Reduced coenzyme q10 crystal and preparation method therefor

Abstract

The present invention provides a reduced coenzyme Q10 crystal which, when measured by means of differential scanning calorimetry at a heating rate of 10 k / min, exhibits an endothermic peak at 52±2°C. Compared with the prior art, the crystal form of reduced coenzyme Q10 provided by the present invention is more stable than reduced coenzyme Q10 crystals reported in the literature, and other physical properties, including water solubility and residual solvents, are also superior; in addition, the crystal form provided by the present invention also overcomes the disadvantages of previously known reduced coenzyme Q10, which is very easily oxidized and has limitations in use; furthermore, the reduced coenzyme Q10 crystal and a crystalline solid containing the crystal provided by the present invention not only have excellent physical properties in terms of stability, but also stand out in terms of high purity and low solvent residue.

Description

A reduced coenzyme Q10 crystal and preparation method thereof

[0001] This application claims priority to the Chinese patent application filed with the China Patent Office on December 19, 2023, with application number 202311751931.0 and invention name “A Reduced Coenzyme Q10 Crystal and Its Preparation Method”, the entire contents of which are incorporated by reference into this application. Technical Field

[0002] The present invention belongs to the technical field of compound crystals, and in particular relates to a reduced coenzyme Q10 crystal and a preparation method thereof. Background Art

[0003] Coenzyme Q10, also known as ubiquinone 10, is chemically named 2,3-dimethoxy-5-methyl-6-decaisopentenylbenzoquinone. Its structure is similar to vitamin K. Scientists discovered and confirmed its chemical structure in the mid-20th century. Coenzyme Q10 is a vitamin-like substance, a fat-soluble organic quinone compound widely distributed throughout the body, with important physiological and pharmacological effects. In the early 1980s, Swedish researcher Ernster revealed the antioxidant and free radical scavenging properties of the vitamin-like substance coenzyme Q10. In 1972, Harman demonstrated the relationship between mitochondrial function and aging. Coenzyme Q10 primarily binds to the mitochondrial membrane, forming a coenzyme in the respiratory chain and participating in hydrogen transfer during energy metabolism. Coenzyme Q10 is a metabolic activator that stimulates cellular respiration, generates cellular power, and accelerates the production of adenosine triphosphate (ATP). Coenzyme Q10 itself is a natural antioxidant produced by cells, inhibiting mitochondrial peroxidation and protecting the integrity of biological membranes. It is also a non-vitamin nutrient that activates cellular respiration, accelerates the production of high-energy ATP, strengthens myocardial metabolism, improves cardiac efficiency, and regulates cellular and tissue hypoxia. It has a protective and beneficial effect on the liver, brain, heart, and nervous system. Coenzyme Q10 also has a highly specific immune-enhancing effect, increasing the phagocytic rate of phagocytes and boosting antibody production. Numerous clinical studies both domestically and internationally have demonstrated that Coenzyme Q10 has excellent therapeutic effects on conditions such as heart disease, hypertension, cerebrovascular disease, scurvy, and viral hepatitis. It can act as a nonspecific immune enhancer and activator of cellular metabolism and respiration. Recently, it has also been shown to have significant therapeutic benefits in the treatment of cancer and AIDS. Therefore, Coenzyme Q10 plays an irreplaceable role and has broad application prospects in human health care, delaying aging, and improving the body's immunity. At the same time, because Coenzyme Q10 is non-toxic, has no side effects, and does not interact with any other drugs, it has become an important medicine and health product.

[0004] Most of the coenzyme Q10 currently sold on the market is oxidized coenzyme Q10, but studies have found that the two-electron reduced form of oxidized coenzyme Q10, reduced coenzyme Q10, shows higher oral absorption than oxidized coenzyme Q10, and the reduced coenzyme Q10 is also the main form that plays a role in the body. The only difference between the two is that one is a benzoquinone form and the other is a benzenediol form.

[0005] Furthermore, among the numerous publicly available literature on reduced oxy-Coenzyme Q10 production technologies, most, with the exception of extraction from fermentation broth (where the majority of the product is reduced Coenzyme Q10 during fermentation, which gradually oxidizes to oxidized Coenzyme Q10 during extraction), use oxidized Coenzyme Q10 as a raw material and reduce it to reduced Coenzyme Q10 using conventional reducing agents. These reducing agents include sodium borohydride, sodium dithionite, ascorbic acid, and certain amino acids, while the solvents used primarily include aliphatic hydrocarbons and fatty acid esters. Furthermore, several methods are known for obtaining reduced Coenzyme Q10 in the form of crystals. For example, a method for producing crystals by precipitating reduced Coenzyme Q10 in an alcohol solution and / or a ketone solution (WO 2003 / 006409) and a method for crystallization by adding a high-concentration liquid phase of reduced Coenzyme Q10 to a poor solvent (Japanese Patent Application Laid-Open No. 2003-089669) have been reported.

[0006] Meanwhile, patent document WO2012 / 176842 reports the observation of polymorphism in reduced coenzyme Q10 and the discovery of a new crystalline form that differs from the aforementioned document. The report also reports that the newly discovered crystalline form is significantly more stable and has superior other physical properties compared to existing reduced coenzyme Q10, and discloses its production method. The report also reports that the newly discovered crystalline form (hereinafter referred to as Form II reduced coenzyme Q10 crystals or Form II crystals) is significantly more stable and has superior other physical properties compared to existing reduced coenzyme Q10 (hereinafter referred to as Form I reduced coenzyme Q10 crystals or Form I crystals).

