composition of BH, compared to the administration of ondansetron 0.1 mg / ml alone.

The results of this study showed that cisplatin significantly reduced weight loss, decreased intestinal flotation, decreased excretion and fecal water content, increased tryptophan hydroxylase activity and decreased monoamine oxidase activity, pyloric gastritis and gastrin immunoreactivity Decreased number of cells, increased pyloric stomach serotonin content, increased number of serotonin immunoreactive cells in pyloric stomach and colon, atrophy and local surface damage of pyloric stomach, stomach or colon mucosa, mucus production on surface mucus and colon mucosa of remaining feces in colon Numerical reduction of cells was observed, and previous reports [Bradner and Schurig, 1981; Kilpatrick et al., 1990; Tyers, 1991; Song et al., 2012], resulting in gastrointestinal injury, particularly gastrointestinal motility disturbances due to endocrine cell disturbances in the gastrointestinal tract, resulting in a significant secondary constipation. In addition, the inhibition of the antioxidant defense system has also been reported in previous reports [Chang et al., 2002], as increased MDA content in pyloric glands, decreased GSH content, and decreased SOD and CAT activity. Chirino and Pedraza-Chaverri, 2009], and was found to be involved in cisplatin-induced digestive tract injury. On the other hand, administration of the combination of BH 40 mg / ml single composition, ondansetron 0.1 mg / ml and BH 40, 20 or 10 mg / ml resulted in inhibition of gastrointestinal motility disorder induced by cisplatin, changes in gastrointestinal endocrine cells, Inhibition of cisplatin-induced gastrointestinal uptake was significantly suppressed in all doses of BH 40, 20, or 10 mg / ml and ondansetron 0.1 mg / ml, Increased inhibitory effects on motor impairment, endocrine cell changes in the digestive tract, and inhibition of the pyloric antioxidant defense system were found to be dependent on the complex BH concentration. Therefore, previous pharmacokinetic experiments by our researchers [CIMI-14-02-11-02 O-BH PK, 2015; As a result of CIMI-14-02-11-04 O-BH PK (2), 2015, it was observed that the complex composition of BH did not affect the bioavailability of ondansetron. Ondansetron and BH composition administration was ondansetron , The combination of ondansetron and BH is considered to increase the antioxidant and gastrin and serotonin production of gastrointestinal endocrine cells by modulating the secretion or function of the gastrointestinal endocrine cells without affecting the bioavailability of ondansetron, And it is expected that it will provide a new effective treatment method which is very useful for the treatment of gastrointestinal disorders caused by chemotherapeutic agents. Ondansetron, on the other hand, is a typical 5-HT3 receptor blocker [Beck et al., 1993; Miyata et al., 1995; Martin et al., 1998; Ozmen and Ku2004; Malik et al., 2007] did not have a significant effect on serotonin production, secretion and degradation in tissues [Song et al., 2012].

Cisplatin acts as a typical redox cycler [Chirino and Pedraza-Chaverri, 2009], resulting in direct damage to various organs of the body and consequently a significant reduction in body weight [Suddek et al., 2011; Khan et al., 2012; Naqshbandi et al., 2012]. In this study, significant weight loss was observed following cisplatin administration. Three weeks after the initiation of cisplatin administration, animals with similar body weights were selected. On the other hand, significant increases in body weight were significantly increased in all of the ondansetron 0.1 mg / ml and BH 40 mg / ml single composition, ondansetron 0.1 mg / ml and BH 40, 20 or 10 mg / 7, and the body weight gain during the experimental period and during the drug administration period was also significantly higher than that of the cisplatin control group. Especially, all concentrations of BH 40, 20 or 10 mg / ml and ondansetron 0.1 mg / , The increase in body weight and body weight gain was more dependent on the concentration of BH than ondansetron 0.1 mg / ml alone. These results indicate that ondansetron and BH are effective as antioxidants or as a result of cisplatin-induced long-term damage in the whole body, and that the ondansetron cisplatin induction by the BH complex composition of 10 mg / It is considered to be one of the evidences that the effect of inhibiting gastrointestinal disorders is remarkably increased synergistically.

Gastrointestinal motility disorder is known to decrease the water content in the feces due to decreased fecal discharge such as constipation and excessive water absorption due to the associated fecal congestion. Therefore, fecal index such as fecal pellet count and fecal water content Have been used to assess the effects of various drugs on gastrointestinal motility disorders [Wintola et al., 2010]. [Wintola et al., 2010], it is well known that the intestinal flotation rate is very useful for diagnosing abnormalities of gastrointestinal motility [Wintola et al., 2010] Decrease indicates stagnation of fecal matter, ie, decreased state of digestive tract motility such as constipation [Sagar et al., 2005; Meite et al., 2010]. Therefore, the increase of the fecal indices and the intestinal flotation rates observed by the administration of ondansetron and BH alone or in combination is a direct evidence that these ondansetron and BH inhibit cisplatin-induced gastrointestinal motility delays very effectively. As a result of this study, it was found that the increase of the stimulating effect of digestive tract peristalsis was more significant than that of the administration of ondansetron 0.1 mg / ml alone in all BH 40, 20 or 10 mg / ml and ondansetron 0.1 mg / Respectively, and it was concluded that the onset of ondansetron was synergistically increased by the BH complex composition of 10 mg / ml or more.

