Toltrazuril nano suspension and preparation method thereof

Abstract

The invention discloses a toltrazuril nano suspension and a preparation method thereof, and belongs to the technical field of veterinary drug preparations. The suspension contains toltrazuril, an iron compound, a bacteriostatic agent, a suspending aid, a stabilizer and the balance of water. According to the invention, an ultrahigh-pressure nano homogenizer technology is adopted, the medicine is prepared into a nano-scale particle size, and the medicine is uniformly dispersed in the preparation, so that the medicine has good stability and dispersibility. The preparation not only can be used for effectively preventing and treating animal coccidiosis, but also can improve animal anemia, and has a wide clinical application prospect.

Description

Technical Field

[0001] This invention belongs to the field of veterinary drug formulation technology, specifically relating to a toltrazuril nano-suspension and its preparation method. Background Technology

[0002] Coccidiosis is a common disease in animal husbandry. Coccidia are highly resilient, and their oocysts are very resistant to the external environment, surviving in soil and bedding for months or even years. Young animals (such as chickens, pigs, cattle, sheep, and rabbits) show more severe symptoms after infection, including diarrhea (which may be bloody), emaciation, anemia, loss of appetite, and lethargy, with a mortality rate of 50%-80%. Adult animals often experience a chronic course of infection, with slower growth, reduced feed conversion ratio (increased feed conversion ratio), and decreased egg production. Globally, coccidiosis causes billions of dollars in losses to the livestock industry annually.

[0003] Iron is an essential trace element for maintaining normal life activities and preventing anemia in animals. Newborn piglets contain only 40-50 mg of total iron, and the iron content in breast milk is only 0.2 mg / 100g, making them prone to iron-deficiency anemia. Intestinal bleeding caused by coccidiosis further promotes the development of anemia. Amino acid chelated iron is a representative of third-generation iron supplements. Compared with traditional iron supplements such as ferrous sulfate, ferrous gluconate, and ferrous fumarate, amino acid chelated iron is absorbed in the small intestine in the form of compounds using the absorption mechanisms of peptides and amino acids. Because no iron ions are produced, it has virtually no irritation to the intestinal mucosa. Numerous experiments have confirmed that amino acid iron has high solubility while avoiding antagonistic effects between iron and other mineral elements and anti-nutritional substances, thus resulting in high absorption and utilization rates. Many amino acids can promote iron absorption; methionine iron, glycine iron, and lysine iron are among the most widely used.

[0004] Toltrazuril is a triazine broad-spectrum anticoccidial drug with a unique mechanism of action. It inhibits cell nuclear division and organelle development in coccidia, interfering with energy metabolism and biosynthesis, thus fundamentally blocking the growth and reproduction of coccidia. Toltrazuril has a good killing effect on Eimeria coccidia and also has some control effect on other coccidia. Toltrazuril has low toxicity and has no adverse effects on the growth and production performance of economically important animals such as chickens and pigs. Currently, commercially available preparations containing toltrazuril include solutions, oral suspensions, and injections. Among them, solutions are used for administration to chickens through drinking water, but their solubility in tap water or local water is low, which cannot meet the needs of centralized administration in large and super-large buildings in large-scale farms. Oral suspensions are for livestock, requiring individual gavage administration, which is inconvenient and labor-intensive. Injections cause significant stress to animals and have high labor costs.

[0005] Patent CN101678031 discloses a formulation containing triazine ketones and dextran iron, which is micron-sized and suitable for individual administration, but not for group administration. Patent CN116672360A is an injectable formulation of toltrazuril and glucohepantran anhydride iron, which is suitable for individual administration and has a high stress response in animals.

[0006] Therefore, there is an urgent need in this field to develop a toltrazuril nano-oral suspension that is convenient to administer, has low stress on animals, is suitable for group administration, has low irritation, has strong stability in group administration, and has good suspension effect, and can significantly prevent and treat coccidiosis, iron deficiency, and hemorrhagic anemia caused by coccidiosis. Summary of the Invention

[0007] The present invention aims to solve the technical problems in the prior art and provide a tortuzumab nanosuspension and its preparation method.