[0007] However, due to its structure, reduced Coenzyme Q10 is primarily present in the fermentation broth, gradually becoming oxidized during extraction. Since reduced Coenzyme Q10 is easily oxidized to oxidized Coenzyme Q10 by atmospheric oxygen during chemical production, complete elimination of molecular oxygen is extremely difficult to achieve on a commercial scale. Consequently, residual oxygen during the manufacturing process exerts a significant negative impact, generating nearly ineliminable oxidized Coenzyme Q10, which subsequently contaminates the product and reduces its purity. Studies have also shown that, while the product is relatively stable in solvents, it is susceptible to oxidation during filtration, drying, and storage in the liquid phase. To obtain high-purity reduced Coenzyme Q10 in crystalline form, it is crucial to adequately protect the reduced Coenzyme Q10 from these oxidation reactions.

[0008] In addition, the literature mentions that the crystal form has a relatively large influence on its oxidation rate, but studies have shown that the crystal form obtained by using different embodiments of the existing literature (WO2012 / 176842, Form II) is actually still Form I, rather than Form II as reported in the literature, according to powder X-ray diffraction. Some literature also reports the use of oil encapsulation or the addition of antioxidants, but these will introduce new substances that are detrimental to the product. In addition, in addition to ethanol, the use of aliphatic hydrocarbon solvents such as n-hexane and n-heptane for purification has the problem that the solvent residue is difficult to meet the requirements. Since the melting point of oxidized or reduced coenzyme Q10 is around 50°C, it is easy to liquefy if the drying temperature is increased. The color of the product obtained by ethanol crystallization is easy to change, the appearance is yellow, and it is in a slurry form, which is difficult to filter.

[0009] Summary of the Invention

[0010] In view of this, the technical problem to be solved by the present invention is to provide a reduced coenzyme Q10 crystal with high stability and purity and a preparation method thereof.

[0011] The invention provides a reduced coenzyme Q10 crystal, which has an endothermic peak at 52±2° C. when the temperature is increased at a speed of 10 k / min as measured by differential scanning calorimetry.

[0012] Preferably, in powder X-ray diffraction measured using Cu-Kα radiation, characteristic peaks are shown at diffraction angles (2θ±0.2°) of 8.95°, 10.04°, 15.09°, 18.65°, 19.03°, 21.61°, and 23.01°.

[0013] Preferably, the infrared absorption spectrum of the reduced coenzyme Q10 crystal measured by the KBr pellet method is at a wave number of 794±1cm -1 、877±1cm -1 、962cm -1 and 1014cm -1It has a characteristic absorption peak.

[0014] Preferably, it has a powder X-ray diffraction pattern measured with Cu-Kα radiation as shown in FIG1 ;

[0015] And / or, having an infrared absorption spectrum measured by the KBr pellet method as shown in FIG2 ;

[0016] And / or, having the differential scanning calorimetry graph shown in FIG3 .

[0017] The present invention also provides a method for preparing reduced coenzyme Q10 crystals, comprising the following steps:

[0018] S) crystallizing the reduced coenzyme Q10 in the presence of a cyclic ether solvent, a fluorinated alcohol solvent, and an organic base to obtain reduced coenzyme Q10 crystals.

[0019] Preferably, the step S) is specifically:

[0020] The reduced coenzyme Q10 and a cyclic ether solvent are mixed and dissolved under heating conditions, and then a mixed solution of a fluorinated alcohol solvent and an organic base is added, and the temperature is lowered for crystallization to obtain reduced coenzyme Q10 crystals;

[0021] The crystallization temperature is 0°C to 20°C.

[0022] Preferably, the cyclic ether solvent is selected from one or more of 2-methyltetrahydrofuran, 3-methyltetrahydrofuran, tetrahydrofuran and 2,5-dimethyltetrahydrofuran;

[0023] The fluorine-containing alcohol solvent is selected from one or more of trifluoromethanol, trifluoroethanol, trifluoropropanol and trifluorobutanol;

[0024] The organic base is selected from one or more of triethylamine, pyridine and piperidine.

[0025] Preferably, the ratio of the reduced coenzyme Q10 to the cyclic ether solvent is 1 g: (1-10) mL;

[0026] The volume ratio of the cyclic ether solvent to the fluorinated alcohol solvent is 1:(1-10);

[0027] The mass of the organic base is 0.1% to 10% of the mass of the reduced coenzyme Q10.

[0028] The present invention also provides a reduced coenzyme Q10 crystalline solid, comprising the above-mentioned reduced coenzyme Q10 crystal.

[0029] The present invention also provides a reduced coenzyme Q10 composition, comprising the above-mentioned reduced coenzyme Q10 crystals;

[0030] and / or the above-mentioned reduced coenzyme Q10 crystalline solid.

