A process for producing ginkgo extracts with a high content of proanthocyanidins and food supplements and medicaments comprising ginkgo extracts

Abstract

The invention relates to a method for producing an extract from Ginkgo biloba leaves, the extract from Ginkgo biloba obtainable by this method, and a medicament or food supplement containing the extract.

Description

[0001] Process for the preparation of ginkgo extracts with a high content of proanthocyanidins and food supplements and medicaments containing ginkgo extracts

[0002] The present invention describes an improved multi-step process for producing an extract from leaves of Ginkgo biloba and its use as a medicament or dietary supplement.

[0003] Extracts from the leaves of Ginkgo biloba have been used medicinally for decades. These medicinal products are currently used to treat various types of dementia and their symptoms, as well as cerebral and peripheral circulatory disorders. The active ingredients are linked to terpene lactones (ginkgolides A, B, C, and bilobalide) and flavonoid glycosides (quercetin, kaempferol, and isorhamnetin).

[0004] Extraction methods relevant for Ginkgo biloba are described in EP 431536 B1 and EP 360556 B1.

[0005] According to claim 1 of EP 431536 B1 / DE 3940092 A1, the process claimed therein for producing an extract from Ginkgo biloba leaves is characterized in that

[0006] (a) fresh or dried green leaves of Ginkgo biloba are extracted with hydrous acetone, a hydrous alkanol with 1 to 3 C atoms or anhydrous methanol at a temperature of about 40 to 100°C,

[0007] (b) the main amount of the organic solvent is separated from the extract to a content of not more than 10% by weight, water being added if necessary during the final distillation stages, (c) the remaining concentrated aqueous solution is diluted with water to a solids content of 5 to 25% by weight, cooled with stirring to a temperature below 25°C, allowed to stand until a precipitate forms and the resulting precipitate, consisting of the lipophilic components which are sparingly soluble in water, is separated,

[0008] (d) the remaining aqueous solution is treated with ammonium sulfate and the resulting solution is extracted with methyl ethyl ketone or a mixture of methyl ethyl ketone and acetone,

[0009] (e) concentrating the resulting extract to a solids content of 50 to 70% and diluting the resulting concentrate with water to a solids content of 5 to 20%,

[0010] (f) the solution thus obtained is extracted in several stages with a water-immiscible butanol or pentanol,

[0011] (g) the butanol or pentanol phases are concentrated to a solids content of 50 to 70%,

[0012] (h) the concentrate is diluted by adding such quantities of water and ethanol that a solution of 5 to 20% by weight of dry extract in 20 to 60% by weight of aqueous ethanol is obtained,

[0013] (i) the aqueous-alcoholic solution is extracted with an aliphatic or cycloaliphatic solvent having a boiling point of 60 to 100°C to further separate the alkylphenol compounds,

[0014] (j) the water phase is concentrated under reduced pressure and dried at a temperature not exceeding 60 to 80°C to give a dry extract with a water content of less than 5%.

[0015] In addition to the compounds mentioned above, Ginkgo biloba leaves also contain the so-called ginkgolic acids (anacardic acids). These compounds are 6-alkylsalicylic acids with n-C13 to n-C19 alkyl radicals with 0 to 3

[0016] Double bonds; cf. J.L. Gellermann et al., Phytochemistry, Vol. 15 (1976), 1959-1961 and Analytic. Chem. Vol. 40 (1968), 739-743. Decarboxylation of ginkgolic acids can produce "ginkgol," a phenol substituted with the corresponding alkyl radical, either biogenetically or during the technical processing of Ginkgo biloba leaves; cf. Kawamura, Japan. J. Chem. Vol. 3 (1928), 91-93.

[0017] The ginkgolic acids and ginkgols in Ginkgo biloba are accompanied by corresponding derivatives with an additional phenolic hydroxyl group in the 4-position, the 6-alkylresorcinol acids and 5-alkylresorcinols, respectively; cf. J. Gellermann et al., Phytochemistry, Vol. 15 (1976), 1959-1961. These resorcinol derivatives are responsible for the toxic effects, particularly the severe allergies and contact dermatitis, induced by plants of the genus Toxicodendron; cf. GA Hill et al., J. Am. Chem. Soc., Vol. 56 (1934), 2736-2738.

[0018] The importance of allergies caused by alkylphenol compounds from Anacardiaceae and Ginkgoaceae is evident from the development of substances and methods for desensitization against allergies caused by alkylphenol compounds, as described in the patent literature (cf. US 4428 965).

[0019] Commercially available extracts from Ginkgo biloba leaves also contain between 50 and 10,000 ppm of ginkgolic acid.

[0020] The extracts from Ginkgo biloba leaves produced by the processes known from DE 17 67 098 and DE 21 17 429 are practically free of alkylphenol compounds because the lipophilic extract components are removed by liquid-liquid extraction of the water-containing acetone extract with a lipophilic solvent that is practically immiscible with water, e.g. a chlorinated lower aliphatic hydrocarbon such as carbon tetrachloride.

[0021] However, the therapeutically valuable ginkgolides and bilobalide are also significantly reduced during this process, so that their content in the final product according to Example 1 of DE 21 17 429 is a maximum of 0.5% for the sum of ginkgolides A, B, C, and J and approximately 0.3% for bilobalide. The sum of the flavonglycosides, however, was significantly enriched by this process, from 3 to 4% in the crude extract to approximately 24% in the final product. The use of substances hazardous to health is naturally undesirable in the production of medicinal products or dietary supplements based on extracts.

[0022] DE 39 40 092 A1, for example, describes a corresponding process in which no lead compounds are used in the removal of undesirable polyphenol compounds with tannin properties (proanthocyanidins). Lead compounds are highly undesirable due to the health risks they pose to the people who work with them and, moreover, incur considerable costs for their proper disposal.

[0023] In recent years, food products containing ginkgo extracts have increasingly appeared on the market. Food products in this context include, in particular, dietary foods, food supplements, and medical foods and dietary supplements.

[0024] In industrialized countries, food has been industrially produced on a large scale for decades and consumed by a large portion of the population every day. It has long been known that industrial production and the associated technical processes such as pasteurization, homogenization, mechanical grinding, separation, heating, or extraction remove a significant portion of the basic components from food. This particularly applies to vitamins in staple foods, which are partially destroyed or rendered ineffective by the technical process, e.g., as a result of heating or other methods. The various vitamins (vitamins B, C, D, A, etc.) are essential for a smooth metabolism.

[0025] Another major problem with today's foodstuffs is the various additives such as preservatives (e.g., benzoic acid), antioxidants, artificial flavorings, sugar, or enzymes. Many of these additives, combined with the technical processing of food, are suspected of being responsible, among other things, for the enormous increase in allergies in recent decades.

[0026] For the reasons listed above, dietary supplements have been offered for years to help supplement today's diets, which are largely based on industrially produced foods, and to improve their deficiencies in essential food components.

[0027] However, many of the dietary supplements available on the market today also contain additives or fillers in the form of milk powder, maltodextrin, starch, gelatin, wheat semolina, whey powder, fructose, glycerin, or sorbitol. Other additives include antioxidants; calcium phosphate, flavorings (usually in the form of fruit flavors), artificial colors, or citrates are also added. The compositions of the various preparations vary only slightly, provided they are vitamin supplements in tablet or powder form.

[0028] These state-of-the-art products are generally mixtures of various vitamins, amino acids, minerals, and / or proteins. What most of these preparations have in common is that they are artificial compounds that non-specifically address possible deficiencies such as vitamin or mineral deficiencies or produce a non-specific improvement in general well-being.

[0029] In addition, body cells are exposed to daily attacks from free radicals – these are aggressive or highly reactive substances with free electrons that arise during metabolic processes or as a result of natural UV or radioactive radiation. If the body's repair mechanisms are no longer fully functional, these radicals can trigger unwanted reactions with genetic material or essential proteins or cause lasting damage, thereby contributing to the development of tumors. This process can be prevented or minimized by administering so-called antioxidants. Among the most effective antioxidants for cells are certain vitamins such as vitamin C, vitamin E, vitamin A, and various B vitamins, as well as the trace element selenium and some phytoflavones.