[0008] To achieve the above objectives, the specific technical solution of the present invention is as follows:

[0009] In a first aspect, the present invention provides a tortezulide nanosuspension containing tortezulide and an iron compound.

[0010] Preferably, the suspension further contains an antibacterial agent, a suspending agent, a stabilizer, and water.

[0011] More preferably, the iron compound is amino acid iron.

[0012] Preferably, the iron compound is one of glycine iron, methionine iron, and lysine iron.

[0013] More preferably, the iron compound is iron glycine.

[0014] Preferably, the suspension comprises, by mass-volume ratio: 1-10% toltrazuril, 10-30% iron compound, 0.1-1% antibacterial agent, 0.1-2% surfactant, 0.1-2% suspending agent, 0.05-3% stabilizer, with the remainder being purified water.

[0015] More preferably, the suspension comprises, by mass-volume ratio: 2.5-10% toltrazuril, 15-30% iron compound, 0.2-0.5% antibacterial agent, 0.2-0.6% surfactant, 0.15-0.5% suspending agent, 0.1-0.6% stabilizer, and the balance being purified water.

[0016] More preferably, the suspension comprises, by mass-volume ratio: 5% toltrazuril, 25% iron compound, 0.3% antibacterial agent, 0.5% surfactant, 0.25% suspending agent, 0.2% stabilizer, and the remainder being purified water.

[0017] Preferably, the antibacterial agent is one or more of benzyl alcohol, sodium benzoate, sodium propionate, potassium sorbate, ethylparaben, and methylparaben; more preferably, the antibacterial agent is potassium sorbate.

[0018] Preferably, the surfactant is one or more of sodium lauryl sulfate, sodium deoxycholate, polyoxyethylene hydrogenated castor oil, polyoxyethylene stearate, sodium dioctyl succinate sulfonate, polyvinylpyrrolidone (PVP K30), and polyethylene glycol 15 hydroxystearate (HS-15); more preferably, the surfactant is sodium dioctyl succinate sulfonate.

[0019] Preferably, the suspending agent is one or more of microcrystalline cellulose, xanthan gum, sodium carboxymethyl cellulose, methyl cellulose, carboxypropyl methyl cellulose E50, polyvinyl alcohol, and carbomer; more preferably, the suspending agent is sodium carboxymethyl cellulose, microcrystalline cellulose, or xanthan gum.

[0020] Preferably, the stabilizer is one or more of vitamin C, butylated hydroxyanisole, propyl gallate, triacetic acid, sodium hydroxymethylsulfonate, sodium bisulfite, and sodium thiosulfate; more preferably, the stabilizer is propyl gallate.

[0021] In a second aspect, the present invention provides a method for preparing tortrolyl nanosuspension, the steps of which are as follows:

[0022] (1) Dissolve the prescribed amount of surfactant, antibacterial agent and stabilizer in a portion of purified water in sequence until completely dissolved to obtain solution A;

[0023] (2) Add to thortrixil to solution A and stir until homogeneous, then add amino acid iron and stir to dissolve, to obtain solution B;

[0024] (3) Dissolve the suspending agent in some purified water and stir until uniform, then add solution B and stir until uniform to obtain solution C;

[0025] (4) Pass liquid C through an emulsifying disperser to obtain a preliminary mixture, and then homogenize the preliminary mixture through an ultra-high pressure nano homogenizer to obtain liquid D;

[0026] (5) Add the remaining purified water to solution D and stir until homogeneous to obtain tortuzumab nano suspension.

[0027] Further, in step (4), the speed of the emulsifying disperser is 10,000 rpm to 20,000 rpm; in the ultra-high pressure nano homogenizer, the homogenization pressure is 15,000 to 40,000 psi, and the number of cycles is 5 to 10, to obtain a nano suspension.

[0028] Beneficial effects:

[0029] 1. The formulation of this invention adds an amino acid iron chelate, which is less irritating to the intestinal mucosa than other iron ions, and at the same time prevents and treats coccidiosis, iron deficiency and enteritis bleeding symptoms caused by coccidiosis in newborn piglets.

[0030] 2. The process of this invention uses ultra-high pressure nano-homogenization technology to control the raw materials at the nanoscale, which improves the bioavailability of the drug while ensuring the formulation is stable, has strong suspension properties, good dispersibility, and excellent resuspensibility.