[0031] The present invention provides reduced coenzyme Q10 crystals, which exhibit an endothermic peak at 52±2°C when heated at a rate of 10 k / min, as determined by differential scanning calorimetry. Compared to existing technologies, the reduced coenzyme Q10 crystals provided by the present invention are more stable than reported reduced coenzyme Q10 crystals and exhibit superior physical properties, including water solubility and residual solvent content. Furthermore, the crystals provided by the present invention overcome the shortcomings of previous reduced coenzyme Q10, which has been susceptible to oxidation and limited in use. Furthermore, the reduced coenzyme Q10 crystals provided by the present invention and crystalline solids containing the same exhibit not only excellent stability but also high purity and low residual solvent content. BRIEF DESCRIPTION OF THE DRAWINGS

[0032] FIG1 is a powder X-ray diffraction pattern of reduced coenzyme Q10 crystals obtained in Example 1 of the present invention;

[0033] FIG2 is an infrared absorption spectrum of the reduced coenzyme Q10 crystals obtained in Example 1 of the present invention;

[0034] FIG3 is a DSC spectrum of the reduced coenzyme Q10 crystals obtained in Example 1 of the present invention;

[0035] FIG4 is a powder X-ray diffraction pattern of the reduced coenzyme Q10 crystals obtained in Comparative Example 1 of the present invention;

[0036] FIG5 is an HPLC chart of the reduced coenzyme Q10 crystals obtained in Example 1 of the present invention after storage for 360 days;

[0037] FIG6 is an HPLC chart of reduced coenzyme Q10 crystals obtained in Comparative Example 1 of the present invention after storage for 360 days;

[0038] FIG7 is an HPLC chart of the reduced coenzyme Q10 crystals obtained in Example 1 of the present invention;

[0039] FIG8 is an HPLC chart of the reduced coenzyme Q10 crystals obtained in Comparative Example 1 of the present invention. DETAILED DESCRIPTION

[0040] The following will be combined with the embodiments of the present invention to clearly and completely describe the technical solutions in the embodiments of the present invention. Obviously, the embodiments described are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without making creative efforts are within the scope of protection of the present invention.

[0041] The present invention provides a novel reduced coenzyme Q10 crystal in a crystal form, the physical and chemical properties and crystal structure of which are significantly different from those of reduced coenzyme Q10 crystals reported in the literature.

[0042] Specifically, the present invention provides a reduced coenzyme Q10 crystal, which has an endothermic peak at 52±2°C when heated at a rate of 10k / min as determined by differential scanning calorimetry. More specifically, it has an endothermic peak at 52±2°C indicating crystal melting.

[0043] More specifically, the reduced coenzyme Q10 crystal has a differential scanning calorimetry curve as shown in FIG3 .

[0044] According to the present invention, the reduced coenzyme Q10 crystal preferably shows characteristic peaks at diffraction angles (2θ±0.2°) of 8.95°, 10.04°, 15.09°, 18.65°, 19.03°, 21.61° and 23.01° in powder X-ray diffraction measured using Cu-Kα rays; in particular, it shows characteristic strong diffraction peaks at diffraction angles (2θ±0.2°) of 8.95°, 19.03° and 23.01°.

[0045] More specifically, the reduced coenzyme Q10 crystals have a powder X-ray diffraction pattern measured using Cu-Kα radiation as shown in FIG1 . The characteristic peaks in the XRD diffraction pattern shown in FIG1 are completely different from the diffraction pattern of crystals reported in the prior art (CN103635452A), clearly indicating that the reduced coenzyme Q10 crystals of the present invention shown in FIG1 are a novel polymorph of reduced coenzyme Q10.

[0046] And / or, more specifically, the infrared absorption spectrum of the reduced coenzyme Q10 crystal measured by the KBr pellet method is at a wave number of 794±1cm -1 、877±1cm -1 、962cm -1 and 1014cm -1 There is a characteristic absorption peak at 794±1cm -1 and 877±1cm -1 The peak near 962cm is a characteristic absorption peak formed by two peaks of equal intensity; -1 and 1014cm -1 The peak near the center of the crystalline structure is a characteristic absorption peak formed by two peaks of approximately equal intensity. Since reduced coenzyme Q10 crystals previously reported in the literature do not have two absorption peaks at this location, this clearly indicates that the reduced coenzyme Q10 crystals provided by the present invention are a novel reduced coenzyme Q10 crystalline form that differs from previously reported crystalline forms.

[0047] More specifically, the reduced coenzyme Q10 crystals provided by the present invention have an infrared absorption spectrum measured by the KBr pellet method as shown in FIG2 .

[0048] According to the present invention, the reduced coenzyme Q10 crystals are novel reduced coenzyme Q10 crystals provided by the present invention as long as they contain one or more of the above-mentioned DSC endothermic peaks, XRD diffraction patterns and IR absorption patterns.

[0049] According to the present invention, the reduced Coenzyme Q10 crystals have good water solubility. The solubility of the reduced Coenzyme Q10 crystals in purified water at a temperature of 25°C is preferably at least 0.01 wt%, more preferably at least 0.1 wt%, and even more preferably at least 0.5 wt%. The solubility of the reduced Coenzyme Q10 crystals provided by the present invention is significantly higher than the solubility of previously known reduced Coenzyme Q10 crystals (less than 0.001 wt%). As described above, because the reduced Coenzyme Q10 crystals provided by the present invention exhibit a higher melting point and higher water solubility than reduced Coenzyme Q10 crystals reported in the literature, they are not only a crystalline form with a different crystal structure from the reduced Coenzyme Q10 crystals reported in the literature, i.e., a novel reduced Coenzyme Q10 crystalline form (or crystals containing this crystalline form), but can also be said to be a more stable form. The stable reduced Coenzyme Q10 crystals provided by the present invention not only have high thermal stability but also high water solubility.