Various toxic substances formed by lipid peroxidation, especially active oxygen species, are known to cause destruction of surrounding tissues [Comporti, 1985]. Cisplatin also acts as a typical strong redox cycler, producing harmful active oxygen species, (Chang et al., 2002; Chirino and Pedraza-Chaverri, 2009]. GSH is a representative endogenous antioxidant, and it is known to inhibit various tissue damages by decreasing active oxygen species formed in the cells, thus acting as a typical antioxidant factor in tissues [Odabasoglu et al., 2006]. In addition, SOD is a typical antioxidant enzyme in cells, and it plays a role in removing various oxidizing substances from cells. CAT is also an enzyme that converts H2O2, which is a strong active oxidizing substance, to H2O [Cheeseman and Slater, 1993]. Therefore, lipid peroxidation, reduction of GSH content, and inhibition of SOD and catalase activity play a crucial role in inhibiting gastric mucosal injury [Su et al., 2009]. As a result of this experiment, it was observed that administration of all BH alone or combination compositions effectively blocked the inhibition of lipid peroxidation and antioxidant defense system by cisplatin, and that the ondansetron 0.1 mg / ml single composition was also obtained from previous studies [Miyata et al. 1995; Ozmen and Ku2004; In ondansetron 0.1 mg / ml and all concentrations of BH 40, 20 or 10 mg / ml combination composition, ondansetron 0.1 mg / ml was found to be effective in the treatment of cisplatin- mg / ml was significantly higher than that of the single composition administration group, and the increase in the activity of the antioxidant defense system was recognized to be dependent on the concentration of BH in the cisplatin-induced digestive tract rats. These results suggest that the BH complex composition of at least 10 mg / ml is a direct evidence of a synergistic increase in the antioxidant effect of ondansetron in cisplatin-induced gastrointestinal tract rats. In addition, active oxygen species induced by cisplatin cause significant digestive disturbances due to significant atrophy of the gastrointestinal mucosa and localized mucosal surface damage by inflammation [Bearcroft et al., 1999; Ju et al., 2008], various 5-HT3 receptors blockers [Yoshida et al., 1992; Kishibayashi et al., 1993], anti-inflammatory agents [Roos et al., 1981] and antioxidants [Sharma and Gupta, 1998; Badary et al., 2006] have shown a beneficial effect on these cisplatin-induced gastrointestinal disorders. The results of this experiment showed that cisplatin treatment resulted in significant erosion of the follicles, pyloric stomach, and colon mucosa, as well as local surface erosive damage. However, administration of ondansetron and BH alone or in combination to the cisplatin- And ondansetron 0.1 mg / ml were significantly lower than those of the administration of ondansetron 0.1 mg / ml alone in all BH 40, 20 or 10 mg / ml and ondansetron 0.1 mg / Increased inhibitory effects on atrophy and local surface damage were found to be dependent on the complexed BH concentration, respectively.

Histopathologically, decreased secretion of intracolonic mucus is directly associated with constipation-like digestive motility disorders [Yang et al., 2008], marked constriction of mucous membranes in colon and decrease of mucus-producing cells are recognized [McCullogh et al., 1998], and a marked increase in the residual fecal mass in the colon during constipation and a decrease in mucus on the feces surface [Yang et al., 2008; Wu et al., 2011; Choi et al., 2014], the increase in the number of mucin-producing cells and the increase in the thickness of the residual mucinous surface mucous layer in the colon by the administration of ondansetron and BH alone or in combination with the combination of ondansetron and BH As a result of this. As a result of this experiment, it was found that in all of the BH 40, 20 or 10 mg / ml composite composition administration group of three concentrations, the thickness of the mucous membrane of the fecal colonic mucosa and the number of mucous producing cells were increased compared to the administration of ondansetron 0.1 mg / The increase in the thickness of the mucous layer on the feces surface was dependent on the complex BH concentration. These results are considered to be one of the direct proofs that the underdosing of ondansetron is synergistically increased by the BH complex composition of at least 10 mg / ml under the conditions of this experiment.

