Patent Application
20230127534
The invention relates to a process for purification of novel mogrosides from Siraitia grosvenorii fruit extract, and their applications in foods, beverage and other consumables.
Luo han guo generally refers to a fruit of Siraitia grosvenorii, a member of the Cucurbitaceae family, which is a plant native to some regions of southern Asia and China. The sweet taste of luo han guo mainly comes from triterpene glycosides generally known as mogrosides or mogrol glycosides. Mogrosides comprise only about 1% of the luo han guo fruit. There are a number of mogrosides identified in luo han guo but generally mogroside V (CAS No: 88901-36-4) has the highest concentration compared to others (Table 1). Mogrol glycosides have the same core molecule—mogrol or oxo-mogrol and differ from each other by number and type of glycosidic residues bonded to mogrol or oxo-mogrol molecules.
Various extraction techniques are used to isolate mogrosides from luo han guo fruits. As a result luo han guo powdered extracts are being prepared which usually contain 30-65% w/w of total mogrosides, and mogroside V content of those materials can vary as much as 18-55%. Such extracts generally cannot be widely used in foods and beverages as they possess undesirable organoleptic properties.
Hence there is a need for compositions comprising novel mogroside molecules with significantly improved organoleptic characteristics allowing their wider usage in foods, beverages and other consumables.
No techniques are currently available for purifying novel mogrosides. Therefore, there is a need for a process of for purification of novel high purity mogrosides from Siraitia grosvenori fruit extract.
The invention relates to a process for preparation of compositions comprising novel mogrosides from Siraitia grosvenori fruit extract.
In one embodiment, the process for purification of novel mogrosides comprises methods described in U.S. Patent Application No. 61/379,729 (filed Sep. 3, 2010), U.S. patent application Ser. No. 13/219,721 (filed Aug. 29, 2011), now patented as U.S. Pat. No. 9,101,162, and U.S. patent application Ser. No. 14/792,594 (filed Jul. 6, 2015), which was published as US Patent Application Publication No. 2015/0305381, which are incorporated by reference herein in their entirety.
In yet another embodiment the process further comprises:
a. providing a mixture of mogrosides;
b. dissolving the mogrosides mixture in solvent to form an initial solution of mogrosides;
c. passing the initial solution through a chromatographic system,
d. separating the fractions comprising novel mogroside compounds 1-15.
The present invention further provides compositions comprising novel mogroside compounds 1-15.
In one embodiment the compositions further comprise another high intensity sweetener selected from the group consisting of stevioside, rebaudioside A, rebaudioside B, rebaudioside C, rebaudioside D, rebaudioside E, rebaudioside F, rebaudioside G, rebaudioside H, rebaudioside I, rebaudioside J, rebaudioside K, rebaudioside L, rebaudioside M, rebaudioside N, rebaudioside O, dulcoside A, dulcoside B, rubusoside, other steviol glycosides occurring in Stevia rebaudiana plant, glycosylated steviol glycosides, biosynthetic steviol glycosides, Luo Han Guo sweetener, siamenoside, mogroside IV, mogroside V, mogroside VI, other mogrosides occurring in Siraitia grosvenorii fruits, monatin and its salts (monatin SS, RR, RS, SR), glycyrrhizic acid and its salts, curculin, thaumatin, monellin, mabinlin, brazzein, hemandulcin, phyllodulcin, glycyphyllin, phloridzin, trilobatin, baiyunoside, osladin, polypodoside A, pterocaryoside A, pterocaryoside B, mukurozioside, phlomisoside I, periandrin I, abrusoside A, cyclocarioside I, and combinations thereof.
The present invention also provides consumables comprising novel mogroside compounds 1-15.
In one embodiment the consumable is selected from the group consisting of food, beverage, pharmaceutical composition, tobacco, nutraceutical, oral hygienic composition, or cosmetic.
Advantages of the present invention will become more apparent from the detailed description given hereinafter. However, it should be understood that the detailed description and specific examples, while indicating preferred embodiments of the invention, are given by way of illustration only, since various changes and modifications within the spirit and scope of the invention will become apparent to those skilled in the art from this detailed description.
The present invention provides a process for purification of novel mogroside compounds 1-15 from Siraitia grosvenorii fruit.
