318 FLORIDA STATE HORTICULTURAL SOCIETY, 1964

from the diphenyl pads into open crates, and these in citrusfruits packed in cartons with diphenyl pads.Proc. Fla. State Hort. Soc. 76: 318-20. residues remain effective in reducing decay. 3. Newhall, W. F., E. J. Elvin, and L. R. Knodel. 1954. Infrared determination of biphenyl in citrus fruits. Anal. 5. All residues were substantially within the Chem. 26: 1234. legal residue tolerance for diphenyl. 4. Rygg, G. L., A. W. Wells, Shirley M. Norman, and E. P. Atrops. 1962. Biphenyl control of lemon spoilage. U.S.D.A. Marketing Research Report No. 569. LITERATURE CITED 5. Rygg, G. L., A. W. Wells, Shirley M. Norman, and E. P. Atrops. 1964. Biphenyl control of citrus spoilage. 1. Grierson, W., F. W. Hayward, and M. F. Oberbacher. U.S.D.A. Marketing Research Report No. 646. 1959. Simulated packing,shipping and marketing experi 6 1956. Tolerance of 110 ppm for biphenyl ments with Valencia oranges. Proc. Fla. State Hort. Soc. in oranges, lemons and grapefruit. Fed. Reg. 21(144): 72: 248-54. 5619-20. 2. Hayward, F. W. and G. J. Edwards. 1963. The effects 7. 1960. Tolerance for biphenyl revisedto of time and temperature of storage on residues of diphenyl include all citrus fruits and hybrids. Fed. Reg. 25: 13631-2.

PHENOLIC AMINES IN CITRUS JUICE

Ivan Stewart and T. A. Wheaton of each amine present. The method used for the quantitative determination of these com University of Florida pounds will be given in detail in a subsequent Florida Citrus Experiment Station publication. In many cases, determinations were run on Lake Alfred only one sample of a variety. For this reason, the data given should not be considered as con The authors have recently demonstrated that clusive proof that a particular compound does the phenolic amines, synephrine, octopamine, not occur in different varieties of fruit or at tyramine, and feruloylputrescine are present in different stages of maturity. citrus juices (4, 5, 7). The purpose of this paper is to report the amounts of these compounds pres ent in various juices and to discuss their possible Results and Discussion significance. The synephrine content of several juices was determined by paper chromatography and reported previously (3). Subsequent studies CHOHCH2NHCH3 showed that substantial quantities of synephrine were lost using this technique. A more reliable method based on ion exchange chromatography Synephrine and spectrophotometric measurement was de veloped and used in obtaining the results reported Synephrine was found in the juice of all va here. rieties of fruit analyzed except lemon, limes, grapefruit, and kumquat (Table 1). The pres ence of synephrine in Meyer, sweet, and rough Materials and Methods lemons would suggest that these varieties are Fresh juice, single strength canned juice, or hybrids having at least some parentage other reconstituted frozen concentrate was centrifuged than lemon or lime. at 20,000 X g for 30 minutes.Aliquots were then The largest amount of synephrine was found puton a resin column (Technicon Type A, NH4 + in Dancy tangerine juice which contained from form) and the phenolic amines were eluted with 97 to 152 mg per liter. There was considerably an ammonium hydroxide-water gradient. The less in the juice of Robinson tangerine. Orange effluent from the column was passed through a juice contained 15 to 30 mg per liter with the flow cell in a spectrophotometer and the absorp lowest amount occurring in Navel juice. How tion of ultraviolet light by the separated amines ever, in other analyses not given in this paper, was recorded. The area under each of the re the amounts found in commercial packs of con corded curves was proportional to the amounts centrate were quite uniform. Previously, it was suggested that synephrine might be used as a means of identifying orange Florida Agricultural Experiment Stations Journal Series No. 1975. juice. This seems even more plausible since a STEWART AND WHEATON: PHENOLIC AMINES 319

Table 1. Phenolic amine content of citrus juices.

Kind of 1 NOo of Oc to- Variety Tyra- Feruloyl- Juice Samples Synephrine

mg/1 single-strength juice Hamlin orange S 1 22 5 Parson Brown orange S 1 30 5

Pineapple orange S 1 27 - - 4

Valencia orange CS 2 19 - - 4 Pope orange C 1 25 - - 8

Navel orange F 1 15 _ _ 4

Temple orange CF 2 27-43 T 1 - Murcott orange CF 2 50-52 T T

Dancy tangerine C 5 97-152 1 1 -

Robinson tangerine F 1 58 1 1 -

_ Orlando tangelo F 1 46 —

Duncan grapefruit S 2 - - - 15-22

Marsh grapefruit C 1 - - - 41

Ruby Red grapefruit C 1 - - - 25

Bearss (common) lemon F 1 - - - - Meyer lemon F 1 2 4 25

Sweet lemon F 1 20 - - -

Rough lemon F 1 11 - _

Persian lime F 1 - - -

Key lime F 1 - - - -

Calamondin F 1 9 - - -

Kumqua t F 1 - - - -

Troyer citrange F 1 1 2 7 —

S - Single-strength canned, C - Frozen concentrate, F - Fresh.