[0030] The antioxidant and neuroprotective properties of proanthocyanidins from ginkgo leaves and various other sources have been described in the literature (Qu'dan et al. 2011, He et al. 2016, Cao et al. 2016, Xie et al. 2014, Yao et al. 2020). References:

[0031] - Fadi Qa'dan, Kenza Mansoor, Ibraheem AL-Adham, Mathias Schmidt & Adolf Nahrstedt (2011 ) Proanthocyanidins from Ginkgo biloba leaf extract and their radical scavenging activity, Pharmaceutical Biology, 49:5, 471-476, https: / / doi.org 10.3109 / 13880209.2010.523831

[0032] - Q. He, S.-Y. Yang, W. Wang, Z.-J. Wu, H.-L. Ma, Y. Lu; Proanthocyanidins affects the neurotoxicity of Aß25-35 on C57 / bl6 mice; Eur. Rev. Med. Pharmacol. Sci 20 (2016): 679-684

[0033] - Wang-li Cao, Hai-bo Huang, Ling Fang, Jiang-ning Hu, Zhu-ming Jin, Ru-wei Wang; Protective effect of ginkgo proanthocyanidins against cerebral ischemia / reperfusion injury associated with its antioxidant effects; Neural Regeneration Research. 2016; 11 (11 ): 1779-1783, https: / / doi.Org / 10.4103 / 1673-5374.194722

[0034] - Xie, H.; Wang, J.-R.; Yau, L.-F.; Liu, Y.; Liu, L; Han, Q.-B.; Zhao, Z.; Jiang, Z - H. Catechins and Procyanidins of Ginkgo biloba Show Potent Activities towards the Inhibition of ß-Amyloid Peptide Aggregation and Destabilization of Preformed Fibrils. Molecules 2014, 19, 5119-5134. https: / / doi.Org / 10.3390 / molecules19045119

[0035] - Jianbiao Yao, Hongxiang Qiao, Zhuming Jin, RuweiWang, Haibo Huang, Ling Fang, Yan Chen, Khalid Tai, Yan Chen, Thorsten Roland Doeppner, Zhong Chen, Kenny Kuchta; Ginkgo biloba and Its Constituent 6-hydroxykynurenic- acid as well as Its Proanthocyanidins Exert Neurorestorative Effects against Cerebral Ischemia; Plant Med 2020; 86(10): 696-707; https: / / doi.Org / 10.1055 / a-1146-2861

[0036] A combined spectrum of effects that can positively influence deficiencies, general well-being, health and even illnesses is therefore desirable.

[0037] Therefore, an increased content of proanthocyanidins in Ginkgo biloba leaf extracts is desirable to reduce the daily attacks of free radicals.

[0038] The object of the present invention is therefore to develop a process for obtaining a Ginkgo biloba extract that contains an increased content of proanthocyanidins and is suitable for use as a medicament, dietary supplement, or food. Preferably, the extract meets all the requirements of the European Pharmacopoeia and preferably has a high content of proanthocyanidins.

[0039] The object is therefore achieved by a process for producing an extract, preferably a dry extract, from Ginkgo biloba leaves comprising the following steps:

[0040] (a) Extraction of fresh or dried leaves of Ginkgo biloba with hydrous acetone, a hydrous alkanol with 1 to 3 C atoms or anhydrous methanol at a temperature of 40 to 100°C to obtain a crude extract solution,

[0041] (b) separating by distillation the acetone, alkanol with 1 to 3 C atoms or anhydrous methanol from the crude extract solution from step (a) to a content of not more than 10% by weight thereof, based on the total weight of the crude extract solution, in order to obtain a concentrated aqueous solution, wherein, in the case of the use of anhydrous methanol in step (a), water is added one or more times at the end of the distillation,

[0042] (c) diluting the concentrated aqueous solution from step (b) with water to a solids content of 10 to 30 wt.%, preferably 15 to 25 wt.%, cooling to a temperature below 25°C, keeping cool until a precipitate forms and separating the resulting precipitate to again obtain an aqueous solution,

[0043] (d) adding ammonium sulfate to the aqueous solution obtained from step (c) and extracting the resulting ammonium sulfate-containing solution with methyl ethyl ketone / acetone in a ratio of 3 / 2 to 1 / 1 (m / m) and separating the aqueous phase from the methyl ethyl ketone-acetone phase to obtain a methyl ethyl ketone-acetone phase, wherein the extract after step (d) after drying to dry extract has a content of 2.0 wt.% to 5.0 wt.% bilobalide, 1.5 wt.% to 5.0 wt.% ginkgolides A, B and C and 25 - 40 wt.%, preferably 30 - 40 wt.%, proanthocyanidins.

[0044] The abbreviation (m / m) advantageously stands for the respective masses, e.g. 3g methyl ethyl ketone to 2g acetone.

[0045] Preferably, the extraction in step (a) is carried out with hydrous acetone, a hydrous alkanol with 1 to 3 carbon atoms, or anhydrous methanol in a ratio of 5-12 (volume of extractant in liters) to 1 (mass of Ginkgo biloba leaves in kg). The ratio of extractant / drug (volume in liters / mass in kg) is determined by generally weighing the drug (Ginkgo biloba leaves) and adding the solvent for the extraction in liters (via a volumetric meter). Preferably, the extraction can be repeated several times in succession.

[0046] Preferably, the method according to the invention comprises the further steps

[0047] (e) concentrating the methyl ethyl ketone-acetone phase from step (d) to a dry extract content of 40 to 80 wt.% to obtain a concentrate,

[0048] (f) optionally diluting the concentrate obtained from step (e) with water to a solids content of 25 - 50 wt.% to obtain a solution (if the concentrate from step (e) has a dry extract content of 40 - 50 wt.%, ie a solids content in the range of 25 - 50 wt.%, the concentrate from step (e) can be used directly in step (g)).

[0049] (g) at least one-stage extraction of the solution obtained in step (f) with a water-immiscible butanol or pentanol to obtain a butanol or pentanol phase,

[0050] (h) concentrating the butanol or pentanol phase from step (g) to a solids content of at least 50% by weight to obtain a concentrate, (i) diluting the concentrate obtained in step (h) by adding such amounts of water and, if appropriate, ethanol that a solution of 5 to 20% by weight of dry extract in water or at most 60% by weight of aqueous ethanol (equivalent to ‘hydrous ethanol with an ethanol content of 60% by weight’) is obtained,

[0051] (j) extracting the aqueous or aqueous-ethanolic solution from step (i) with an aliphatic solvent having a boiling point of 60 to 100°C, and separating the aqueous phase from the aliphatic solvent phase to obtain an aqueous phase,

[0052] (k) concentrating the aqueous phase obtained in step (j) under reduced pressure and at a temperature of at most 60 to 80°C to obtain a dry extract with a water content of less than 5% by weight.

[0053] Preferably, in step (d) of the process, the volume of methyl ethyl ketone / acetone used is in the range of 0.3 to 1.8 times the volume of the aqueous solution. More preferably, in step (d) of the process, the volume of methyl ethyl ketone / acetone used is in the range of 0.6 to 1.8 times the volume of the aqueous solution, even more preferably in the range of 0.8 to 1.5 times the volume of the aqueous solution.

[0054] Particularly preferably, in step (d) in the process, the volume of methyl ethyl ketone / acetone used is in the range of 0.3 to 0.4; 0.4 to 0.5; 0.5 to 0.6; 0.6 to 0.7; 0.7 to 0.8; 0.8 to 0.9; 0.9 to 1.0; 1.0 to 1.1; 1.1 to 1.2; 1.2 to 1.3; 1.3 to 1.4; 1.4 to 1.5; 1.5 to 1.6 or 1.6 to 1.7 times the volume of the water phase (aqueous solution).