Hereinafter the term “novel mogroside(s)” refers to compound(s) 1-15 (—
The process of purification of novel mogrosides of the present invention comprises methods described in U.S. Patent Application No. 61/379,729 (filed Sep. 3, 2010), U.S. patent application Ser. No. 13/219,721 (filed Aug. 29, 2011), now patented as U.S. Pat. No. 9,101,162, and U.S. patent application Ser. No. 14/792,594 (filed Jul. 6, 2015), which was published as US Patent Application Publication No. 2015/0305381, which are incorporated by reference herein in their entirety.
The process of the present invention may further comprise isolation and purification of novel mogrosides using chromatography system.
In one embodiment of present invention a solution comprising at least one novel mogroside was passed through a HPLC column and fractions comprising novel mogrosides were collected. The HPLC column can be any suitable HPLC analytical or preparative scale column. The fractions may be eluted by adding an appropriate eluent. The eluent can be any suitable solvent or combination of solvents. In one embodiment, the eluent is water and/or acetonitrile. The method may optionally comprise additional steps, such as removal of solvents from the eluted solution to provide a concentrate comprising at least one novel mogroside.
In one embodiment, the following chromatography systems were used for isolation and purification of novel mogrosides. The mogrosides sample from a natural source was subjected to preparative HPLC using a commercial polyamine column, eluting with a mobile phase comprising acetonitrile and water. The mogrosides fraction from this step was further subjected to preparative HPLC using a commercial C18 column, eluting with a mobile phase comprising acetonitrile and water. Individual fractions from this step were further subjected to analytical-preparative HPLC using a commercial C18 column, eluting with a mobile phase comprising acetonitrile and water. The results of isolation and purification are summarized in Table 2. The purity of the isolated mogroside samples was suitable for NMR and HPLC-MS analysis.
In one embodiment, the fractions comprising novel mogrosides were dried to produce compositions comprising at least one novel mogroside.
The compositions comprising at least one novel mogroside can be also produced from new cultivars of Siraitia grosvenorii plant which have increased content of novel mogrosides.
The compositions comprising at least one novel mogroside can be also produced by fermentation of recombinant microbial hosts selected from genera Candida, Cyberlindnera, Kluyveromyces, Meyerozyma, Pischia, Rhodosporidium, Zygosaccharomyces, Saccharomyces, Aspergillus, Hansenula, Humicola, Trichosporon, Brettanomyces, Pachysolen, Yarrowia, Yamadazyma, Schizosaccharomyces, Ashbya, Cyberlindnera, Pichia, Arxula, Xanthophyllomyces or Escherichia. Non-limiting examples of microbial strains include Arthrobacter globiformis, Aspergillus niger, Aspergillus oryzae, Bacillus licheniformis, Bacillus sphaericus, Bacillus subtilis, Brevibacterium linens, Candida utilis, Candida vini, Corynebacterium glutamicum, Cyberlindnera jadinii, Cyberlindnera sp., Debaryomyces hansenii, Fusarium semitectum, Hypomyces anneniacus, Kluyveromyces lactis, Kluyveromyces marxianus, Kocuria rhizophila, Lactobacillus brevis, Lactobacillus casei, Lactobacillus pentoses, Lactobacillus plantarum, Lactobacillus reuteri, Meyerozyma guilliennondii, Microbacterium sp., Micrococcus luteus, Mucor hiemalis, Mucor racemosus, Penicillium roqueforti, Pichia guilliermondii, Pichia jadinii, Pichia pastoris, Pseudomonas jluorescens, Pseudomonas stutzeri, Rhodosporidium sp., Rhodosporidium toruloides, Rhodotorula mucilaginosa, Rhodotorula rubra, Rhodotorula sp., Saccharomyces bayanus, Saccharomyces cerevisiae, Saccharomyces pastorianus, Streptomyces a/bus, Streptomyces coelicolor, Streptomyces griseus, Streptomyces lividans, Torulaspora delbrueckii, Trichosporon laibachii, Trichosporon oleaginosus, Yarrowia lipolytica, Zygosaccharomyces rouxii, Zymomonas mobilis.
The compositions comprising at least one novel mogroside can be also produced by biotransformation using suitable biocatalysts.
The compositions comprising at least one novel mogroside can be also produced by chemical synthesis from precursor compounds.
One embodiment of present invention is a composition comprising at least one novel mogroside.
In some embodiments, at least one novel mogroside imparts sweet taste.