[0031] 3. The nano-sized oral suspension of this invention has a smaller active ingredient particle size than ordinary micron-sized suspensions, making it easier to absorb. It can be mixed with water in any proportion and then uniformly suspended in water without precipitation, blockage of dosing equipment and water lines, and is not affected by local water quality.

[0032] 4. The nanoscale oral suspension of this invention does not require injection, has less stress than injection-based administration, and is convenient to administer. It can be administered orally individually or via drinking water for groups, meeting the needs of large-scale farms for group medication. Detailed Implementation

[0033] The above content will be further described in detail below through specific embodiments. However, this should not be construed as limiting the scope of the above subject matter to the following embodiments. All technologies implemented based on the content of this invention fall within the scope.

[0034] Example 1

[0035] A toltrazuril nano-oral suspension, the formulation and preparation method of which are as follows:

[0036] composition Element Weight (per 100ml) Active ingredients Tochuzhuli 5g Amino acid iron Iron glycine 15g antibacterial agent Potassium sorbate 0.3g surfactants HS-15 0.2g stabilizer propyl gallate 0.5g Suspending agent Sodium carboxymethyl cellulose 0.3g water Purified water Up to 100ml

[0037] Preparation method:

[0038] (1) Dissolve the prescribed amount of surfactant, antibacterial agent and stabilizer in a portion of the prescribed amount of purified water in sequence until completely dissolved to obtain solution A.

[0039] (2) Add to thortrixol to solution A and stir to disperse evenly, then add amino acid iron and stir to dissolve, to obtain solution B.

[0040] (3) Dissolve the suspending agent in some purified water and stir until dissolved. Then add solution B and stir until homogeneous to obtain solution C.

[0041] (4) The C liquid is passed through an emulsifying disperser to obtain a preliminary mixture, and the preliminary mixture is ground by an ultra-high pressure nano homogenizer to obtain the D liquid; the speed of the emulsifying disperser is 12000 rpm; in the ultra-high pressure nano homogenizer, the homogenization pressure is 15000 psi, and the number of cycles is 10 to obtain a nano suspension.

[0042] (5) Add the remaining purified water to solution D and stir until homogeneous to obtain the finished nano suspension.

[0043] Example 2

[0044] A toltrazuril nano-oral suspension, the formulation and preparation method of which are as follows:

[0045] composition Element Weight (per 100ml) Active ingredients Tochuzhuli 5g Amino acid iron Iron glycine 25g antibacterial agent Potassium sorbate 0.3g surfactants Sodium lauryl sulfate 0.5g stabilizer Nitroglycerin 0.2g Suspending agent Xanthan Gum 0.3g water Purified water Up to 100ml

[0046] Preparation method:

[0047] (1) Dissolve the prescribed amount of surfactant, antibacterial agent and stabilizer in a portion of the prescribed amount of purified water in sequence until completely dissolved to obtain solution A.

[0048] (2) Add to thortrixol to solution A and stir to disperse evenly, then add amino acid iron and stir to dissolve, to obtain solution B.

[0049] (3) Dissolve the suspending agent in some purified water and stir until dissolved. Then add solution B and stir until homogeneous to obtain solution C.

[0050] (4) The initial mixture of liquid C is obtained by passing it through an emulsifying disperser. The initial mixture is then homogenized by an ultra-high pressure nano-homogenizer to obtain liquid D. The speed of the emulsifying disperser is 15000 rpm. The homogenization pressure in the ultra-high pressure nano-homogenizer is 22000 psi, and the number of cycles is 6 to obtain a nano-suspension.

[0051] (5) Add the remaining purified water to solution D and stir until homogeneous to obtain the finished nano suspension.

[0052] Example 3

[0053] A toltrazuril nano-oral suspension, the formulation and preparation method of which are as follows:

[0054]

[0055] The preparation method is as follows:

[0056] (1) Dissolve the prescribed amount of surfactant, antibacterial agent and stabilizer in a portion of the prescribed amount of purified water in sequence until completely dissolved to obtain solution A.