Gastrin is a typical hormone that produces, secretes, and promotes gastric acid secretion in the gastric mucosa, especially in endocrine cells that are located locally in the pyloric mucosa [Lee et al., 2010]. Cisplatin results in a decrease in gastrointestinal motility disturbances, particularly in gastrointestinal tract excretion [Bradner and Schurig, 1981], resulting in a marked decrease in gastrin production, resulting in indigestion or increased diarrhea [Wang and Aggarwal, 1997 ; Wang et al., 2000; Song et al., 2012]. As a result of this experiment, the administration of all BH alone, 0.1 mg / ml ondansetron and BH 40, 20 or 10 mg / ml combination composition resulted in significant decrease in gastrin content and gastrin immunoreactive cells in the pyloric gastric mucosa by cisplatin In contrast, the concentration of gastrin in the pyloric gastric mucosa and the number of gastrin-immunoreactive cells increased significantly compared to the administration of ondansetron 0.1 mg / ml alone in all three concentrations of all BH and ondansetron 0.1 mg / Respectively. These results suggest that the administration of BH resulted in restoration of gastrointestinal motility disorder and secondary recovery of gastrin production and gastrin immunoreactive cells. However, in the case of ondansetron, despite decreased gastrointestinal motility and gastric mucosal damage, It was observed that the gastrin content in the pyloric gastric mucosa and the number of gastrin - immunoreactive cells did not affect the change significantly. Therefore, more systematic research is needed.

At present, more than 90% of serotonin is known to be present in the digestive tract [Erspamer and Testinini, 1959]. Serotonin in the gastrointestinal tract is synthesized by tryptophan hydroxylase and then stored in gastrointestinal endocrine cells, enterochromaffin cells [Yu et al., 1999], secreted from the gastrointestinal endocrine cells into the interstitial space by various stimuli [Rackeand Schwo1991]. The secreted serotonin was first introduced into the epithelial cells of the gastrointestinal tract by the serotonin reuptake transporter [Wada et al., 1996], and by the intracellular enzyme monoamine oxidase present in the gastric mucosa and submucosa, 5-hydroxyindoleacetic acid It is known to be degraded and removed [Saura et al., 1992]. Cisplatin, on the other hand, inhibits the activity of monoamine oxidase, an intracellular enzyme responsible for the synthesis of serotonin and the serotonin 5-HT3 receptors in endocrine cells in the gastrointestinal tract, or the degradation of serotonin, (Bradner and Schurig, 1981; Kilpatrick et al., 1990; Tyers, 1991]. As a result of this study, it was observed that administration of all BH alone and ondansetron 0.1 mg / ml and BH combination composition normalized tryptophan hydroxylase activity and inhibition of monoamine oxidase activity by cisplatin. Especially, ondansetron 0.1 mg / In the all BH 40, 20 and 10 mg / ml combination composition administration groups, the decrease in activity of tryptophan hydroxylase and the increase in monoamine oxidase activity were found to be dependent on the complex BH concentration, compared with the administration of ondansetron 0.1 mg / The effect of synergistic ondansetron sublingual enhancement on the activity of tryptophan hydroxylase and monoamine oxidase is thought to be partially mediated by the activity of tryptophan hydroxylase and monoamine oxidase. Ondansetron, on the other hand, is a typical 5-HT3 receptor blocking drug [Beck et al., 1993; Miyata et al., 1995; Martin et al., 1998; Ozmen and Ku2004; Malik et al., 2007] did not significantly affect the activity of tryptophan hydroxylase and monoamine oxidase.

Thus, the complex composition of BH 40, 20 or 10 mg / ml significantly increases the pharmacological effect of ondansetron on cisplatin-induced gastrointestinal tract disorders through antioxidant, gastrin and serotonin-producing gastrointestinal endocrine cell synthesis, secretion or function regulation Respectively. Thus, a BH complex composition at a concentration of 10 mg / ml or greater did not affect the bioavailability of ondansetron [CIMI-14-02-11-02 O-BH PK, 2015; CIMI-14-02-11-04 O-BH PK (2), 2015] provides a new approach to cancer therapy that can control the digestive tract disorders caused by various chemotherapeutic agents including cisplatin in cancer patients. It is expected to be able to give. In this study, ondansetron was shown to be a typical 5-HT3 receptor blocker and did not significantly affect the production, secretion and degradation of serotonin in tissues.

It will be understood by those skilled in the art that the foregoing description of the present invention is for illustrative purposes only and that those of ordinary skill in the art can readily understand that various changes and modifications may be made without departing from the spirit or essential characteristics of the present invention. will be. It is therefore to be understood that the above-described embodiments are illustrative in all aspects and not restrictive.

Claims (6)

A composition for enhancing the therapeutic effect of nausea and vomiting, which comprises Ondansetron and royal jelly extract, and alleviates side effects due to nausea and vomiting treatment. The method according to claim 1,
Wherein said nausea and vomiting are induced by chemotherapy. delete The method according to claim 1,
Characterized in that the ondansetron and royal jelly extract are pre-mixed and formulated or separately formulated. The method according to claim 1,
Characterized in that the ondansetron and royal jelly extract is administered parenterally, orally, locoregionally, or percutaneously. The method according to claim 1,
Wherein the administration of the royal jelly extract begins between 5 minutes and 4 hours after administration of the ondansetron.

Images