May 2-7, 1947. R, H, H|GBY_ 2,421,061 METHODS FOR RECOVERY OF Filed March 11. 1944

/ FLAl/ANO/VE GLYCOS/DE PLANT MATER/AL $00205 MA Z'ER/AL _ A6 c/mus PULP

2 FIRM 0R HARD/5N ~45 W/TH LIME a ALKALINE EXTRA CT/ON — AS WITH Na 0/)’ 0/? All/4 0/,’

4 L/QUOR REMOVAL —A6 BY PRL'SS/NG

5 LOQWER pH OF LIQUOR - A6 W/Th' 5c‘!

6 CEYSTALL/ZAT/O/V ‘

7 SEPARATION 0/‘ CRYSTALS

8 REC‘EYSTALL/ZAT/ON — //" DES/RED

//V l/ENTOR> RALPH H. H/GBY

I ‘i. a A T TOENE Y Patented May 27, 1947 2,421,061 UNITED STATES PATENT OFFICE 2,421,061 METHODS FOR RECOVERY OF FLAVANONE GLYCOSIDES Ralph H. Higby, Ontario, Calif., assignor to Cali fornia Fruit Growers Exchange, Los Angeles, Calif., a corporation of California Application March 11, 1944, Serial No. 526,087

4 Claims. (Cl. 260-210) _

This invention relates to a new and useful which the mass is handled through various types process for the recovery of ?avanone glycosides of equipment. from plant source material. This produces a representative source material, An object of this invention is the recovery of indicated at l on the drawing. The pulp is then flavanone glycosides from their source materials, mixed with a su?icient amount of calcium hy e. g., citrus fruits. It involves alkaline extraction droxide to cause a hardening, indicated at 2. of the ?avanone glycosides from their source Since a portion of the calcium hydroxide will be materials. ' neutralized by any acid present, the particular These and further objects and advantages will amounts of calcium hydroxide to be added will appear more fully to those skilled in the art from 10 vary, depending upon the acidity of the mass be a consideration of the invention as set forth in ing treated. Where the preliminary process has the following description and in the appended been such as to include in this pulp a fair amount claims. of the natural acid of the fruit, as in a bulk or The drawing is a ?ow sheet containing an mechanical juicing operation, and particularly illustrative showing of my preferred process. in those cases in which there is a moderately large Convenient sources of certain of the commer proportion of natural acid in the fruit, as in the cially useful ?avanone glycosides are the wastes case of grapefruit or some of the more acidic accumulating in those establishments which are oranges, I ?nd that the amount of lime added engaged in assembling and processing citrus at this point may suitably run as high as ?fteen fruits to recover other valuables, such as essen 20. pounds per ton in order to alkalinize the pulp tial oil, pectin, citric acid, and many of the su?iciently to give the best v‘hardening. Where numerous types of juice products. The two most the peel has been separated with little or no commonly identi?ed ?avanone glycosides occur admixture of juice, and in those cases where in citrus. They are and . the fruit is naturally very low in acid, amounts Eriodictin (also referred to in the literature as 25 as low as ?ve or six pounds per ton will produce or “glucoside”) has also been suitable hardening. Intimate mixing of the lime reported as occurring in citrus. A number of with the peel will promote more rapid harden other ?avanone glycosides have been identi?ed, me’. among which may be mentioned sakuranin, liq In carrying out step 2 of the process. I have uiritin, citronin, and . 30 found it desirable to use a tank of suitable capac My process will be illustrated as applied to ity, as for example 500 gallons and one equipped the extraction of ?avanone glycosides from cit with an agitator. In this tank are placed ap rus, and particularly from the fruit portion proximately 200 gallons of water to which I add thereof. about 10 pounds of slaked lime. This is suffi The fruits are ?rst preferably treated to re 35 cient water to suspend approximately one ton move the juice therefrom, or the juice portion of the chopped pulp, which is agitated therein of the fruit may be removed or separated from for approximately 15 minutes, after which there the peel in its entirety. The juice may be re is added, as indicated at 3, a 25% caustic soda moved by reaming from the fruit or passing the 40 solution until the pH value of the liquor, when whole fruit through a pulping machine or a suit in equilibrium with the peel, remains de?nitely able press. In some commercial processes for the alkaline. The preferred extraction range will canning of grapefruit hearts or sections, the peel vary somewhat from member to member of the is removed by hand from the juice ball. group. One well-known member of the group If the peel is separated in large pieces from 45 namely, naringin, will be extracted to some ex the juice portion, the peel should now preferably tent at the very low alkalinity of pH 7.5, and be disintegrated to such an extent as to be of exhibits a preferred extraction range under com uniformly small particle size so as to permit con mercial operating conditions of about pH 8.4 to venient access by the reagent. I have found that about pH 9.1; while another member of the group, if