In one embodiment, the present invention is a sweetener composition comprising at least one novel mogroside.
In another embodiment, the present invention is a flavor-enhancing composition comprising at least one novel mogroside, wherein the novel mogroside is present in an amount effective to provide a concentration at or below the threshold flavor recognition level of the novel mogroside when the flavor-enhancing composition is added to a consumable. In a particular embodiment, the novel mogroside is present in an amount effective to provide a concentration below the threshold flavor recognition level of the novel mogroside when the flavor-enhancing composition is added to a consumable. In one embodiment, the novel mogroside is present in an amount effective to provide a concentration at least about 1%, at least about 5%, at least about 10%, at least about 15,% at least about 20% or at least about 25% or more below the threshold flavor recognition level of the novel mogroside when the flavor-enhancing composition is added to a consumable.
In yet another embodiment, the present invention is a sweetness-enhancing composition comprising at least one novel mogroside, wherein the novel mogroside is present in an amount effective to provide a concentration at or below the threshold sweetness recognition level of the novel mogroside when the sweetness-enhancing composition is added to a consumable. In a particular embodiment, the novel mogroside is present in an amount effective to provide a concentration below the threshold sweetness recognition level of the novel mogroside when the sweetness-enhancing composition is added to a consumable. In one embodiment, the novel mogroside is present in an amount effective to provide a concentration at least about 1%, at least about 5%, at least about 10%, at least about 15,% at least about 20% or at least about 25% or more below the threshold sweetness recognition level of the novel mogroside when the sweetness-enhancing composition is added to a consumable.
In yet another embodiment, the present invention is a consumable comprising at least one novel mogroside. Suitable consumables include, but are not limited to, liquid-based or dry consumables, such as, for example, pharmaceutical compositions, edible gel mixes and compositions, dental compositions, foodstuffs, beverages and beverage products.
In a particular embodiment, the present invention is a beverage comprising a at least one novel mogroside. In a particular embodiment, the novel mogroside is present in the beverage at a concentration that is above, at or below the threshold sweetness recognition concentration of the novel mogroside.
In another particular embodiment, the present invention is a beverage product comprising a at least one novel mogroside. In a particular embodiment, the novel mogroside is present in the beverage product at a concentration that is above, at or below the threshold flavor recognition concentration of the novel mogroside.
In another aspect, the present invention is a method of preparing a consumable comprising (i) providing a consumable matrix and (ii) adding at least one novel mogroside to the consumable matrix to provide a consumable. In a particular embodiment, the novel mogroside is present in the consumable in a concentration above, at or below the threshold sweetness recognition of the novel mogroside. In another particular embodiment, the novel mogroside is present in the consumable in a concentration above, at or below the threshold flavor recognition of the novel mogroside.
In a particular embodiment, the present invention is a method of preparing a beverage comprising (i) providing a beverage matrix and (ii) adding at least one novel mogroside to the consumable matrix to provide a beverage. In a particular embodiment, the novel mogroside is present in the consumable in a concentration above, at or below the threshold sweetness recognition of the novel mogroside. In another particular embodiment, the novel mogroside is present in the consumable in a concentration above, at or below the threshold flavor recognition concentration of the novel mogroside.
In another aspect, the present invention is a method of enhancing the sweetness of a consumable comprising (i) providing a consumable comprising at least one sweet ingredient and (ii) adding at least one novel mogroside to the consumable to provide a consumable with enhanced sweetness, wherein the novel mogroside is present in the beverage with enhanced sweetness at a concentration at or below the threshold sweetness recognition concentration of the novel mogroside.
In a particular embodiment, the present invention is a method of enhancing the sweetness of a beverage comprising (i) providing a beverage comprising at least one sweet ingredient and (ii) adding at least one novel mogroside to the beverage to provide a beverage with enhanced sweetness, wherein the novel mogroside is present in the beverage with enhanced sweetness at a concentration below the threshold sweetness recognition concentration of the novel mogroside. In one embodiment, the concentration of the novel mogroside is present in the beverage with enhanced sweetness at a concentration that is at least about 1%, at least about 5%, at least about 10%, at least about 15%, at least about 20%, or at least about 25% or more below the threshold sweetness recognition concentration of the novel mogroside.