[0055] The extract after step (d) preferably has a content of 10.0 wt.% to 27.0 wt.% flavonoids, calculated as flavonglycosides, 2.0 wt.% to 5.0 wt.% bilobalide, 1.5 wt.% to 5.0 wt.% ginkgolides A, B and C and 30 - 40 wt.% proanthocyanidins. The percentages refer to the dry extract. The extract after step (d) preferably has a content of 10.0 wt.% to 27.0 wt.% flavonoids, calculated as flavonglycosides, 2.1 wt.% to 5.0 wt.% bilobalide, 2.1 wt.% to 5.0 wt.% ginkgolides A, B and C and 30 - 40 wt.% proanthocyanidins. The percentages refer to the dry extract.

[0056] Preferably, the extract after step (d) contains 10.0% to 27.0% by weight of flavonoids, calculated as flavonglycosides, 2.1% to 3.6% by weight of bilobalide, 2.1% to 3.8% by weight of ginkgolides A, B, and C, and 30-40% by weight of proanthocyanidins. The percentages refer to the dry extract.

[0057] Preferably, the extract after step (d) contains 10.0 wt.% to 27.0 wt.% flavonoids, calculated as flavonglycosides, 2.1 wt.% to 3.2 wt.% bilobalide (particularly preferably 2.6 wt.% to 3.2 wt.% bilobalide), 2.1 wt.% to 3.4 wt.% ginkgolides A, B, and C (particularly preferably 2.8 wt.% to 3.4 wt.% ginkgolides A, B, and C), and 30-40 wt.% proanthocyanidins. The percentages refer to the dry extract.

[0058] Preferably, the extract after step (g) contains 20.0 wt.% to 27.0 wt.% flavonoids, calculated as flavonglycosides, 2.6 wt.% to 5.0 wt.% bilobalide (preferably 2.6 wt.% to 3.6 wt.% bilobalide), 3.0 wt.% to 5.0 wt.% ginkgolides A, B and C (preferably 3.0 wt.% to 3.8 wt.% ginkgolides A, B and C) and 5.5-25 wt.%, preferably 5.5-20 wt.%, more preferably 5.5-18 wt.%, most preferably 7-14 wt.%, proanthocyanidins. The percentages refer to the dry extract.

[0059] Preferably, the extract according to step (g) contains 20.0% to 27.0% by weight of flavonoids, calculated as flavonglycosides, 2.6% to 3.2% by weight of bilobalide, 2.8% to 3.4% by weight of ginkgolides A, B, and C, and 5.5-18% by weight, preferably 7-14% by weight, of proanthocyanidins. The percentages refer to the dry extract.

[0060] Preferably, the extract according to step (k) or (j) contains 21.0% to 27.0% by weight of flavonoids, calculated as flavonglycosides, 2.6% to 5.0% by weight of bilobalide, 2.8% to 5.0% by weight of ginkgolides A, B, and C, as well as a maximum of 5 ppm of ginkgolic acids and 5.5-25% by weight, most preferably 6.5-20% by weight, of proanthocyanidins. The percentages refer to the dry extract.

[0061] Preferably, the extract according to step (k) or (j) contains 21.0% to 27.0% by weight of flavonoids, calculated as flavonglycosides, 2.6% to 3.6% by weight of bilobalide (preferably 2.6% to 3.2% by weight of bilobalide), 2.8% to 3.6% by weight of ginkgolides A, B and C (preferably 2.8% to 3.4% by weight of ginkgolides A, B and C) as well as a maximum of 5 ppm of ginkgolic acids and 6.5-20% by weight of proanthocyanidins. The percentages refer to the dry extract.

[0062] The extract after step (k) or (j) preferably has a proanthocyanidin content of 7.0 to 8.0; 8.0 to 9.0; 9.0 to 10.0; 10.0 to 11.0; 11.0 to 12.0; 12.0 to 13.0; 13.0 to 14.0; 14.0 to 15.0, 15.0 to 16.0, 16.0 to 17.0, 17.0 to 18.0 or 18.0 to 19.0 wt.%. The extract after step (k) or (j) particularly preferably has a proanthocyanidin content of 7.0 to 10 wt.%. The percentages refer to the dry extract.

[0063] Preferably, the method according to the invention comprises the further steps

[0064] (e1) Concentration of the methyl ethyl ketone-acetone phase from step (d) to a solids content of at least 40 wt.%, preferably 45 to 95 wt.%, to obtain a concentrate,

[0065] (f1) diluting the concentrate obtained in step (e1) by adding such amounts of water and optionally ethanol that a solution of 5 to 20% by weight of dry extract in water or at most 60% by weight of aqueous ethanol is obtained,

[0066] (g1) Extraction of the aqueous or aqueous-ethanolic solution from step (f1) with an aliphatic solvent having a boiling point of 60 to 100°C and separation of the aqueous phase from the aliphatic solvent phase to obtain an aqueous phase, (hi) Concentration of the aqueous phase obtained in step (g1) under reduced pressure and at a temperature of at most 60 to 80°C to obtain a dry extract with a water content of less than 5 wt.%, wherein the dry extract after step (h1) has a content of 2.0 wt.% to 5.0 wt.% bilobalide (preferably 2.0 wt.% to 3.6 wt.%, more preferably 2.6 wt.% to 3.6 wt.%, particularly preferably 2.6 wt.% to 3.2 wt.% bilobalide), 2.0 wt.% to 5.0 wt.% ginkgolides A, B and C (preferably 2.0 wt.% to 3.6 wt.%, more preferably 2.8 wt.% to 3.6 wt.%, particularly preferably 2.8 wt.% to 3.4 wt.% ginkgolides A, B and C) and at most 5 ppm ginkgolic acids and 25 - 40 wt.-%, preferably 30-40 wt.%, of proanthocyanidins. The content of flavonoids, calculated as flavonglycosides, is preferably 21.0 wt.% to 27.0 wt.%, particularly preferably 22.0 wt.% to 27.0 wt.%.

[0067] Steps (e1) to (h1) are to be understood as alternative process steps to steps (e) to (k).

[0068] A particularly preferred method is for producing an extract, preferably a dry extract, from Ginkgo biloba leaves comprising the following steps:

[0069] (a) Extraction of fresh or dried leaves of Ginkgo biloba with hydrous acetone, a hydrous alkanol with 1 to 3 C atoms or anhydrous methanol at a temperature of 40 to 100°C to obtain a crude extract solution,

[0070] (b) separating by distillation the acetone, alkanol with 1 to 3 C atoms or anhydrous methanol from the crude extract solution from step (a) to a content of not more than 10% by weight thereof, based on the total weight of the crude extract solution, in order to obtain a concentrated aqueous solution, wherein, in the case of the use of anhydrous methanol in step (a), water is added one or more times at the end of the distillation,

[0071] (c) diluting the concentrated aqueous solution from step (b) with water to a solids content of 10 to 30 wt.%, preferably 15 to 25 wt.%, cooling to a temperature below 25°C, keeping cool until a precipitate forms and separating the resulting precipitate to again obtain an aqueous solution,

[0072] (d) adding ammonium sulfate to the aqueous solution obtained from step (c) and extracting the resulting ammonium sulfate-containing solution with methyl ethyl ketone / acetone in a ratio of 3 / 2 to 1 / 1 (w / w) and separating the aqueous phase from the methyl ethyl ketone / acetone phase to obtain a methyl ethyl ketone / acetone phase, wherein the extract preferably after step (d) contains 10.0 wt.% to 27.0 wt.% flavonoids, calculated as flavonglycosides, 2.0 wt.% to 5.0 wt.% bilobalide, 1.5 wt.% to 5.0 wt.% ginkgolides A, B and C, 30-40 wt.% proanthocyanidins, and the percentages refer to the dry extract, e) concentrating the methyl ethyl ketone / acetone phase from step (d) to a dry extract content of 40 to 80 wt.%, to obtain a concentrate,

[0073] (f) optionally diluting the concentrate obtained from step (e) with water to a solids content of 25 - 50 wt.% to obtain a solution,

[0074] (g) at least one-stage extraction of the solution obtained in step (f) with a water-immiscible butanol or pentanol to obtain a butanol or pentanol phase, wherein preferably in step (g) the aqueous solution is extracted in one stage.