the peel is passed through a press of the food 50 which from its structural formula appears to be chopper variety in which the face plate has open very much like naringin, namely, hesperidin, can ings of about % inch in diameter, the peel par be extracted satisfactorily at the very much higher ticles will be satisfactory for my purpose. It alkalinity of pH 11.5, and seems to exhibit a pre should be understood that as much of the rag and ferred extraction range under commercial oper interior pulp as may be desired may be included 55 ating conditions of about pH 10.8 to about pH 11.4. with the peel in my ?avanone glycoside recovery This suspension or slurry is preferably agitated process. The particular manner in which the for a further period of time, usually about 30 peel and pulp is comminuted is of signi?cant minutes, in order to give the best extraction and importance only insofar as it may affect the is then subjected to a pressing operation, indi access of reagents to the pulp and the ease with 60 cated at 4, for the purpose of separating the 2,421,081 3 4 liquor containing the ?avanone glycoside from has reported naringin as appearing in Indian the pulp. shaddock, which is apparently a clearly distin The liquor should be adjusted to within a range guished variety of Citrus maxima (Webster’s New of about pH 4 to about pH 5, which I find gives International Dictionary, second edition). good crystallization or precipitation with the Eriodictin (also referred to as eriodictyol gly members of the group generally. Some variation coside) , which is so far only reported as occurring among the members with respect to crystalliza in lemon peel, appears not to have been de?nitely tion is also found. Referring again speci?cally to identi?ed as to structure but may be presumed to hesperidin and naringin, hesperidin will be found have the empirical formula C27Ha201s and the to begin to crystallize slowly when pH 9 is reached 10 structural formula: and to crystallize at a commercially satisfactory rate at pH 6, while naringin will begin to crystal lize at about pH '7 and increases to a moderately Sugar residue-O satisfactory rate at about pH 6. This acidi?ca tion is conveniently and economically effected 15 with hydrochloric acid, and is indicated at 5. After adjustment of the pH, the flavanone HO O glycoside is allowed to crystallize from the liquor. Eriodictin according to this View, is theoretically The crystallization step is indicated generally at capable of being formed by the demethylation of 8. The ?avanone glycoside separation step may 20 hesperidin. be carried out in any desired and known manner, Sakuranin, having the empirical formula and is indicated on the flow sheet at ‘I. If it is CHI-124010, and the probable structural formula: desired to improve the purity of this product, puri?cation may be effected by subsequent re 0 H crystallization steps, which are all well known, 25 H300 —-- OH and are indicated at 8. Hesperidin has the empirical formula C28H34015 H: and can now with little or no doubt be assigned the following structural formula: Gluco (I) on 30 has been isolated from the bark of the Prunus o H 'Jedoensis Matsumura. Sugar residue-0 0 CH: , with the empirical formula C21H22O9, and the apparent structural formula: H: 35 0 H H t H0 #00 Gluco In this formula the sugar residue contains one Hi molecule of rhamnose and one molecule of glu ll cose. Hesperidin is a natural glycoside appar 0 ently occurring in most citrus fruits. While per 40 haps the most proli?c source of hesperidin is the is obtained from the root of the licorice, Glycyr albedo of the citrus fruit, some has been found rhz'za glabra L. var. glandulifera. in the juice portion of the fruit, and traces have Citronin, with the empirical formula C2aHa4O14, also been found in the leaves, twigs, and bark of and the probable structural formula: citrus trees. This flavanone glycoside has been OCH: reported as occurring rather widely in nature. However, the de?nite possibility exists that the Sugar residue-O identi?cation was not sufficiently certain in all cases and that in some of these instances, at least, other but closely related substances were reported as hesperidin. Naringin is a natural glycoside having the em pirical formula C2'1H32O14 and apparently having in which the sugar residue is apparently a the structural formula: rhamno-glucoside, as in hesperidin, and in which there is some question as to whether the methoxyl 0 11 group, —OCH3, is in the 2’ or the 3’ position, is Sugar residue-0 obtained from the peel of lemon ponderosa, Citrus limon. Burm. f. ponderosa Hort. Neohesperidin, isolated from bitter oranges, ap H 60 H 0 parently has the same empirical formula as hesperidin, C2sH34015, having also an identical It apparently differs from hesperidin only in the aglycon and the same sugar groups united there absence of the methoxyl group at the 4' position to, but appearing to have these bound differently, and the shift of the hydroxyl group to this posi the substance possibly having