[0050] The reduced Coenzyme Q10 crystals provided herein exhibit excellent stability against molecular oxygen. Reduced Coenzyme Q10 is known to be easily oxidized by molecular oxygen in the air. However, as demonstrated in the Examples described below, the novel reduced Coenzyme Q10 crystal form discovered herein and the reduced Coenzyme Q10 crystals containing it as the primary component exhibit greater stability than previously reported crystal forms, even when exposed to air without any protection against molecular oxygen. Furthermore, the high oxidative stability of the reduced Coenzyme Q10 polymorphs of the present invention is demonstrated even in the presence of previously reported reduced Coenzyme Q10 crystals and other amorphous components. This demonstrates an oxidative stability not previously considered to be inherent in the reduced Coenzyme Q10 crystalline solids of the present invention. The oxidative stability of the reduced coenzyme Q10 crystals and crystalline solids of the present invention cannot be generalized due to the content of the novel reduced coenzyme Q10 crystal form in the crystals or crystalline solids, storage conditions, and other factors. For example, the reduced coenzyme Q10 retention rate (%) after storage for a specified period at 25°C in air and protected from light is generally about 80% or higher, preferably about 85% or higher, more preferably about 90% or higher, and particularly preferably 95% or higher. It should be noted that the retention rate referred to herein is the value calculated by the ratio of the absolute content of reduced coenzyme Q10 (or concentration in the crystalline solid) after storage for a specified period to the absolute content of reduced coenzyme Q10 in the composition (or concentration in the crystalline solid) before storage. The specified period is not particularly limited, but is, for example, 6 months, preferably 12 months, and more preferably 36 months.

[0051] The crystalline form of reduced coenzyme Q10 provided by the present invention is more stable than reduced coenzyme Q10 crystals reported in the literature, and has other physical properties, including water solubility and residual solvent, that are also superior. At the same time, the crystalline form provided by the present invention overcomes the shortcomings of previous reduced coenzyme Q10, which is very easy to oxidize and has limitations in use. In addition, the reduced coenzyme Q10 crystals provided by the present invention and the crystalline solid containing the crystals not only have excellent stability but also are outstanding in terms of high purity and low residual solvent.

[0052] The present invention also provides a method for preparing reduced coenzyme Q10 crystals, comprising the following steps: S) crystallizing reduced coenzyme Q10 in the presence of a cyclic ether solvent, a fluorinated alcohol solvent, and an organic base to obtain reduced coenzyme Q10 crystals.

[0053] The present invention has no particular limitation on the sources of all raw materials, and any commercially available raw materials may be used.

[0054] According to the present invention, it is preferred to first mix and dissolve reduced coenzyme Q10 with a cyclic ether solvent under heating conditions, then add a mixed solution of a fluorinated alcohol solvent and an organic base, cool the mixture and crystallize to obtain reduced coenzyme Q10 crystals.

[0055] In the present invention, the reduced coenzyme Q10 can be any reduced coenzyme Q10 known to those skilled in the art without any particular limitation; the purity of the reduced coenzyme Q10 is preferably greater than or equal to 99%, more preferably greater than or equal to 99.5%.

[0056] As the solvent used for crystallization and subsequent processing, since reduced coenzyme Q10 is difficult to dissolve in alcohol solvents, the alcohol reported in the literature for recrystallization is not appropriate. However, as a poor solvent, the crystallization system is easy to slurry after addition, making filtration difficult, and the resulting crystal form is still the crystal form originally reported in the literature. When a single fatty acid ester or ether is used as a solvent, the crystallization yield is low due to the solubility of the sample. The yield can only be improved by reducing the solvent ratio or lowering the temperature. However, this brings another problem. Due to the low temperature or low solvent ratio, the system is relatively viscous and difficult to filter, and the resulting crystal form is still the crystal form reported in the literature. Using alkanes, such as n-hexane, n-heptane, n-propane, n-butane, etc., with the appropriate crystallization temperature and solvent-to-crude product ratio, a high yield of product can be obtained, but the crystal form obtained is confirmed to be the reduced coenzyme Q10 Form I reported in the literature, and there is a situation where the solvent residue remains after drying for 48 hours and it is still difficult to meet the requirements.

[0057] After extensive research, the present invention adopts good cyclic ether solvents, bad fluorinated alcohol solvents and a small amount of organic base. In addition to high yield, the stability of the precipitated crystal is higher than that reported in the literature, and the impurity removal effect is better.

[0058] Reduced coenzyme Q10 and a cyclic ether solvent are mixed and dissolved under heating conditions; the cyclic ether solvent is preferably one or more of 2-methyltetrahydrofuran, 3-methyltetrahydrofuran, tetrahydrofuran and 2,5-dimethyltetrahydrofuran; the ratio of reduced coenzyme Q10 to cyclic ether solvent is preferably 1 g: (1-10) mL, more preferably 1 g: (1-8) mL, more preferably 1 g: (1-5) mL, and most preferably 1 g: (1-3) mL; in some embodiments provided by the present invention, the ratio of reduced coenzyme Q10 to cyclic ether solvent is specifically 1 g: 2 mL or 1 g: 10 mL; the temperature of the mixed dissolution is preferably 30°C to 50°C, more preferably 35°C to 45°C, and more preferably 40°C.