[0057] (2) Add to thortrixol to solution A and stir to disperse evenly, then add amino acid iron and stir to dissolve, to obtain solution B.

[0058] (3) Dissolve the suspending agent in some purified water and stir until dissolved. Then add solution B and stir until homogeneous to obtain solution C.

[0059] (5) The C liquid is passed through an emulsifying disperser to obtain a preliminary mixture, and the preliminary mixture is homogenized by an ultra-high pressure nano homogenizer to obtain the D liquid; the speed of the emulsifying disperser is 15000 rpm; in the ultra-high pressure nano homogenizer, the homogenization pressure is 20000 psi, and the number of cycles is 10 to obtain a nano suspension.

[0060] (4) Add the remaining purified water to solution D and stir until homogeneous to obtain the finished nano suspension.

[0061] Example 4

[0062] A toltrazuril nano-oral suspension, the formulation and preparation method of which are as follows:

[0063]

[0064] Preparation method:

[0065] (1) Dissolve the prescribed amount of surfactant and antibacterial agent in a portion of the prescribed amount of purified water in sequence until completely dissolved to obtain solution A.

[0066] (2) Add to thortrixol to solution A and stir to disperse evenly, then add amino acid iron and stir to dissolve, to obtain solution B.

[0067] (3) Dissolve the suspending agent in some purified water and stir until dissolved. Then add solution B and stir until homogeneous to obtain solution C.

[0068] (4) The initial mixture of liquid C is obtained by passing it through an emulsifying disperser. The initial mixture is then homogenized by an ultra-high pressure nano-homogenizer to obtain liquid D. The speed of the emulsifying disperser is 18000 rpm. The homogenization pressure in the ultra-high pressure nano-homogenizer is 20000 psi, and the number of cycles is 8 to obtain a nano-suspension.

[0069] (5) Add the remaining purified water to solution D and stir until homogeneous to obtain the finished nano suspension.

[0070] Example 5

[0071] A toltrazuril nano-oral suspension, the formulation and preparation method of which are as follows:

[0072]

[0073]

[0074] Preparation method:

[0075] (1) Dissolve the prescribed amount of surfactant and antibacterial agent in a portion of the prescribed amount of purified water in sequence until completely dissolved to obtain solution A.

[0076] (2) Add to thortrixol to solution A and stir to disperse evenly, then add amino acid iron and stir to dissolve, to obtain solution B.

[0077] (3) Dissolve the suspending agent in some purified water and stir until dissolved. Then add solution B and stir until homogeneous to obtain solution C.

[0078] (4) The initial mixture of liquid C is obtained by passing it through an emulsifying disperser. The initial mixture is then homogenized by an ultra-high pressure nano-homogenizer to obtain liquid D. The speed of the emulsifying disperser is 18000 rpm. The homogenization pressure in the ultra-high pressure nano-homogenizer is 35000 psi, and the number of cycles is 8 to obtain a nano-suspension.

[0079] (5) Add the remaining purified water to solution D and stir until homogeneous to obtain the finished nano suspension.

[0080] Example 6: Settlement Investigation

[0081] The sedimentation volume ratio and redispersibility of the formulations in Examples 1, 2, 3, 4, and 5 were investigated.

[0082] Sedimentation volume ratio: Measure 50ml of suspension using a stoppered graduated cylinder, stopper tightly, shake vigorously for 1min, record the initial height H0 of the suspension, and after standing for 3h, record the final height H of the sediment. Then the sedimentation volume ratio F = H / H0, and the F value should not be lower than 0.9.

[0083] Redispersibility: Take an appropriate amount of the preparation into a centrifuge tube, centrifuge at 500 rpm for 10 min, remove the tube, invert it with constant force, and record the time required for the preparation to disperse evenly. The shorter the time, the better the dispersibility.