the following tion. One of the most proli?c sources is the peel structural formula: and rag of the grapefruit. It is also reported to . OH occur in relatively large quantities in the leaves. H This glycoside seems so far only to have been re 0 ported as obtained from the grapefruit, which is Gluco-O variously identi?ed as Citrus grandis, Osbeck, and Citrus decumana, Linn. (The Standard Cyclo I pedia of Horticulture, by L. H. Bailey, The Mac Rhamno~0 0 millan Company, 1935) and identi?ed as derived from Citrus mamima (Webster’s New International It is thus seen that the principal commercial Dictionary, second edition). One recent worker sources of the ?avanone glycosides are citrus 2,421,061 5 6 fruits. My method is admirably adapted to ex range in accordance with the disclosures above to traction from these, and can be applied to the effect the desired pH of extraction. However it other sources. While in the above description I is to be understood that any alkali-acting sub have in general referred to different species of stance which is capable of effecting the desired citrus as sources of individual ?avanone glyco pH and which will solubilize the ?avanone gly sides, it is known that more than one of these coside, i. e., form soluble compounds of ‘the ?ava compounds occur in particular fruits; and it is none glycoside, may be used for this purpose. to be expected, as identi?cations of individual Examples of commercially convenient substances members of this group of compounds becomes are potassium hydroxide, ammonium hydroxide, more certain, that more instances will become sodium carbonate, calcium hydroxide, etc. While de?nitely known of the occurrence of more than these alkali-acting substances form compounds one member of the group in individual species of with ?avanone glycosides which are often re citrus. Where two or more members do occur ferred to as salts, they are more precisely called together, they probably are in my process pre phenolates. v cipitated together as complexes. When this oc I have found that ammonium‘ hydroxide is curs, it seems likely that at least one of the very convenient for use in the extraction since ' ?avanone glycosides will be precipitated in the after the extraction has taken place, the alkalin chalcone form. . ity of the material may be reduced by merely sub Several methods have been heretofore known jecting the solution to aeration or to reduced for the extraction of ?avanone glycosides from pressure to remove ammonia. This variation of their source materials. One almost universally the process is not only convenient, to employ, but reported in the literature depends upon the solu is also economical in that considerably less acid bility of the ?avanone glycosides in some organic is required for the subsequent neutralization solvent. While this method is satisfactory for which effects the precipitation of the ?avanone laboratory procedures, it is ordinarily found pro glycoside. hibitively expensive for use in commercial operat Performance of the extraction step with alkali ing conditions. Another method which can be acting substances which form soluble pectic com used with those members of the group which have pounds without having ?rst performed the firm considerably greater solubility in hot water than ing or hardening step hinders crystallization and in cold, consists essentially of boiling up the 30 will result in the recovery of pectic material with source material with water, followed by straining the crude ?avanone glycoside. and. cooling. Manifestly this method is capable While I have mentioned hydrochloric acid as of introducing considerable quantities of impuri being satisfactory for use in adjusting the pH of ties with the recovered products, and indeed the the liquor to within the range of about pH 4 to ‘ other method mentioned immediately above will about pH 5 for purposes of crystallization, many also introduce various impurities which may be mineral or organic acids may be used for this ad soluble in the particular solvent employed, unless justment, for example, sulphuric acid, nitric acid, special precautionary steps are taken to prevent citric acid, acetic acid, carbonic acid, etc. Where this result, which will naturally further increase the extraction, at step 3, has been performed with the cost of the operation. 