[0075] (h) concentrating the butanol or pentanol phase from step (g) to a solids content of at least 50% by weight to obtain a concentrate,

[0076] (i) Diluting the concentrate obtained in step (h) by adding such amounts of water and optionally ethanol that a solution of 5 to 20 wt.% dry extract in water or at most 60 wt.% aqueous ethanol (equivalent to "hydrous ethanol with an ethanol content of 60 wt.%) is obtained, (j) Extracting the aqueous or aqueous-ethanolic solution from step (i) with an aliphatic solvent having a boiling point of 60 to 100°C, and separating the aqueous phase from the aliphatic solvent phase to obtain an aqueous phase,

[0077] (k) Concentration of the aqueous phase obtained in step (j) under reduced pressure and at a temperature of at most 60 to 80°C in order to obtain a dry extract with a water content of less than 5% by weight, wherein the extract after step (k) or (j) has a content of 21.0% by weight to 27.0% by weight of flavonoids, calculated as flavonglycosides, 2.6% by weight to 5.0% by weight of bilobalide, 2.8% by weight to 5.0% by weight of ginkgolides A, B and C and at most 5 ppm of ginkgolic acids and 5.5 - 25% by weight, very particularly preferably 6.5 - 20% by weight, of proanthocyanidins, and wherein the percentages relate to the dry extract.

[0078] Preferably, the extract according to step (d) has a content of 10.0 wt.% to 27.0 wt.% flavonoids, calculated as flavonglycosides, 2.1 wt.% to 3.6 wt.% bilobalide (particularly preferably 2.1 wt.% to 3.2 wt.% bilobalide), 2.1 wt.% to 3.8 wt.% ginkgolides A, B and C (particularly preferably 2.1 wt.% to

[0079] 3.4 wt% ginkgolides A, B and C), 30 - 40 wt% proanthocyanidins, whereby the percentages refer to the dry extract.

[0080] Preferably, the extract according to step (k) or (j) has a content of 21.0 wt.% to 27.0 wt.% flavonoids, calculated as flavonglycosides, 2.6 wt.% to 3.6 wt.% bilobalide, 2.8 wt.% to 3.6 wt.% ginkgolides A, B and C and at most 5 ppm ginkgolic acids and 5.5 - 25 wt.%, very particularly preferably

[0081] 6.5 - 20 wt.% proanthocyanidins, whereby the percentages refer to the dry extract.

[0082] Preferably, the extraction solvent used in step (a) is water-containing acetone having an acetone content of about 50 to 70 wt.%, particularly preferably having an acetone content of about 60 wt.%.

[0083] Preferably, the extraction solvent used in step (a) is a water-containing alkanol selected from the group consisting of methanol, ethanol, 1-propanol, 2-propanol or mixtures thereof, with an alkanol content of 50 wt.% to 70 wt.%, particularly preferably water-containing ethanol with an ethanol content of 60 wt.%.

[0084] Preferably, the aqueous solution diluted in step (c) is cooled to a temperature below 12°C, in particular to 0°C to 11°C (normal physical conditions).

[0085] Preferably, at least 30 wt.% ammonium sulfate, particularly preferably 30 to 50 wt.% ammonium sulfate, based on the aqueous solution from step (c) is added in step (d).

[0086] Preferably, in step (e), the methyl ethyl ketone-acetone phase from step (d) is concentrated to a dry extract content of 45 to 75 wt.% in order to obtain a concentrate.

[0087] Preferably, in step (f), the concentrate obtained from step (e) is optionally diluted with water to a solids content of 30 - 45 wt.% to obtain a solution,

[0088] Preferably, in step (g) extraction is carried out with 1-butanol, preferably once or once to three times.

[0089] Preferably, in step (g), the aqueous solution is extracted in one stage and in a further step (j) extracted with heptane, particularly preferably with n-heptane or a heptane isomer mixture with a proportion of more than 25% by weight of n-heptane.

[0090] After step (c), the content of PACs is preferably 8.5 to 12 wt.%, based on the dry extract.

[0091] After step (g), the content of PACs is preferably 5.5 to 20 wt.%, particularly preferably 5.5 to 18 wt.%, based on the dry extract.

[0092] In addition, in a further embodiment, an extract from Ginkgo biloba is provided, which is obtainable by the process according to the invention.

[0093] For the production of medicaments, the Ginkgo biloba extract according to the invention can be processed in the usual way, e.g., into solutions, coated tablets, or injection preparations. The medicaments of the invention are preferably used for the treatment of peripheral and cerebral arterial circulatory disorders.

[0094] Therefore, in a further embodiment, a medicament or dietary supplement is provided containing the extract according to the invention.

[0095] Preferably, the dietary supplement comprises vital substances selected from the group consisting of vitamins, minerals, trace elements, plant pigments, coenzymes, amino acids, peptides or plant extracts.

[0096] The dietary supplement preferably contains:

[0097] (i) at least one vitamin selected from the group consisting of vitamin A (retinol), vitamin B1 (thiamine), vitamin B2 (riboflavin), vitamin B3 (niacin), vitamin B4 (adenine, choline), vitamin B5 (pantothenic acid), vitamin B6 (pyridoxine), vitamin B7 (biotin), vitamin B8 (adenosine phosphate), vitamin B9 (folic acid), vitamin B10 (para-aminobenzoic acid, mixture of B vitamins), vitamin B11 (folic acid), vitamin B12 (cobalamin), vitamin B13 (orotic acid), vitamin B14 (mixture of vitamins B10 and B11), vitamin B15 (pangamic acid), vitamin B16 (pyridoxine), vitamin B17 (amygdalin), vitamin B22, vitamin B1T (carnitine), vitamin BX (para-aminobenzoic acid), vitamin C (ascorbic acid), vitamin D (Cholecalciferol), Vitamin E (Tocopherol), Vitamin H (Biotin), Vitamin K (Phylloquinin, Menaquinone) Vitamin Q (Ubiquinone, Coenzyme Q10) and their derivatives; and

[0098] (ii) optionally a mineral component comprising at least one component selected from the group consisting of calcium, iron, potassium, copper, magnesium, molybdenum, selenium, silicon, zinc and their salts,

[0099] (iii) optionally an added natural substance or natural substance mixture comprising at least one natural substance or natural substance mixture selected from the group consisting of carotenoids, in particular lutein, lycopene, lipids, in particular medium-chain triglycerides, phospholipids and omega-3 fatty acids and omega-6 fatty acids, resveratrol and curcumin,

[0100] (iv) a plant extract component comprising at least two different plant extracts, each of which is an extract from a plant or plant parts, independently selected from the group consisting of the ginkgo extract according to the invention and additionally ginseng, green tea, Rhodiola rosea, lavender oil (essential oil), saffron, lemon balm, valerian, withania or combinations thereof,

[0101] (v) optionally at least one probiotic selected from the group consisting of Streptococcus thermophilus, Lactobacillus strains (e.g. Lactobacillus plantarum and Lactobacillus helveticus), Bifidobacterium strains (e.g. Bifidobacterium breve strain A1), Bacillus subtilis, Anaerostipes strains, Oscillibacter strains, Lachnospiraceae strains, Akkermansia strains, Christensenellaceae strains, Firmicutes strains, Ruminococcus strains, Clostridium strains (e.g. Clostridium butyricum), Eubacterium strains (e.g. Eubacterium Hallii, Eubacterium Rectale and Eubacterium plexicaudatum), Coprococcus strains, Faecalibacterium strains, Butyrivibrio strains, Actinomycetes strains, Bacteroidetes strains, Proteobacteria strains, Spirochetes strains, Blautia strains (e.g. Blautia hansenii) and Muribaculum strains (e.g. Muribaculum intestinale).