2 T - Less than 1 mg/liter of juice.

suitable procedure has been developed for its de termination. This compound is not known to oc cur in plants other than citrus and it seems to CHOHCH2NH2 occur in fairly uniform amounts in orange juice. The pharmacological properties of synephrine are of interest. The drug is used as a stimulant Octopamine and has decongestant properties similar to neo- Octopamine was isolated recently from Meyer synephrine. Synephrine in oral form is recom lemon and is not known to occur in any plants mended both for children and adults (1). The other than citrus (5). In addition to being in minimum dose is 100 mg of the dl tartrate. This Meyer lemon, it is also found in detectable quan is believed to be equivalent to about 17 mg of tities in Temple, Murcott, tangerine, and Troyer the 1 isomer found in citrus. Four to six citrange. Octopamine, which is closely related to ounces of tangerine juice contain this amount of synephrine, was first found in the octopus from synephrine. which its name is derived. More recently, it has 320 FLORIDA STATE HORTICULTURAL SOCIETY, 1964

been found in a number of animals including grapefruit (7) and is also found in lesser humans. It, like synephrine, has interesting amounts in orange varieties. This compound was pharmacological properties; however, because of reported previously by the Russians in Salsola the small amounts which occur in citrus, it is subaphylla, a plant unrelated to citrus (2). Very mainly of academic interest. little is known of its pharmacological properties although it appears to have hypotensive activity.

Summary CH2CH2NH2 The phenolic amines, synephrine, octopamine, tyramine, and feruloylputrescine were determined in citrus juices by separation with an ion ex Tyramine change chromatographic column and spectropho-

Tyramine, unlike synephrine and octopamine, tometric measurement. occurs fairly commonly in plants. It was re Synephrine was present in orange, tangerine, tangelo, calamondin, citrange, and in sweet, ported previously that oranges contained 10 mg/ kg of this compound (6). However, the methods rough, and Meyer lemons but not in Bearss lemon, used for identification probably did not distin limes, grapefruit, or kumquat. Tangerine juice guish between tyramine and synephrine and it is contained 97 to 152 mg/liter of this compound. believed that synephrine was found and erron Octopamine and tyramine were found in eously reported as tyramine. In the present Meyer lemon, Temple, Murcott, citrange, and study, tyramine isolated from Meyer lemon was tangerine. Feruloylputrescine occurred in great positively identified by infrared spectroscopy. est amounts in grapefruit juice and in lesser Meyer lemon juice contains the largest amount of amounts in orange varieties. this compound. Detectable quantities occur in Temple, Murcott, tangerine, and Troyer citrange LITERATURE CITED juice. The absence of tyramine in lemon and lime 1. Personal communication from Dr. A. M. Lands, Sterling Winthrop Research Institute. would make this a useful compound for deter 2. Ryabinin, A. A. and E. M. Il'ina. 1949. The alkaloid mining if Meyer lemon juice had been mixed of Salsola subaphylla. Doklady Akad. Nauk S.S.S.R. 67: 513-516. with that of other varieties. 3. Stewart, Ivan. 1963. An Ephedra alkaloid in citrus juices. Proc. Fla. State Hort. Soc. 76: 242-245. 4. Stewart, Ivan, W. F. Newhall, and G. J. Edwards. CH=CH-CO-NH- 1964. The isolation and identification of 1-synephrine in the leaves and fruit of citrus. Jour. Bio. Chem. 239: 930-932. 5. Stewart, Ivan and T. A. Wheaton. 1964. 1-octopamine in citrus: Isolation and identification. Science 145: 60-61. 6. Udenfriend, S., W. Lovenberg and A. Sjoerdsma. 1959. Physiologically active amines in common fruits and vege tables. Archives Biochem. Biophy. 85: 487-490. 7. Wheaton, T. A. and Ivan Stewart. 1964. Feruloyl OCH3 putrescine: Isolation and identification from the leaves and fruit of citrus. Nature. In press. Feruloylputrescine

Feruloylputrescine is an amide of ferulic acid and putrescine. It was recently isolated from