[0059] Then, a mixed solution of a fluorinated alcohol solvent and an organic base is added, and the temperature is lowered for crystallization; the fluorinated alcohol solvent is preferably one or more of trifluoromethanol, trifluoroethanol, trifluoropropanol and trifluorobutanol; the volume ratio of the cyclic ether solvent to the fluorinated alcohol solvent is preferably 1: (1-10), more preferably 1: (1-8), more preferably 1: (1-7), and most preferably 1: (1-5); in some embodiments provided by the present invention, the volume ratio of the cyclic ether solvent to the fluorinated alcohol solvent is specifically 1:5 or 1:1; the mass of the organic base is preferably 0.1% to 10% of the mass of the reduced coenzyme Q10, more preferably 0.5% to 10%, more preferably 1% to 10%, and most preferably 5% to 10%; in some embodiments provided by the present invention, the mass of the organic base is preferably 5%, 7% or 10% of the mass of the reduced coenzyme Q10. %; the crystallization can be room temperature crystallization or cooling crystallization, preferably cooling crystallization or cooling crystallization in combination with other crystallization methods; specifically, the crystallization temperature is preferably 0°C to 20°C; more specifically, in the present invention, the crystallization temperature is preferably 0°C to 10°C or 10°C to 20°C, more preferably 0°C to 5°C or 10°C to 15°C; the crystallization time is preferably more than 6 hours, more preferably 6 to 24 hours; specifically, in the present invention, when the crystallization temperature is 0°C to 10°C, the crystallization time is 6 to 12 hours, and when the crystallization temperature is 10°C to 20°C, the crystallization time is 12 to 24 hours; the crystallization can be carried out under static conditions or under stirring conditions, without special restrictions. In the present invention, it is preferably carried out under stirring conditions; the stirring speed is preferably not less than 200r / min. In the present invention, seed crystals may be added after cooling; the seed crystals are crystals of reduced coenzyme Q10; the present invention has no particular limitation on the crystal form of the reduced coenzyme Q10 crystals used as seed crystals, and the crystals may be the crystals provided by the present invention or the Form I type prepared in existing literature, without any particular limitation; crystallization is induced by adding seed crystals, and the amount of the seed crystals added is not particularly limited. In the present invention, the mass of the seed crystals is preferably 0.1% or more of the mass of the reduced coenzyme Q10, more preferably 0.1% to 5%, more preferably 0.1% to 3%, more preferably 0.5% to 3%, more preferably 1% to 2%, and most preferably 1% to 1.5%.

[0060] Under the above crystallization conditions, in order to improve the solubility, concentration, yield, slurry properties or crystal properties of reduced coenzyme Q10 and other conditions that affect crystallization, the above solvents can be mixed in preferred proportions according to the characteristics of each solvent.

[0061] In a specific embodiment provided herein, the crystallization concentration or the holding time after crystallization can be appropriately determined, taking into account factors such as the solubility of reduced coenzyme Q10 in the solvent, to obtain the desired reduced coenzyme Q10 crystals. For example, when using 2-methyltetrahydrofuran as the solvent, a 50% reduced coenzyme Q10 solution in 2-methyltetrahydrofuran is prepared at a temperature below 40°C, then a fluorinated alcohol and an organic base are added, and the solution is cooled to 10°C for crystallization. After the reduced coenzyme Q10 crystals are precipitated, the precipitated reduced coenzyme Q10 crystals can be directly held in the solvent at this temperature for at least 6 hours, preferably at least 8 hours, and more preferably at least 12 hours. During this holding step, the mixed solution of the precipitated reduced coenzyme Q10 crystals and the solvent can be stirred or left at rest, but is preferably stirred at a rate of not less than 200 r / min.

[0062] After crystallization, the solid can be recovered as needed after solid-liquid separation and drying, using conventionally known methods such as those described in patent literature. For example, solid-liquid separation can be performed using pressure filtration or centrifugal filtration, typically using inert gas filter press. The drying temperature is determined based on the boiling point of the added solvent and is generally not greater than 45°C. The dried crystalline solid can be crushed or classified (sieved) as needed and then recovered to obtain reduced coenzyme Q10 crystals.

[0063] It should be noted that the crystallization and post-treatment steps are preferably carried out in a deoxygenated environment. The deoxygenated environment can be replaced by an inert gas such as nitrogen, helium, argon, and carbon dioxide, preferably nitrogen or argon.

[0064] Furthermore, once reduced coenzyme Q10 crystals are produced or obtained, reduced coenzyme Q10 crystals of the present invention can be produced under normal conditions by adding the reduced coenzyme Q10 crystals of the present invention as seed crystals during crystallization. In this case, either room temperature or cooling crystallization methods can be used. The preferred crystallization method is cooling crystallization, or a combination of cooling crystallization and other crystallization methods.

[0065] The present invention also provides a reduced coenzyme Q10 crystalline solid, comprising the above-mentioned reduced coenzyme Q10 crystal.