[0102] "Extraction" in the sense of the present invention is understood to mean a separation process in which one or more components are extracted from a mixture of solid, liquid, or gaseous individual substances with the aid of an extractant or mixtures thereof. The extractant used in the process according to the invention is an extraction solvent, for example hydrous acetone, a hydrous alkanol having 1 to 3 C atoms (preferably methanol, ethanol, 1-propanol, 2-propanol, or mixtures thereof), anhydrous methanol, a methyl ethyl ketone-acetone mixture (in step (d) in a ratio of 3 / 2 to 1 / 1 (m / m)), water-immiscible butanol or pentanol, or aliphatic solvents (preferably having a boiling point of 60 to 100°C).

[0103] Water-immiscible butanol includes n-butanol, iso-butanol, and sec-butanol. Water-immiscible pentanol includes pentan-1-ol, pentan-2-ol, pentan-3-ol, 2-methylbutan-1-ol, 2-methylbutan-2-ol, 3-methylbutan-1-ol, 3-methylbutan-2-ol, and 2,2-dimethylpropan-1-ol.

[0104] A preferred aliphatic solvent is heptane, particularly preferably n-heptane or a heptane isomer mixture containing more than 25 wt.% n-heptane.

[0105] "Extract" within the meaning of the present invention refers to the one or more components extracted from a mixture of substances during extraction. The extract is preferably a dry extract.

[0106] "Dried leaves of Ginkgo biloba" are obtained by a process known to those skilled in the art. They are preferably obtained by drying in a drum oven at temperatures up to several hundred °C, preferably at temperatures above 60 °C, and drying times ranging from 10 minutes to several hours. Air drying (without technical aids) is also possible, as is the use of other technical drying processes or combinations of air drying and technical drying.

[0107] According to the current European Pharmacopoeia (Version 11.0; Monograph 04 / 2008: 1827 “Ginkgo dry extract, refined and quantified”), which is binding for all Ginkgo extracts that can be approved as medicinal products within the scope of the Pharmacopoeia, Ginkgo extract contains 22.0% to 27.0% flavonoids, calculated as flavonglycosides, 2.6% to 3.2% bilobalide, 2.8% to 3.4% ginkgolides A, B and C and a maximum of 5 ppm ginkgolic acids. The production is described in the “Notice on the approval and registration of medicinal products (preparation monographs for human medicine) - Ginkgo folium (Ginkgo biloba leaves)” (Federal Gazette 46, (133), 7360-7361 (1994)) as follows: “A dry extract produced from the dried leaves of Ginkgo biloba LINNE with acetone water and subsequent purification steps without the addition of concentrates or isolated ingredients”.

[0108] Ginkgolides A, B, and C, and bilobalide are well-known compounds from the group of terpene lactones. Ginkgolides A, B, and C are hexacyclic diterpenes, and bilobalide is a tetracyclic sesquiterpene.

[0109] The flavonoid content is calculated as the content of flavonglycosides. Flavonglycosides primarily include the glycosides of flavones (also called flavonoids) quercetin, camphor oil, and isorhamnetin, which are well-known compounds.

[0110] For the purposes of the present invention, "plant extract" refers to an extract obtained from a plant or plant parts as a starting material. The plant extract is preferably a dry extract.

[0111] For the purposes of the present invention, "dry extract" refers to the residue obtained after an extraction, after which the extraction agent has been removed. The resulting solid residue is preferably subjected to an additional drying step. Dry extracts are typically characterized by the drug-to-extract ratio (DV). Dry extracts preferably have a water content of at most 5 wt.%, preferably at most 4 wt.%, at most 3 wt.%, at most 2 wt.%, or at most 1 wt.%.

[0112] For the purposes of the present invention, “crude extract solution” is understood to mean the mixture of extract and extraction solvent obtained during an extraction.

[0113] In step (b), "water is added once or several times at the end of the distillation" preferably means the addition of water as soon as the dry extract content of the concentrated extract solution has reached at least 20 wt.%, which can be achieved by adding 25–150 wt.% water once or several times, based on the mass of the concentrated extract solution, and further distillation, which can reduce the methanol content in the concentrated extract solution. "Proanthocyanidins" (PAC) in the sense of the present invention are secondary plant substances from the group of tannins, which, when heated with dilute mineral acids, produce a color reaction, the so-called proanthocyanidin reaction. Proanthocyanidins are preferably polymerization products of flavan-3-ols and flavan-3,4-diols.Common proanthocyanidins include the condensation products of flavan-3,4-diols, which condense via 4^8 or 4^6 bonds to form di-, tri-, and oligomers, and the condensation products of flavan-3-ol units linked via positions 4 and 8. Proanthocyanidins preferably include dimeric proanthocyanidins, oligomeric proanthocyanidins (OPC), and polymeric proanthocyanidins (PPC).

[0114] Flavan-3,4-diol unit Flavan-3-ol unit

[0115] For the purposes of the present invention, “plant or parts of a plant” means a whole plant (e.g. ginkgo, ginseng, green tea) or a part of this plant, for example one or more parts selected from the group consisting of root, stem, leaf, flower, fruit, bud, shoot or seed.

[0116] For the purposes of the present invention, “fresh or dried leaves of Ginkgo biloba” preferably refers to the green leaves as fresh leaves of the tree, and when these are gently dried, they produce the dried leaves.

[0117] For the purposes of the present invention, “Ginkgo” refers to the plant Ginkgo biloba, a tree species belonging to the order Ginkgoales.

[0118] For the purposes of the present invention, “ginkgolides” preferably refers to ginkgolides A, B, and C. The term “hydrated” (e.g., hydrated acetone, hydrated alkanol) for the purposes of the present invention means that a chemical compound, preferably a solvent, contains a minimum amount of water, typically at least 1% by weight, preferably 10-90% by weight, more preferably 20-80% by weight, more preferably 30-80% by weight, more preferably 40-80% by weight, more preferably 50-70% by weight. For example, hydrated acetone preferably has an acetone content of about 50 to 70% by weight, particularly preferably about 60% by weight. For example, hydrated alkanol preferably has an alkanol content of about 50 to 70% by weight, particularly preferably about 60% by weight.

[0119] The term "anhydrous" in the context of the present invention means that a chemical compound, preferably a solvent, contains virtually no water. Commercially available anhydrous methanol that can be used in the present invention contains, for example, a maximum of 0.8 wt.%, 0.2 wt.%, 0.1 wt.%, 0.02 wt.%, 0.01 wt.%, or 0.002 wt.% water.

[0120] “Concentration” in the sense of the present invention means the partial or complete removal of a solvent (extractant) or a solvent mixture from a mixture of substances by means of distillation.

[0121] For the purposes of the present invention, "dry extract content" refers to the non-volatile portion of an extract solution. This is generally expressed in wt.%. The term "solids content" 1 is used synonymously.

[0122] For the purposes of the present invention, "extract solution" refers to a solution of the solvent-soluble components of the fresh or dried leaves of Ginkgo biloba. This term is also used for further processed solutions of the intermediate stages.

[0123] For the purposes of this invention, "mineral component" refers to both macroelements and trace elements. These are essential for the human organism. Macroelements are present at a concentration of at least 50 mg / kg body weight. Trace elements are present at a concentration of less than 50 mg / kg body weight. Examples:

[0124] Overview of analytical methods

[0125] Photometric determination of proanthocyanidins

[0126] The content of proanthocyanidins is determined photometrically using a proanthocyanidin fraction obtained from the Ginkgo biloba extract as a calibration substance using a 1-point calibration.

[0127] 50 mg of extract, accurately weighed, are diluted to 25 mL with a mixture of isopropanol / 3 M HCl [5:1 (v / v)] and dissolved. 5.0 mL of this solution is transferred to a sealable vial and hydrolyzed for 45 min in a water bath at 100°C.

[0128] The cooled sample is filled up to 10 mL with the isopropanol / HCl mixture and measured on the photometer (wavelength: 556 nm; path length: 1 cm).

[0129] A suitable amount of proanthocyanidin fraction is measured analogously.