[0084] The sedimentation and dispersion are shown in Table 1:

[0085] Table 1. Settlement and Dispersion

[0086] Example Particle size (nm) D90 Redispersion(s) Settlement volume ratio 1 288 48±5 <0.90 2 281 36±3 1.0 3 284 28±5 1.0 4 279 30±3 1.0 5 280 24±3 1.0

[0087] Example 7 Stability Study

[0088] The stability and quality of the formulations in Examples 2, 3, 4, and 5 were investigated. Samples from the formulations of these examples were placed under high-temperature conditions (60°C) for 10 days and under accelerated conditions (40°C ± 2°C, RH 75% ± 5%) for 1 month, 2 months, 3 months, and 6 months, respectively. Particle size, toltrazuril content, and total iron content were measured. The results are shown in Table 2.

[0089] Table 2 Stability test results

[0090]

[0091] The stability and quality studies of the formulations showed that, for Examples 2, 3, 4, and 5, the content and particle size of toltrazuril did not change significantly after being placed at high temperature (60°C) for 10 days and under accelerated conditions (40°C ± 2°C, RH 75% ± 5%) for 1 month, 2 months, 3 months, and 6 months, respectively. However, the total iron content of Example 2 decreased more rapidly compared to Examples 3, 4, and 5.

[0092] Example 8: Clinical Efficacy Trial

[0093] The clinical efficacy of the nanosuspensions obtained in Examples 3-5 of this invention was verified based on conventional anticoccidial drugs.

[0094] One hundred and fifty 3- to 5-day-old piglets infected with *Isospora suis* were randomly divided into five groups: a positive control group, Example 3 group, Example 4 group, Example 5 group, and Comparative Example 1 group (patent EP2276347A1 group), with thirty piglets in each group. The positive control group received no treatment. The treatment groups received a single dose of toltrazuril at 20 mg / kg body weight for 21 consecutive days. Clinical symptoms, diarrhea, number of deaths, body weight, intestinal lesions, number of oocysts, hemoglobin concentration, diarrhea rate, mortality rate, cure rate, and average daily weight gain were recorded. The experimental results are shown in Tables 3, 4, and 5.

[0095] Table 3 Health and survival status of each group

[0096] Group positive group Example 3 Example 4 Example 5 Comparative Example 1 mortality rate(%) 6.67 0.00 0.00 0.00 0.00 Cure rate (%) / 100 100 100 100 Diarrhea rate (%) 96.43 0.00 0.00 0.00 3.33

[0097] Table 4. Results of intestinal lesions, ovarian cysts, and hemoglobin detection in each group.

[0098] Group positive group Example 3 Example 4 Example 5 Comparative Example 1 Intestinal lesion value 2.33 0.00 0.00 0.00 0.33 ovarian cyst reduction rate / 95.67％ 95.82％ 96.52％ 93.63％ Hemoglobin concentration (g / L) 74.4±3.1 101.2±3.2 108.7±6.4 112.0±4.6 89.5±4.7

[0099] Table 5. Average daily weight gain (g / day / head) for each group at different time periods.

[0100] time positive group Example 3 Example 4 Example 5 Comparative Example 1 0-7 days 116.32 139.56 140.38 140.22 124.72 7-14 days 122.43 147.25 146.52 151.47 142.51 14-21 days 186.84 237.34 242.51 245.13 215.67

[0101] Average daily weight gain: Total weight gain per period / Number of days / Number of pigs

[0102] Mortality rate: Number of pigs that died at the end of the trial / Number of pigs × 100%.

[0103] Cure rate: (Number of cured piglets / Number of piglets) × 100% at the end of the experiment. Piglets are considered cured when diarrhea stops, they are in good spirits, and their feed intake returns to normal.

[0104] Diarrhea rate: Statistical analysis on day 7, calculated as number of pigs with diarrhea / number of pigs × 100%.

[0105] Oocyst reduction rate: Feces were collected once a day to calculate the number of coccidia oocysts (OPG) in pig feces. The total number of oocysts detected in each group from 1 to 11 days after drug administration was counted. Oocyst reduction rate = (OPG of positive group - OPG of drug group) / OPG of positive group × 100%.

[0106] The scoring criteria for intestinal lesions were as follows: on the 7th day after drug administration, 3 pigs from each group were randomly dissected to examine the intestinal lesions.