In a further aspect, the present invention is a method of enhancing the flavor of a consumable comprising (i) providing a consumable comprising at least one flavor ingredient and (ii) adding at least one novel mogroside to the consumable to provide a consumable with enhanced flavor, wherein the novel mogroside in present in the consumable with enhanced flavor at a concentration at or below the threshold flavor recognition concentration of the novel mogroside.
In a particular embodiment, the present invention is a method of enhancing the flavor of a beverage comprising (i) providing a beverage comprising at least one flavor ingredient and (ii) adding at least one novel mogroside to the beverage to provide a beverage with enhanced flavor, wherein the novel mogroside is present in the beverage with enhanced flavor in a concentration at or below the threshold flavor recognition concentration of the novel mogroside. In one embodiment, the concentration of the novel mogroside is present in the beverage with enhanced sweetness at a concentration that is at least about 1%, at least about 5%, at least about 10%, at least about 15%, at least about 20%, or at least about 25% or more below the threshold flavor recognition concentration of the novel mogroside.
In the above methods, the novel mogroside may be added as such, or in the form of a composition comprising the novel mogroside. When the novel mogroside is provided as a composition, the concentration of the novel mogroside in the composition is effective to provide a concentration above, at or below the threshold flavor or sweetener composition of the novel mogroside, when the composition is added to the consumable, e.g., the food or beverage.
In some embodiments, the compositions of the present invention comprise one or more additional mogrosides, where the additional mogrosides are selected from, but not limited to, the group consisting of Luo han guo extract, by-products of other mogrosides' isolation and purification processes, a commercially available Luo han guo extract, mogroside IIE, mogroside IIB, mogroside III, mogroside IV, mogroside V, 11-oxo-mogroside V, mogroside VI, siamenoside I, grosmomoside I, neomogroside, and other mogrol and oxo-mogrol glycosides occurring in Siraitia grosvenorii fruit and combinations thereof.
In other embodiments, the compositions of the present invention comprise one or more sweeteners or additional sweeteners. In one embodiment, the additional sweetener is a natural sweetener or a synthetic sweetener. In a particular embodiment, the additional sweetener is a high intensity sweetener. In a particular embodiment, the additional sweetener is a mogroside.
In some embodiments, the compositions of the present invention comprise one or more additives. In a particular embodiment, the additive is selected from the group consisting of carbohydrates, polyols, amino acids and their corresponding salts, poly-amino acids and their corresponding salts, sugar acids and their corresponding salts, nucleotides, organic acids, inorganic acids, organic salts including organic acid salts and organic base salts, inorganic salts, bitter compounds, flavorants and flavoring ingredients, astringent compounds, proteins or protein hydrolysates, surfactants, emulsifiers, flavonoids, alcohols, polymers and combinations thereof.
In some embodiments, the compositions of the present invention comprise one or more functional ingredients. In a particular embodiment, the functional ingredient is selected from the group consisting of caffeine, saponins, antioxidants, dietary fiber sources, fatty acids, vitamins, glucosamine, minerals, preservatives, hydration agents, probiotics, prebiotics, weight management agents, osteoporosis management agents, phytoestrogens, long chain primary aliphatic saturated alcohols, phytosterols and combinations thereof.
In a particular embodiment, the present invention is a consumable comprising a novel mogroside and one or more additional mogrosides, sweeteners, additional sweeteners, additives or functional ingredients.
In another particular embodiment, the present invention is a beverage comprising at least one novel mogroside and one or more additional mogrosides, sweeteners, additional sweeteners, additives or functional ingredients.