[0130] The sample weights are adjusted so that the absorbance is between 0.2 and 1.

[0131] HPLC determination of terpene lactones (ginkgolides A, B, C and bilobalide)

[0132] The determination of terpene lactones is carried out in accordance with the method described in Pharm. Eur. 11 for the determination of terpene lactones in Ginkgo-

[0133] Dry extract.

[0134] HPLC determination of flavonoids

[0135] The determination of flavonoids is based on the method described in Pharm. Eur. 11 for the determination of flavonoids in ginkgo dry extract.

[0136] HPLC determination of ginkgolic acids

[0137] The determination of ginkgolic acids is carried out in accordance with the method described in Pharm. Eur. 11 for the determination of ginkgolic acids in ginkgo-

[0138] Dry extract. Corresponding determination methods can be found in "European Pharmacopoeia 11.0" under the heading "Ginkgo dry extract, refined and quantified."

[0139] Optimization of the parameters of process step (c)

[0140] Preparation of the starting solution for optimization of the parameters of process step (c)

[0141] (corresponding to process step (a); see also “Preparation of the starting solution”)

[0142] 600 g of dried and crushed Ginkgo biloba leaves were extracted with 4.5 kg of 60 wt% acetone for 30 min at 60 °C. The plant material was filtered off, extracted again with 4.5 kg of 60 wt% acetone for 30 min at 60 °C, and filtered again. The two resulting extract solutions were combined, homogenized, and then aliquots were used for the individual experiments.

[0143] Reference example 1 : Cold precipitation at TE (dry extract content) = 5 %

[0144] (according to process steps (b) and (c))

[0145] 0.68 kg of the above starting solution was concentrated to 41.7 g and adjusted to 291 g (dry extract content: 5%) with water. The mixture was cooled to 12°C for 1 h. The precipitate was filtered off, and the crude extract solution was evaporated and dried (analysis sample KF-05%).

[0146] Example 2: Cold precipitation at TE = 15%

[0147] (according to process steps (b) and (c))

[0148] 6.78 kg of the above starting solution was concentrated to 412.5 g and diluted with water to 962 g (dry extract content: 15%). The mixture was cooled to 12°C for 1 h. The precipitate was filtered off, and 10 g of the crude extract solution was evaporated and dried (analysis sample KF-15%). Example 3: Cold precipitation at TE = 25%

[0149] (according to process steps (b) and (c))

[0150] 0.68 kg of the above starting solution was concentrated to 37.4 g and adjusted to 58 g (dry extract content: 25%) with water. The mixture was cooled to 12°C for 1 h. The precipitate was filtered off, and the crude extract solution was evaporated and dried (analysis sample KF-25%).

[0151] Analysis results: Process step (c) “Cold precipitation”

[0152] *all percentages in weight percent

[0153] Optimization of the parameters of the process step (d)

[0154] Optimization of the methyl ethyl ketone - acetone ratio

[0155] Preparation of the starting solution

[0156] (according to process steps (a) to (c))

[0157] 600 g of dried and crushed Ginkgo biloba leaves were extracted with 4.5 kg of 60 wt% acetone for 30 min at 60°C. The plant material was filtered off, extracted again with 4.5 kg of 60 wt% acetone for 30 min at 60°C, and filtered off again. The two resulting extract solutions were combined and homogenized. 6.78 kg of the above extract solution was concentrated to 412.5 g and adjusted to 962 g (dry extract content: 15%) with water. The mixture was cooled to 12°C for 1 h. The precipitate was filtered off, and the crude extract solution was aliquoted for further experiments.

[0158] Reference Example 4: MEK : Acetone distribution 0.75 / 1 (m / m)

[0159] (according to process step (d))

[0160] 100 g of filtrate (crude extract solution from "Preparation of the starting solution") were mixed with 30 g of ammonium sulfate, which was dissolved. This solution was extracted twice with 50 mL of methyl ethyl ketone / acetone 0.75 / 1 (w / w) each. The methyl ethyl ketone / acetone phases were combined, homogenized, evaporated, and dried (analysis sample: MEK:acetone 0.75 / 1).

[0161] Example 5: MEK : Acetone distribution 6 / 5 (m / m)

[0162] (according to process step (d))

[0163] 656 g of filtrate were mixed with 197 g of ammonium sulfate, which was dissolved. This solution was extracted twice with 330 mL of methyl ethyl ketone / acetone 6 / 5 (w / w) each. 100 g of the combined and homogenized methyl ethyl ketone / acetone phases were evaporated and dried (analysis sample: MEK:acetone 6 / 5).

[0164] Reference Example 6: MEK : Acetone distribution 2.25 / 1 (m / m)

[0165] (according to process step (d))

[0166] 52 g of filtrate were mixed with 16 g of ammonium sulfate, which was dissolved. This solution was extracted twice with 25 mL of methyl ethyl ketone / acetone 2.25 / 1 (w / w) each. The methyl ethyl ketone / acetone phases were combined, homogenized, evaporated, and dried (analysis sample: MEK:acetone 2.25 / 1). Analysis results:

[0167] Process step (d) "Optimization of the methyl ethyl ketone - acetone ratio"

[0168] Optimization of the volume ratio of the upper phase to the lower phase

[0169] The upper phase is the upper phase formed in the separating funnel (methyl ethyl ketone - acetone phase). The lower phase is the lower phase formed in the separating funnel (aqueous phase).

[0170] Preparation of the starting solution

[0171] (according to process steps (a) to (c))

[0172] The procedure for preparing the starting solution was analogous to the general experimental procedure, but the cold precipitation was carried out at a TE of 25%. This value corresponds to the preferred method.

[0173] Reference Example 7 MEK: Acetone distribution 0.25 times the volume of upper phase to lower phase

[0174] (according to process step (d)) 92 g of filtrate were mixed with 28 g of ammonium sulfate, which was dissolved. This solution was then treated twice with 11.5 mL of methyl ethyl ketone / acetone 6 / 5 (w / w). After shaking, a three-phase mixture formed (analysis sample: MEK:acetone 0.25).

[0175] Example 8 MEK: Acetone distribution 0.5 times the volume of upper phase to lower phase

[0176] (according to process step (d))

[0177] 52 g of filtrate were treated with 16 g of ammonium sulfate, which was dissolved. This solution was extracted twice with 12.5 mL of methyl ethyl ketone / acetone 6 / 5 (w / w) each. The combined and homogenized methyl ethyl ketone / acetone phases were evaporated and dried (analysis sample: MEK:acetone 0.5).

[0178] Reference example 9 MEK: Acetone distribution 2 times the volume of upper phase to lower phase

[0179] (according to process step (d))

[0180] 52 g of filtrate were treated with 16 g of ammonium sulfate, which was dissolved. This solution was extracted twice with 50 mL of methyl ethyl ketone / acetone 6 / 5 (w / w) each. The combined and homogenized methyl ethyl ketone / acetone phases were evaporated and dried (analysis sample: MEK:acetone 2).

[0181] Analysis results:

[0182] Process step ( d ) "Optimization of the volume ratio of the upper phase to

[0183] subphase"

[0184] *all percentages in weight percent

[0185] Optimization of the parameters of the process step (q)

[0186] Production of the initial dry extract

[0187] (according to process steps (a) to (e))

[0188] 1500 g of dried and crushed Ginkgo biloba leaves were extracted with 11.25 kg of 60 wt% acetone for 1 h at 60°C. The plant material was filtered off, extracted again with 11.25 kg of 60 wt% acetone for 1 h at 60°C, and filtered off again. The two resulting extract solutions were combined and homogenized. The starting solution (21.85 kg) was concentrated to 1.2 kg (dry extract content 35%) and adjusted to 2545 g (dry extract content 15%) with water. The mixture was cooled to <12°C for 16 h. The precipitate was filtered off.

[0189] 2402 g of filtrate were mixed with 721 g of ammonium sulfate, which was dissolved. This solution was extracted twice with 2066 g of methyl ethyl ketone / acetone 5.4 / 6 (w / w) each. The methyl ethyl ketone / acetone phases were combined, homogenized, and dried.