[0107] 0 points, no visible lesions;

[0108] +1 point, very few scattered lesions;

[0109] +2 points, with more sparse lesions, such as thinning of the intestinal wall, intestinal gas, bleeding spots on the mucosa, and redness and swelling of the mesenteric lymph nodes;

[0110] +3 points for catarrhal inflammation, thinning of the intestinal wall, gas filling the intestines, mucosal hemorrhage or hemorrhagic spots, and edematous enlargement of mesenteric lymph nodes; +3 points are also awarded for pig deaths.

[0111] The results showed that, compared with the comparative examples, toltrazuril nanosuspension could better improve diarrhea in piglets, reduce oocyst excretion, and reduce intestinal lesions. Groups 3-5 had higher hemoglobin levels, lower diarrhea rates, lower intestinal lesion values, higher oocyst reduction rates, and faster growth and development, indicating that the toltrazuril nanosuspension described in this invention can better prevent and treat coccidiosis in piglets and protect the intestinal mucosa from coccidial damage.

[0112] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, and not to limit them. Although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art should understand that modifications can still be made to the technical solutions described in the foregoing embodiments, or equivalent substitutions can be made to some or all of the technical features therein; these modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the scope of the technical solutions of the embodiments of the present invention.

Claims

1. A toltrazuril nanosuspension, characterized in that, The suspension contains toltrazuril and an iron compound; the suspension also contains an antibacterial agent, a suspending agent, a stabilizer, and water; the iron compound is amino acid iron.

2. The suspension according to claim 1, characterized in that, The iron compound is one of glycine iron, methionine iron, and lysine iron; preferably glycine iron.

3. The suspension according to claim 1, characterized in that, The suspension, by mass-volume ratio, comprises: 1-10% toltrazuril, 10-30% iron compound, 0.1-1% antibacterial agent, 0.1-2% surfactant, 0.1-2% suspending agent, 0.05-3% stabilizer, with the remainder being purified water.

4. The suspension according to claim 3, characterized in that, The suspension, by mass-volume ratio, comprises: 2.5-10% toltrazuril, 15-30% iron compound, 0.2-0.5% antibacterial agent, 0.2-0.6% surfactant, 0.15-0.5% suspending agent, 0.1-0.6% stabilizer, and the remainder being purified water.

5. The suspension according to claim 4, characterized in that, The antibacterial agent is one or more of benzyl alcohol, sodium benzoate, sodium propionate, potassium sorbate, ethylparaben, and methylparaben.

6. The suspension according to claim 4, characterized in that, The surfactant is one or more of sodium lauryl sulfate, sodium deoxycholate, polyoxyethylene hydrogenated castor oil, polyoxyethylene stearate, sodium dioctyl succinate sulfonate, PVP K30, and HS-15.

7. The suspension according to claim 4, characterized in that, The suspending agent is one or more of microcrystalline cellulose, xanthan gum, sodium carboxymethyl cellulose, methyl cellulose, carboxypropyl methyl cellulose E50, polyvinyl alcohol, and carbomer.

8. The suspension according to claim 4, characterized in that, The stabilizer is one or more of vitamin C, butylated hydroxyanisole, propyl gallate, triacetic acid, sodium hydroxymethylsulfonate, sodium bisulfite, and sodium thiosulfate.

9. A method for preparing a tortezulil nanosuspension as described in any one of claims 1-8, comprising the following steps: (1) Dissolve the prescribed amount of surfactant, antibacterial agent and stabilizer in a portion of purified water in sequence until completely dissolved to obtain solution A; (2) Add to thortrixil to solution A and stir until homogeneous, then add amino acid iron and stir to dissolve, to obtain solution B; (3) Dissolve the suspending agent in some purified water and stir until uniform, then add solution B and stir until uniform to obtain solution C; (4) Pass liquid C through an emulsifying disperser to obtain a preliminary mixture, and then homogenize the preliminary mixture through an ultra-high pressure nano homogenizer to obtain liquid D; (5) Add the remaining purified water to solution D and stir until homogeneous to obtain tortuzumab nano suspension.

10. The preparation method according to claim 9, characterized in that, The speed of the emulsifying disperser in step (4) is 10,000 rpm to 20,000 rpm; the homogenization pressure in the ultra-high pressure nano homogenizer is 10,000 to 40,000 psi, and the number of cycles is 5 to 10, so as to obtain a nano suspension.

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