The compositions comprising novel mogrosides can be used either alone or in combination with at least one other sweetener in consumables including food, beverage, pharmaceutical composition, tobacco, nutraceutical, oral hygienic composition, or cosmetic. The other sweeteners are selected from the group including sucrose, glyceraldehyde, dihydroxyacetone, erythrose, threose, erythrulose, arabinose, lyxose, ribose, xylose, ribulose, xylulose, allose, altrose, allulose, galactose, glucose, gulose, idose, mannose, talose, fructose, psicose, sorbose, tagatose, mannoheptulose, sedoheltulose, octolose, fucose, rhamnose, arabinose, turanose, sialose, inulin, inulooligosaccharides, fructooligosaccharides, high fructose corn syrup (HFCS), maltodextrin, coupling sugar, honey, stevia, rebaudioside A, rebaudioside B, rebaudioside C, rebaudioside D, rebaudioside E, rebaudioside F, rebaudioside G, rebaudioside H, rebaudioside I, rebaudioside J, rebaudioside K, rebaudioside L, rebaudioside M, rebaudioside N, rebaudioside O, dulcoside A, dulcoside B, rubusoside, steviolbioside, stevioside, other steviol glycosides occurring in Stevia rebaudiana plant, biosynthetic steviol glycosides, glycosylated steviol glycosides, glucosylated steviol glycosides (GSGs), mogroside N, mogroside V, mogroside VI, Luo han guo, siamenoside, other mogrosides occurring in Siraitia grosvenorii fruits, monatin and its salts, curculin, glycyrrhizic acid and its salts, thaumatin, monellin, mabinlin, brazzein, hemandulcin, phyllodulcin, glycyphyllin, phloridzin, trilobatin, baiyunoside, osladin, polypodoside A, pterocaryoside A, pterocaryoside B, mukurozioside, phlomisoside I, periandrin I, abrusoside A, and cyclocarioside I, sugar alcohols, sucralose, potassium acesulfame, acesulfame acid and salts thereof, aspartame, alitame, saccharin and salts thereof, neohesperidin dihydrochalcone, cyclamate, cyclamic acid and salts thereof, neotame, advantame, and combinations thereof.
The following examples are provided for illustrating the process and column system of the present invention.
Purification of Novel Mogrosides
Luo Han Guo fruit extract, commercialized by PureCircle Sdn Bhd (Malaysia) containing 50.72% Mogroside V, 8.58% 11-oxo-Mogroside V, 5.67% Siamenoside I, 5.28% Grosmomoside I and about 3% of novel mogrosides was separated by a column system described in U.S. 61/379,729, U.S. Pat. No. 9,101,162 and U.S. Ser. No. 14/792,594 to obtain an enriched composition comprising about 20% of novel mogrosides, 31.1% Mogroside V, 2.8% 11-oxo-Mogroside V, 1.6% Siamenoside I, and 1.1% Grosmomoside I, all percentages being on w/w dried basis. The enriched composition was concentrated to approximately 50% solids content and subjected to preparative HPLC using YMC Polyamine II column (250×20 mm; 5 μm) at ambient temperature, eluting with a mobile phase consisting of acetonitrile and water with 75:25 (vol/vol) ratio (23 mL/min flowrate).
The mogrosides fraction from the preceding step was concentrated to about 7% solids content and was further subjected to preparative HPLC using Zorbax SB-C18 (9.4×50 mm; 5 μm) column, eluting with a mobile phase consisting of acetonitrile and water with 14:86 vol/vol ratio (35 mL/min flowrate). Five fractions were collected (Table 2) and analysed by analytical-preparative HPLC for purity assessment. Fraction 1 contained two peaks corresponding to mogroside compounds 8 and 9, fraction 2 contained two peaks corresponding to mogroside compounds 10 and 11, fraction 3 contained one major peak corresponding to mogroside compound 12, fraction 4 contained at least two peaks corresponding to mogroside compound 13 and 14, and fraction 5 contained one major peak corresponding to mogroside compound 15. Fractions 1, 2 and 4 were further subjected to analytical-preparative separation described below, while fractions 3 and 5 were concentrated, dried and subjected to NMR and HPLCMS analyses.
Fractions 1, 2 and 4 from the preceding step were subjected to analytical-preparative HPLC, eluting with a mobile phase consisting of a mixture of acetonitrile and water with 19:81 (vol/vol) ratio for fraction 1 and 25:75 (vol/vol) ratio for fractions 2 and 4. The column was Poroshell 120 SB-C18 (4.6×150 mm; 2.7 μm), operated at 0.5 mL/min flowrate. The purified fractions containing individual peaks were concentrated, dried and subjected to NMR and HPLCMS analyses.
NMR samples were dissolved in methanol-d4 and analysed by ID and 2D NMR methods. Noteworthy in the NMR analysis is the assignment of oxo- or hydroxy-functional group at Cl 1-for oxo-a typical 13C chemical shift around 190-220 ppm was seen, whereas for hydroxy—a typical 13C chemical shift of around 65-85 ppm was seen. Also noteworthy is the assignment of alpha-glycosidic linkages for some mogroside compounds where a coupling constant of approximately 3.5 Hz is seen (for beta-glycosidic linkages the coupling constant is usually approximately 7-9 Hz). All novel mogroside compounds claimed in this application showed good purity required for structural elucidation. The acquired data was used for structure elucidation.