[0190] Example 10: 1 -Butanol distribution 3-fold distribution

[0191] (according to process steps (f) and (g))

[0192] 15 g of dry extract was adjusted to a dry extract content of 40% with water. This phase was extracted three times with 42 mL of 1-butanol. The 1-butanol phases were combined and dried (analysis sample: butanol 3x).

[0193] Example 11: 1 -Butanol distribution 1-fold distribution

[0194] (according to process steps (f) and (g))

[0195] 5 g of dry extract was adjusted to a dry extract content of 40% with water. This phase was extracted once with 70 mL of 1-butanol. The 1-butanol phase was dried (analysis sample: 1x butanol).

[0196] Analysis results: Process step ( g ) "1-butanol distribution"

[0197] *all percentages in weight percent

[0198] The process according to the invention is explained and described in more detail using Comparative Example 1 according to EP 431 536 B1 and Example 1 according to the invention.

[0199] In the following examples, the term “heptane” refers to a saturated aliphatic hydrocarbon mixture with a boiling range of 94 - 100 °C and an n-heptane content of more than 25 wt.%.

[0200] Starting solution for Example 1 and Comparative Example 1

[0201] (corresponding to process step (a) of EP 431 536 B1 and the process according to the invention)

[0202] 750 g of dried and crushed Ginkgo biloba leaves were extracted with 5.63 kg of 60 wt% acetone for 1 h at 60 °C. The plant material was filtered off, extracted again with 5.63 kg of 60 wt% acetone for 1 h at 60 °C, and filtered again. The two resulting extract solutions were combined, homogenized, and then divided.

[0203] Comparative example 1 according to EP 431536 B1

[0204] (according to process steps (b) to (k)) Half of the above starting solution (5162 g) was concentrated to 300 g (dry extract content 35%) and adjusted to 716 g (dry extract content 15%) with water. The mixture was cooled to 12 °C for 1 h. The precipitate was filtered off.

[0205] An analytical sample was taken from the solution, concentrated, and dried (analytical sample V 1, sample after step (c)).

[0206] 680 g of filtrate were treated with 204 g of ammonium sulfate, which was dissolved. This solution was extracted twice with 340 mL of methyl ethyl ketone / acetone 0.75 / 1 (w / w) each. The methyl ethyl ketone / acetone phases were combined and homogenized.

[0207] An analytical sample was taken from the solution, concentrated, and dried (analytical sample V 2, sample after step (d)).

[0208] 584 g of the combined methyl ethyl ketone / acetone phase was concentrated and adjusted to a dry extract content of 10% with water. This phase was extracted three times with 55 mL of 1-butanol. The 1-butanol phases were combined and homogenized.

[0209] An analytical sample was taken from the solution, concentrated, and dried (analytical sample V 3, sample after step (g)).

[0210] 96 g of 1-butanol phase was freed from 1-butanol and adjusted with water and ethanol to a dry extract content of 10% and an ethanol content of 30%. This solution was extracted three times with 25 mL of heptane, concentrated, and dried: 5.1 g (analytical sample V 4, sample after step (j), comparable to step (k) in the process according to the invention).

[0211] *all percentages in weight percent

[0212] The content of ginkgolic acids in the analysis sample V 4 was determined to be less than 5 ppm.

[0213] Inventive Example 1

[0214] (according to the process steps (b) to (k) according to the invention)

[0215] The second half of the above starting solution (5161 g) was concentrated to 300 g (dry extract 35%) and diluted with water to 430 g (dry extract 25%). The mixture was cooled to 12 °C for 1 h. The precipitate was filtered off.

[0216] An analytical sample was taken from the solution, concentrated, and dried (analytical sample E 1 , sample after step (c)).

[0217] 393 g of filtrate was diluted to 665 g with water, 200 g of ammonium sulfate was added, and this dissolved. This solution was extracted twice with 330 mL of methyl ethyl ketone / acetone 6 / 5 (w / w) each. The methyl ethyl ketone / acetone phases were combined and homogenized.

[0218] An analytical sample was taken from the solution, concentrated, and dried (analytical sample E 2, sample after step (d)).

[0219] 495 g of the combined methyl ethyl ketone / acetone phase was concentrated and adjusted to a dry extract content of 30% with water. This phase was extracted once with 130 mL of 1-butanol. The 1-butanol phase was homogenized.

[0220] An analytical sample was taken from the solution, concentrated, and dried (analytical sample E 3, sample after step (g)).

[0221] 89 g of 1-butanol phase was freed from 1-butanol and adjusted with water and ethanol to a dry extract content of 10% and an ethanol content of 30%. This solution was extracted three times with 27 mL of heptane, concentrated, and dried: 5.8 g (analytic sample E 4, sample after step (j)).

[0222] *all percentages in weight percent

[0223] The content of ginkgolic acids in the analysis sample E 4 was determined to be less than 5 ppm.

[0224] In the extract according to Example 1 according to the invention (analysis sample E 4), a proanthocyanidin (PAC) content of 8.43 wt.% was determined, while in the extract according to Comparative Example 1 only 5.50 wt.% PAC was found, which corresponds to an enrichment by a factor of 1.53.

[0225] Figure 1 : Inhibition of the production of reactive oxygen species by different groups of ingredients isolated from EGb 761

[0226] Pharmacological studies demonstrate an increased antioxidant potential of extracts with a proanthocyanidin content of 6.5–9.5 wt.% compared to proanthocyanidin-free extracts. Figure 1 shows the antioxidant properties of PACs (proanthocyanidins). These were investigated in a cellular system in comparison to other known constituents of Ginkgo biloba extract (flavones and terpene lactones). For this purpose, the spontaneous production of reactive oxygen species in the rat neuroblastoma cell line RN46A was determined using dihydrofluorescein conversion. The addition of increasing concentrations of Ginkgo biloba constituents led to a concentration-dependent reduction in the fluorescence signal, indicating inhibition of the production of reactive oxygen species. The PAC fraction represented the most active constituent group.

[0227] Figure 2: Inhibition of the production of reactive oxygen species by EGb 761 compared to a PAC-depleted Ginkgo biloba extract

[0228] Figure 2 describes the results of a study of the antioxidant properties of EGb 761 with a PAC content of 8 wt.% compared to a Ginkgo biloba extract (with comparable production and specifications), but with an additional process step for preparative depletion of the PACs (PAC content < 1 wt.%). The antioxidant effect of the PAC-depleted extract was reduced by a factor of 2.6 compared to the PAC-rich EGb 761.

Claims

CLAIMS 1 . A process for the preparation of an extract from Ginkgo biloba leaves comprising the following steps: (a) Extraction of fresh or dried leaves of Ginkgo biloba with hydrous acetone, a hydrous alkanol with 1 to 3 C atoms or anhydrous methanol at a temperature of 40 to 100°C to obtain a crude extract solution, (b) separating by distillation the acetone, alkanol with 1 to 3 C atoms or anhydrous methanol from the crude extract solution from step (a) to a content of not more than 10% by weight thereof, based on the total weight of the crude extract solution, in order to obtain a concentrated aqueous solution, wherein, in the case of the use of anhydrous methanol in step (a), water is added one or more times at the end of the distillation, (c) diluting the concentrated aqueous solution from step (b) with water to a solids content of 10 to 30 wt.%, preferably 15 to 25 wt.%, cooling to a temperature below 25°C, keeping cool until a precipitate forms, and separating the resulting precipitate to obtain an aqueous solution, (d) Adding ammonium sulfate to the aqueous solution obtained from step (c) and extracting the resulting ammonium sulfate-containing solution with methyl ethyl ketone / acetone in a ratio of 3 / 2 to 1 / 1 (m / m) and separating the aqueous phase from the methyl ethyl ketone-acetone phase to obtain a methyl ethyl ketone-acetone phase, wherein the extract after step (d) after drying to dry extract has a content of 2.0 wt.% to 5.0 wt.% bilobalide, 1.5 wt.% to 5.0 wt.% ginkgolides A, B and C and 25 - 40 wt.% proanthocyanidins. Method according to claim 1, comprising the further steps (e) concentrating the methyl ethyl ketone-acetone phase from step (d) to a dry extract content of 40 to 80 wt.% to obtain a concentrate, (f) optionally diluting the concentrate obtained from step (e) with water to a solids content of 25-50 wt.% to obtain a solution, (g) at least one-step extraction of the solution obtained in step (f) with a water-immiscible butanol or pentanol to obtain a butanol or pentanol phase, (h) concentrating the butanol or pentanol phase from step (g) to a solids content of at least 50% by weight to obtain a concentrate, (i) diluting the concentrate obtained in step (h) by adding such amounts of water and, if appropriate, ethanol that a solution of 5 to 20% by weight of dry extract in water or at most 60% by weight of aqueous ethanol is obtained, (j) extracting the aqueous or aqueous-ethanolic solution from step (i) with an aliphatic solvent having a boiling point of 60 to 100°C, and separating the aqueous phase from the aliphatic solvent phase to obtain an aqueous phase, (k) concentrating the aqueous phase obtained in step (j) under reduced pressure and at a temperature of at most 60 to 80°C to obtain a dry extract with a water content of less than 5% by weight.