[0066] Specifically, the reduced coenzyme Q10 crystalline solid can be obtained by adding reduced coenzyme Q10 of a crystalline form other than the reduced coenzyme Q10 crystal provided in this application as a seed crystal during the crystallization process.

[0067] More specifically, the content of the reduced coenzyme Q10 crystals in the reduced coenzyme Q10 crystalline solid is preferably 0.1 wt% or more, more preferably 1 wt% or more, even more preferably 10 wt% or more, even more preferably 30 wt% or more, even more preferably 50 wt% or more, even more preferably 70 wt% or more, and most preferably 85 wt% or more. When the lower limits of the reduced coenzyme Q10 crystal content provided by the present invention are the above values, the upper limits corresponding to the respective lower limits are naturally 100 wt%. Whether the reduced coenzyme Q10 crystals and crystalline solids provided by the present invention are mixed with the crystalline forms previously reported in the literature, and the ratio thereof, can be determined, for example, by DSC measurement at a heating rate of 1 k / min. Under these conditions, the endothermic peaks indicating the melting of conventionally known reduced coenzyme Q10 crystals and the reduced coenzyme Q10 crystals provided by the present invention are clearly separated. Since the magnitude of these peaks is correlated with the mixing ratio, even when reduced coenzyme Q10 crystals reported in the literature are mixed with the reduced coenzyme Q10 crystals or crystalline solids of the present invention, the presence or content of the novel reduced coenzyme Q10 crystal form can be clearly determined.

[0068] As long as the reduced coenzyme Q10 crystals provided by the present invention contain reduced coenzyme Q10 crystals having the above-mentioned DSC endothermic peak, XRD diffraction pattern, or IR absorption pattern, they can coexist with reduced coenzyme Q10 crystals previously reported in the literature as a crystalline solid. In addition, as long as it is a crystalline solid containing the reduced coenzyme Q10 crystals of the present invention, whether or not it coexists with other solid forms of reduced coenzyme Q10 is within the scope of the present invention. It should be noted that because the reduced coenzyme Q10 crystals contained in the reduced coenzyme Q10 crystalline solid of the present invention are more stable than the crystal forms reported in the literature, even if only a small amount of such reduced coenzyme Q10 crystals are present in the reduced coenzyme Q10 crystalline solid of the present invention, all crystal forms may transition to the reduced coenzyme Q10 crystals provided by the present invention over time.

[0069] The present invention also provides a reduced coenzyme Q10 composition, comprising the above-mentioned reduced coenzyme Q10 crystals and / or the above-mentioned reduced coenzyme Q10 crystalline solid.

[0070] The reduced coenzyme Q10 crystals, reduced coenzyme Q10 crystalline solids, and reduced coenzyme Q10 compositions provided by the present invention can be used in foods, nutritional functional foods, specific health foods, nutritional supplements, animal drugs, beverages, feeds, cosmetics, pharmaceuticals, therapeutic drugs, preventive drugs, or pet medicines and foods.

[0071] To further illustrate the present invention, the following describes in detail a reduced coenzyme Q10 crystal and a preparation method thereof provided by the present invention in conjunction with examples.

[0072] In particular, the measurement conditions of powder X-ray diffraction (XRD), differential scanning calorimetry (DSC) and infrared spectroscopy (IR) analysis in the examples are as follows:

[0073] XRD measurement conditions:

[0074] Equipment model: X-ray powder diffractometer / SmartLab SE;

[0075] X-ray tube: Cu target;

[0076] Speed: 10° / min;

[0077] Collection angle: 3°~60°;

[0078] Step size: 0.02°;

[0079] Slit width: 10mm;

[0080] Output voltage: 40kV;

[0081] Output current: 40mA.

[0082] DSC measurement conditions:

[0083] Equipment model: Differential calorimetry scanner / Netzsch DSC214

[0084] Sample crucible: Concavus Al;

[0085] Heating rate: 10K / min;

[0086] Test range: 30℃~300℃;

[0087] Purge gas / protective gas: N2.

[0088] IR measurement conditions:

[0089] Equipment model: Shimadzu Fourier transform infrared spectrometer IRTracer-100;

[0090] Determination method: KBr tablet method.

[0091] Example 1

[0092] Under nitrogen, 100 g of commercially available reduced coenzyme Q10 (99.5% purity) and 200 mL of 2-methyltetrahydrofuran were added to a 3 L three-necked flask. The mixture was heated to 40°C with stirring until completely dissolved. The temperature was then lowered to 10°C, and 1000 mL of trifluoroethanol and 5 g of triethylamine were slowly added dropwise. After cooling to 2°C, the mixture was kept warm and stirred for 12 hours. The mixture was filtered, washed with 20 mL of trifluoroethanol, and dried under reduced pressure at 40°C for 6-8 hours to obtain granular crystals, which are reduced coenzyme Q10 crystals.

[0093] As a result of DSC analysis, it was confirmed that when the temperature was increased at a rate of 10 k / min, an endothermic peak of melting was observed at 52.2° C. as shown in FIG3 .

[0094] The analysis results of powder X-ray diffraction are shown in Figure 1. Characteristic peaks were found at diffraction angles (2θ±0.2°) of 8.95°, 10.04°, 15.09°, 17.36°, 18.65°, 19.03°, 20.19°, 21.61°, and 23.01°.