HPLCMS Analysis of the Novel Mogrosides
HPLCMS were performed on Shimadzu MS 2020 on Cortecs UPLC C18 1.6 μm, 50×2.1 mm column, mobile phase A: 5 mM ammonium formate +0.1% formic acid, B: methanol:acetonitrile (1:1)+5 mM ammonium phosphate buffer +0.1% formic acid, injection volume 0.1-50, detection ELSD (Sedex 85, pressure 4 bar, nebulizer temperature 35° C.), PDA 210-400 nm, gradient 5% B to 100% B in 4 min followed by 2 min 100% B, tlowrate 1 ml/min, scan 100-1600 amu, pos/neg switch. All mogroside compounds claimed in this application showed good purity required for HPLCMS analysis.
Based on NMR data acquired in Example 2 and the data acquired during HPLCMS analysis of novel mogrosides, the structures of novel mogroside compounds 1-15 were proposed.
Novel Mogrosides Organoleptic Characteristics
The organoleptic properties of the enriched composition comprising about 20% (w/w) novel mogrosides (Example 1), the individual novel mogroside compounds, obtained from individual peak fractions of Example 1, were assessed in aqueous solutions isosweet to 5% sucrose. The assessment was conducted by 20 panelists in comparison with 5% isosweet aqueous solutions of commercial Luo Han Guo fruit extract of Example 1, highly purified (96.6% pure) Mogroside V sample (sourced from ChromaDex Inc., USA) and sucrose (5%). The results are summarized in Table 3.
The sensory evaluation results show all novel mogrosides possess significantly better taste profile compared to commercially available mogroside samples. The data for enriched composition (containing about 20% novel mogrosides) also shows that novel mogrosides can improve the overall taste characteristics of mogrosides mixtures comprising less pleasant mogrosides such as Mogroside V.
Consumable Comprising Novel Mogrosides.
Carbonated beverage samples were prepared according to formula presented in Table 4.
The following samples were used as “mogroside composition” in the formula. The enriched composition of Example 1 (comprising about 20% (w/w) novel mogrosides), the individual novel mogroside compounds, obtained from individual peak fractions of Example 1, commercial Luo Han Guo fruit extract of Example 1, highly purified (96.6% pure) Mogroside V sample (from ChromaDex Inc., USA). The sensory assessment of beverage samples was conducted by 20 panelists. The results are summarized in Table 5.
The results showed the beverages prepared using the composition comprising novel mogrosides possessed the best organoleptic characteristics.
Consumable Comprising Novel Mogrosides.
Chocolate samples were papered according to formula in Table 6.
Chocolate liquor, cocoa butter, milk powder, sorbitol, salt, and the “mogroside composition” were kneaded sufficiently, and the mixture was then placed in a refiner to reduce its particle size for 24 hours. Thereafter, the content was transferred into a conche, the lecithin was added, and the composition was kneaded at 50° C. for 48 hours. Then, the content was placed in a shaping apparatus, and solidified.
The following samples were used as “mogroside composition” in the formula of Table 6. Enriched composition comprising about 20% (w/w) novel mogrosides (Example 1), the individual novel mogroside compounds, obtained from individual peak fractions of Example 1, commercial Luo Han Guo fruit extract of Example 1, highly purified (96.6% pure) Mogroside V sample sourced from ChromaDex Inc., (USA). The sensory assessment of chocolate samples was conducted by 20 panelists. The results are summarized in Table 7.
The results showed the chocolate samples prepared using the composition comprising novel mogrosides possessed the best organoleptic characteristics.
While the present invention has been described with reference to particular embodiments, it will be understood that the embodiments are illustrative and that the invention scope is not so limited. Alternative embodiments of the present invention will become apparent to those having ordinary skill in the art to which the present invention pertains. Such alternate embodiments are considered to be encompassed within the spirit and scope of the present invention. Accordingly, the scope of the present invention is described by the appended claims and is supported by the foregoing description.
| Number | Date | Country | |
|---|---|---|---|
| 62419238 | Nov 2016 | US |
| Number | Date | Country | |
|---|---|---|---|
| Parent | 16348444 | May 2019 | US |
| Child | 17987071 | US |