3. Method according to claim 1, comprising the further steps (e1) Concentration of the methyl ethyl ketone-acetone phase from step (d) to a solids content of at least 40 wt.% to obtain a concentrate, (f1) diluting the concentrate obtained in step (e1) by adding such amounts of water and optionally ethanol that a solution of 5 to 20% by weight of dry extract in water or at most 60% by weight of aqueous ethanol is obtained, (g1) Extraction of the aqueous or aqueous-ethanolic solution from step (f1) with an aliphatic solvent having a boiling point of 60 to 100°C and separation of the aqueous phase from the aliphatic solvent phase to obtain an aqueous phase, (h1) Concentration of the aqueous phase obtained in step (g1) under reduced pressure and at a temperature of at most 60 to 80°C in order to obtain a dry extract with a water content of less than 5% by weight, wherein the dry extract after step (h1) has a content of 2.0% by weight to 5.0% by weight of bilobalide, preferably 2.0% by weight to 3.6% by weight of bilobalide, 2.0% by weight to 5.0% by weight of ginkgolides A, B and C, preferably 2.0% by weight to 3.6% by weight of ginkgolides A, B and C, and at most 5 ppm of ginkgolic acids and 25 - 40% by weight of proanthocyanidins.

4. The process according to any one of claims 1 to 3, wherein in step (d) the volume of methyl ethyl ketone / acetone used is in the range of 0.3 to 1.8 times the volume of the aqueous solution.

5. The process according to claim 2, wherein the extract after step (j) or (k) has a content of 21.0 wt.% to 27.0 wt.% flavonoids, calculated as flavonglycosides, 2.6 wt.% to 5.0 wt.% bilobalide, 2.8 wt.% to 5.0 wt.% ginkgolides A, B and C as well as a maximum of 5 ppm ginkgolic acids and 6.5 - 20 wt.% proanthocyanidins.

6. The method according to claim 5, wherein the extract after step (j) or (k) has a content of 22.0 wt.% to 27.0 wt.% flavonoids, calculated as flavonglycosides, 2.6 wt.% to 3.2 wt.% bilobalide, 2.8 wt.% to 3.4 wt.% ginkgolides A, B and C, and at most 5 ppm ginkgolic acids and 6.5 - 20 wt.% proanthocyanidins.

7. The process according to any one of claims 1 to 6, wherein the extraction solvent used in step (a) is water-containing acetone having an acetone content of about 50 to 70% by weight, particularly preferably having an acetone content of about 60% by weight.

8. The process according to any one of claims 1 to 7, wherein the extraction solvent used in step (a) is a water-containing alkanol selected from the group consisting of methanol, ethanol, 1-propanol, 2-propanol or mixtures thereof, having an alkanol content of 50% by weight to 70% by weight, particularly preferably water-containing ethanol having an ethanol content of 60% by weight.

9. A process according to any one of claims 1 to 8, wherein the aqueous solution diluted in step (c) is cooled to a temperature below 12°C.

10. The process according to claim 1 to 9, wherein in step (d) at least 30% by weight of ammonium sulfate, particularly preferably 30 to 50% by weight of ammonium sulfate, based on the aqueous solution from step (c) is added.

11. Process according to claims 1 to 10, wherein in step (g) extraction is carried out with 1-butanol.

12. The method according to claim 1 to 11, wherein in step (g) the aqueous solution extracted in 1 stage and / or extracted in step (j) with heptane, particularly preferably with n-heptane or a heptane isomer mixture with a proportion of more than 25 wt.% n-heptane.

13. Extract of Ginkgo biloba, obtainable by the process according to any one of claims 1 to 12.

14. A medicinal product or dietary supplement containing an extract according to claim 13.

15. Dietary supplement according to claim 14, characterized in that the dietary supplement comprises vital substances selected from the group consisting of vitamins, minerals, trace elements, plant pigments, coenzymes, amino acids, peptides or plant extracts.

16. Dietary supplement according to claim 15 comprising: (i) at least one vitamin selected from the group consisting of vitamin A (retinol), vitamin B1 (thiamine), vitamin B2 (riboflavin), vitamin B3 (niacin), vitamin B4 (adenine, choline), vitamin B5 (pantothenic acid), vitamin B6 (pyridoxine), vitamin B7 (biotin), vitamin B8 (adenosine phosphate), vitamin B9 (folic acid), vitamin B10 (para-aminobenzoic acid, mixture of B vitamins), vitamin B11 (folic acid), vitamin B12 (cobalamin), vitamin B13 (orotic acid), vitamin B14 (mixture of vitamins B10 and B11), vitamin B15 (pangamic acid), vitamin B16 (pyridoxine), vitamin B17 (amygdalin), vitamin B22, vitamin B1T (carnitine), vitamin BX (para-aminobenzoic acid), vitamin C (ascorbic acid), vitamin D (Cholecalciferol), Vitamin E (Tocopherol), Vitamin H (Biotin), Vitamin K (Phylloquinin, Menaquinone) Vitamin Q (Ubiquinone, Coenzyme Q10) and their derivatives; and (ii) optionally a mineral component comprising at least one component selected from the group consisting of calcium, iron, potassium, copper, magnesium, molybdenum, selenium, silicon, zinc and their salts, (iii) optionally an added natural substance or natural substance mixture comprising at least one natural substance or natural substance mixture selected from the group consisting of carotenoids, in particular lutein, lycopene, lipids, in particular medium-chain triglycerides, Phospholipids and omega-3 and omega-6 fatty acids, Resveratrol and curcumin, (iv) a plant extract component comprising at least two different plant extracts, each of which is an extract from a plant or plant parts independently selected from the group consisting of the ginkgo extract according to claim 13 and additionally ginseng, green tea, Rhodiola rosea, lavender oil (essential oil), saffron, lemon balm, valerian, withania or combinations thereof, (v) optionally at least one probiotic selected from the group consisting of Streptococcus thermophilus, Lactobacillus strains (e.g. Lactobacillus plantarum and Lactobacillus helveticus), Bifidobacterium strains (e.g. Bifidobacterium breve strain A1), Bacillus subtilis, Anaerostipes strains, Oscillibacter strains, Lachnospiraceae strains, Akkermansia strains, Christensenellaceae strains, Firmicutes strains, Ruminococcus strains, Clostridium strains (e.g. Clostridium butyricum), Eubacterium strains (e.g. Eubacterium Hallii, Eubacterium Rectale and Eubacterium plexicaudatum), Coprococcus strains, Faecalibacterium strains, Butyrivibrio strains, Actinomycetes strains, Bacteroidetes strains, Proteobacteria strains, Spirochetes strains, Blautia strains (e.g. Blautia hansenii) and Muribaculum strains (e.g. Muribaculum intestinale).