40 such a reagent or under such conditions that , My method, in contradistinction, is very eco there is present in the liquor removed, at step 4, nomical to operate and produces a very satisfac any appreciable quantity of alkaline earth, the tory yield commercially and a product which is acidifying step 5, will ordinarily be performed relatively free of impurities. with acid which will yield a soluble alkaline earth I prefer, as an ordinary commercial embodi salt, so as not to cause at this stage an alkaline ment, to use lime, i. e., calcium hydroxide, for the earth precipitate which would introduce a ?ltra hardening step, for reasons of convenience and tion step or a recrystallization step into the pro economy. But it should be noted that any alka cedure. Where ammonia has been used as the line earth compound which will give su?icient extraction agent, heat or aeration will go far cations in the aqueous medium will function to towards completing the lowering of the pH of harden the pulp satisfactorily. Barium hydroxide the liquor, indicated at 5. and strontium hydroxide are chemically effective. While the desired pH range for crystallization Whether they will desirably be utilized will de has been indicated as lying Within the range of pend upon the subsequent use, if any, to which about pH 4 to about pH 5, the variation among it is intended to put the residual pulp. members of the group referred to above will nec While magnesium acts in some respects as an essarily be borne in mind. This acidi?cation alkaline earth, it has been forcefully argued that should not, of course, be carried on to such a it should not be considered as belonging to the point as to cause destruction or breakdown of the alkaline earth group. Notwithstanding, I have ?avanone glycoside, and for this purpose I prefer found that, for example, Mg(OH)2 or MgCOs til) not to go below about pH 2.5. ' may ?rst be used to harden the peel, and the Furthermore, in the above examples, I have solubilization and extraction of the ?avanone mentioned the use of only one extraction of the glycoside may then be effected by adding any ap source material. It is obvious, however, that re propriate alkaline reagent to obtain the desired peated extractions may be carried out on the pH of extraction. It therefore follows that for iii) source material in a manner well known to those the purpose of this process, magnesium may be skilled in the art. Furthermore I have found that considered as an alkaline earth at the hardening additional amounts of ?avanone glycoside may step, but it is not preferred for use in the extrac be recovered by washing the extracted source ma tion step. While it is possible with Mg(OH)z terial with water after the ?rst extraction and to alkalinize into the lower portion of the oper then using this wash water to make up sub ative extraction range, uneconomical quantities sequent batches of fresh peel for extraction pur of reagent are required. poses. In step 3 of the above description, as an ex Having thus described my invention in such ample, I have given the use of caustic soda for full, clear, concise, and exact language as to en the purpose of adjusting the pH to a suitable able others skilled in the art to make and use 2,421,061 7 8 the same, I claim as my invention and desire to 3. A method of recovering naturally occurring secure by Letters Patent the following: ?avanone glycoside from plant materials, which 1. A method for the recovery of ?avanone comprises treating the plant material with water glycosides which comprises the steps of adding to and su?iclent of an alkali compound to adjust plant material containing ?avanone glycosides the pH of the mixture to above 7.5 to solubilize lime in amount su?icient to coagulate slimy com the flavanone glycoside, removing liquor contain ponents of the plant material and adjusting the ing the solubilized ?avanone glycoside, acidifying alkalinity of an aqueous suspension of said plant the liquor to below pH6 to precipitate the ?ava material to a point suillciently alkaline to result none glycoside and separating the precipitated in the formation of soluble compounds of ?ava 10 material from the liquor. ‘ none glycosides, allowing the mixture to stand 4. A method of recovering naturally occurring for a time suiilcient to solubilize and extract flavanone glycoside from plant materials, which ?avanone glycoside, recovering a liquor containing comprises treating the plant material with water solubilized ?avanone glycoside from said mate and suf?cient of an alkaline ammonia compound rial, adjusting the pH of the liquor downward to to adjust the pH of the mixture to above 7.5 to promote crystallization of ?avanone glycoside, al solubilize the flavanone glycoside, removing liquor lowing the ?avanone glycoside to crystallize from containing the solubilized ?avanone glycoside, re the liquor and recovering crystallized flavanone ducing the pH of the liquor to below pH6 to glycoside therefrom. precipitate the flavanone glycoside, and separat 2. A method of recovering ?avanone glycosides 20 ing the precipitated material from the liquor. from naturally occurring source materials, which comprises treating the source materials to harden RALPH H. HIGBY. slimy components thereof, alkalinizing an aque ous suspension of the material to above pH '7 .5 to REFERENCES CITED solubilize the ?avanone glycoside, removing from 25 The following references are of record in the the material liquor containing solubilized flava file of this patent: none glycoside, acidifying the liquor to below pH6 to precipitate the ?avanone glycoside, and UNITED STATES PATENTS removing the precipitated material from the solu Number Name Date tion. 30 2,348,215 Higby ______-_ May 9, 1944