[0095] Further IR analysis results, as shown in Figure 2, are different from the reduced coenzyme Q10 crystals in the literature. -1 、877±1cm -1 and 962cm -1 、1014cm -1 There is a characteristic absorption peak near it.

[0096] The above analysis results confirm that the reduced coenzyme Q10 crystals obtained in this example are a different crystal form from the reduced coenzyme Q10 described in the literature. The solubility of the obtained crystals in purified water was measured to be 0.5% by weight at 25°C.

[0097] Example 2

[0098] Under nitrogen protection, 100 g of commercially available reduced coenzyme Q10 (purity 99.5%) and 1000 mL of 2,5-dimethyltetrahydrofuran were added to a 3 L three-necked flask and heated to 40°C with stirring until completely dissolved. 1000 mL of trifluoropropanol and 7 g of triethylamine were slowly added dropwise. After cooling to 10°C, 1 g of seed crystals (obtained in Example 1) was added. After stirring for 1 hour, the temperature was lowered to 2°C, and the mixture was kept warm and stirred for 12 hours. The mixture was filtered, washed with 20 mL of trifluoropropanol, and dried under reduced pressure at 40°C for 6 to 8 hours to obtain crystals.

[0099] As a result of DSC analysis, it was confirmed that each of the samples showed an endothermic melting peak at 50.2° C. when the temperature was increased at a rate of 10 k / min.

[0100] The crystals obtained in Example 2 were analyzed and tested using powder X-ray diffraction and infrared spectroscopy, and the test results were similar to those in Example 1, with the error being within the specified range. It was determined that the crystals obtained in Example 2 had the same crystal form as that of the crystals in Example 1.

[0101] The above analysis results confirm that the reduced coenzyme Q10 crystals obtained in this example are a different crystal form from the reduced coenzyme Q10 described in the literature. The solubility of the obtained crystals in purified water was measured to be 0.5% by weight at 25°C.

[0102] Example 3

[0103] Under nitrogen protection, 100 g of commercially available or homemade reduced coenzyme Q10 and 1000 mL of tetrahydrofuran were added to a 3 L three-necked flask. The mixture was heated to 40°C with stirring until completely dissolved. 1000 mL of trifluoroethanol and 10 g of triethylamine were added dropwise. After cooling to 10°C, 1 g of seed crystals (obtained in Example 1) was added. After stirring for 1 hour, the temperature was lowered to 2°C and the mixture was kept warm and stirred for 12 hours. The mixture was filtered, washed with 20 mL of trifluoroethanol, and dried under reduced pressure at 40°C for 6 to 8 hours to obtain crystals.

[0104] DSC analysis confirmed a melting endothermic peak at 52.2°C when the temperature was raised at a rate of 10 k / min. Furthermore, powder X-ray diffraction analysis confirmed that the reduced coenzyme Q10 crystals obtained in this example were the same crystalline form as the reduced coenzyme Q10 in Example 1.

[0105] Comparative Example 1 (CN103635452A Example 1)

[0106] After nitrogen purge, a 300 mL reaction flask (made of heat-resistant glass) was filled with 40 g of commercially available reduced coenzyme Q10 (manufactured by Kaneka Corporation, conventionally known reduced coenzyme Q10 crystals) and 60 g of n-hexane. The mixture was heated to 40°C while stirring until complete dissolution. The solution was cooled to 25°C at a cooling rate of 10°C / hour and then maintained at 25°C for 96 hours with continued stirring. The solution was then filtered and dried (under reduced pressure, 20°C to 40°C) to obtain crystals.

[0107] As a result of DSC analysis, it was confirmed that each of the samples showed an endothermic melting peak at 50.2° C. when the temperature was increased at a rate of 10 k / min.

[0108] Furthermore, powder X-ray diffraction analysis results, shown in Figure 4, confirm that the reduced coenzyme Q10 crystals obtained in Comparative Example 1 are identical to the reduced coenzyme Q10 Form I reported in the literature, rather than Form II. Experiments also confirmed that using other solvents reported in patents, such as ethanol and heptane, also resulted in Form I crystals as shown in Figure 4. Therefore, conventional solvent crystallization may present difficulties in improving the stability of reduced coenzyme Q10.

[0109] Example 4

[0110] The reduced coenzyme Q10 crystals obtained in Example 1 and Comparative Example 1 were respectively placed in vacuum bags and stored at 25°C in the dark. The weight ratio of reduced coenzyme Q10 to oxidized coenzyme Q10 was determined by HPLC analysis as follows. The results are shown in Table 1. The HPLC chart of the reduced coenzyme Q10 crystals obtained in Example 1 after 360 days of storage is shown in Figure 5, and the HPLC chart of the reduced coenzyme Q10 crystals obtained in Comparative Example 1 after 360 days of storage is shown in Figure 6.

[0111] HPLC analysis conditions

[0112] Chromatographic column: (Agilent) ZORBAX Extend C18 4.6×150mm, 5μm;

[0113] Mobile phase: Mobile phase A: acetonitrile, mobile phase B: isopropanol, mobile phase C: methanol;

[0114] Detection wavelength: 290nm;

[0115] Flow rate: 1.5 mL / min;

[0116] Column temperature: 35°C;

[0117] Injection concentration: 1 mg / mL;

[0118] Injection volume: 20 μL.

[0119] The elution program is shown in Table 2.

[0120] Table 1 Stability test results

[0121] Table 2 Elution program

[0122] As shown in the above results, it is confirmed that the reduced coenzyme Q10 crystals of the present invention have higher stability than the crystal forms in the literature, and higher purity can be obtained by using this crystallization method.

[0123] Comparative Example 3

[0124] The liquid phase method of patent CN103635452 was used for detection, and the results are shown in Figures 7 and 8, where Figure 7 is an HPLC graph of the reduced coenzyme Q10 crystals obtained in Example 1, and Figure 8 is an HPLC graph of the reduced coenzyme Q10 crystals obtained in Comparative Example 1, indicating that the results obtained by the existing method and the literature method are close.

[0125] Example 5

[0126] The reduced coenzyme Q10 crystals obtained in Example 1 and Comparative Example 1 were subjected to solvent residue measurement. The results are shown in Table 3.

[0127] GC analysis conditions

[0128] Chromatographic column: PEG-20M 30m×0.53mm, 1.0μm or equivalent capillary column;

[0129] Heating rate: Start at 35°C, maintain for 10 minutes, then increase to 220°C at a rate of 20°C per minute;

[0130] Inlet temperature: 200°C;

[0131] Detector temperature: 250°C;

[0132] Headspace time: 20 min;

[0133] Headspace temperature: 70°C

[0134] Injection volume: 1 mL.

[0135] Table 3 Solvent residual determination results

[0136] The preferred embodiments of the present invention disclosed above are merely intended to help illustrate the present invention, but the present invention is not limited thereto. Those skilled in the art will appreciate that, within the scope of the technical concept of the present invention, the technical solutions of the present invention may be modified, or some of the technical features may be combined in any other manner. Such modifications or combinations do not deviate from the essence of the corresponding technical solutions from the spirit and scope of the various technical solutions of the present invention, and should be regarded as the contents disclosed by the present invention and fall within the scope of protection of the present invention.

Claims

1. A reduced coenzyme Q10 crystal, characterized in that: The differential scanning calorimetry showed that the product had an endothermic peak at 52±2°C when the temperature was increased at a rate of 10 k / min.

2. The reduced coenzyme Q10 crystal according to claim 1, characterized in that: In powder X-ray diffraction measured using Cu-Kα rays, characteristic peaks were shown at diffraction angles (2θ±0.2°) of 8.95°, 10.04°, 15.09°, 18.65°, 19.03°, 21.61°, and 23.01°.

3. The reduced coenzyme Q10 crystal according to claim 1, characterized in that: The infrared absorption spectrum of the reduced coenzyme Q10 crystal measured by the KBr tablet method is at a wave number of 794±1cm -1 、877±1cm -1 、962cm -1 With 1014cm -1 It has a characteristic absorption peak.

4. The reduced coenzyme Q10 crystal according to claim 1, characterized in that: Having a powder X-ray diffraction pattern measured by Cu-Kα radiation as shown in FIG1; And / or, having an infrared absorption spectrum measured by a KBr tablet method as shown in FIG2 ; And / or, having a differential scanning calorimetry graph as shown in FIG. 3 .

5. A method for preparing reduced coenzyme Q10 crystals, characterized in that: The following steps are involved: S) crystallizing the reduced coenzyme Q10 in the presence of a cyclic ether solvent, a fluorinated alcohol solvent and an organic base to obtain reduced coenzyme Q10 crystals.

6. The preparation method according to claim 5, characterized in that: The step S) is specifically: The reduced coenzyme Q10 and the cyclic ether solvent are mixed and dissolved under heating conditions, and then a mixed solution of a fluorinated alcohol solvent and an organic base is added, and the temperature is lowered for crystallization to obtain reduced coenzyme Q10 crystals; The crystallization temperature is 0°C to 20°C.

7. The preparation method according to claim 5, characterized in that: The cyclic ether solvent is selected from one or more of 2-methyltetrahydrofuran, 3-methyltetrahydrofuran, tetrahydrofuran and 2,5-dimethyltetrahydrofuran; The fluorine-containing alcohol solvent is selected from one or more of trifluoromethanol, trifluoroethanol, trifluoropropanol and trifluorobutanol; The organic base is selected from one or more of triethylamine, pyridine and piperidine.

8. The preparation method according to claim 5, characterized in that: The ratio of the reduced coenzyme Q10 to the cyclic ether solvent is 1 g: (1-10) mL; The volume ratio of the cyclic ether solvent to the fluorinated alcohol solvent is 1:(1-10); The mass of the organic base is 0.1% to 10% of the mass of the reduced coenzyme Q10.

9. A reduced coenzyme Q10 crystalline solid, characterized in that: The invention comprises the reduced coenzyme Q10 crystals according to any one of claims 1 to 4 and / or the reduced coenzyme Q10 crystals prepared by the preparation method according to any one of claims 5 to 8.

10. A reduced coenzyme Q10 composition, characterized in that: The reduced coenzyme Q10 crystal comprising any one of claims 1 to 4; And / or, reduced coenzyme Q10 crystals prepared by any one of the preparation methods of claims 5 to 8; And / or, the reduced coenzyme Q10 crystalline solid according to claim 9.

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