E. M. Bickoff

Detection of coumestrol in leguminous plants

Abstract A procedure has been described whereby it is possible to qualitatively detect small amounts of a new plant estrogen, coumestrol, in plant material. The method consists of chromatographing an extract of the plant sequentially in two solvent systems on heavy filter paper, then in four solvent systems on silicic acid chromatostrips. The progress of the purification was followed by the blue fluorescence of the compound when the chromatograms were exposed to ultraviolet light. The identity of coumestrol was established by its characteristic fluorescence and R f and by comparison of the spectrophotometric curve with that of pure coumestrol. By this procedure, coumestrol has been detected in seven leguminous plants, and the estimated potency based on this qualitative procedure was compared with the values obtained by mean uterine weight assays.

Trifoliol, a new coumestan from ladino clover

Abstract A new coumestan, C16H10O6, related to coumestrol, has been isolated from ladino clover. Degradation and synthesis of the compound established its structure as 3,7-dihyroxy-9-methoxy-6H-benzofuro[3,2,c][1]-benzopyran-6-one.

Estrogenic activity of coumestrol and related compounds.

Abstract A study was made of the estrogenic activity of 29 compounds related to the plant estrogen, coumestrol. A number of correlations of structure with activity were observed. Acetylation of both hydroxyl groups did not decrease activity. Etherification of both hydroxyl groups decreased estrogenic activity by 2 3 . The 4′-O-methylcoumestrol was less active than the 7-O-methylcoumestrol, and neither was as active as coumestrol. Opening the lactone ring of coumestrol to form the potassium salt of the resulting o-hydroxycinnamic acid did not decrease activity. Formation of the o-methoxycinnamic acid derivative decreased activity greatly. Opening of the furan ring also greatly decreased estrogenic activity. Removal of one of the two hydroxyl groups of coumestrol decreased activity to about 1 6 of the original, and removal of both hydroxyl groups completely eliminated the estrogenic response. The presence of additional hydroxyl groups on the coumestrol molecule greatly diminished estrogenic activity.

Estrogen-like Activity in Vegetable Oils and Mill By-products

By the immature female mouse bioassay technique, an increased uterine weight was observed when certain vegetable oils were fed or injected. Byproducts from the milling of cereals were also capable of eliciting a uterine response.

Bisphenol derivatives as antioxidants for carotene

SummaryRelative values for the carotene-stabilizing effects in mineral oil solutions of a number of bisphenol derivatives were determined. Certain relationships between molecular structure and antioxidant activity were observed. Alkyl substitutions in the reactiveortho andpara positions were shown to improve the stabilizing efficiency of the compounds studied. High activities were obtained with bisphenols linked by methylene or sulfur. Several of the most effective antioxidants were found to be also effective for stabilizing carotene in alfalfa meal.

Effect of antioxidants, individually and in combination, on stability of carotene in cottonseed oil

Summary and ConclusionsBy an accelerated test, l-ascorbyl palmitate, alphatocopherol, hydroquinone, and phospholipids from cottonseed and soybean oil have been evaluated singly and in combinations, for their antioxidant effect on carotene in mineral oil and cottonseed oil solutions. Their effects in retarding the formation of peroxides in the refined cottonseed oil were also determined, and similar studies were made on cottonseed oil after the addition of 1.2 mg. of carotene per gram of oil.In the refined cottonseed oil the formation of peroxides occurred concurrently with the destruction of carotene and was greatly accelerated as the carotene disappeared. In mineral oil solutions, no peroxides were found until after the carotene had become completely decolorized.Although alpha-tocopherol was very effective in stabilizing carotene when added to mineral oil solutions neither the naturally present nor added tocopherol was effective in a more unstable solvent such as refined cottonseed oil. To make the tocopherol effective in such a solvent it is necessary first to stabilize the solvent. When the combination of phospholipid and hydroquinone was added to the refined cottonseed oil, the oil was stabilized and a very marked increase in carotene stability was obtained.While the combinations AH, APH, TAH and TAPH did not stabilize the carotene during the early part of the storage tests, they did become effective after about 20 per cent of the carotene had been destroyed and were as effective as PH and TPH when measured at the 50 per cent point. The reason for this ineffectiveness during the preliminary period of storage is not apparent.

Comparative evaluation of antioxidants for carotene

SummaryA variety of compounds were evaluated as antioxidants for carotene in mineral oil solution by an accelerated stability test. These included derivatives of aromatic amines, substituted dihydroquinolines, ureas, phenothiazine, azobenzene, and diphenylmethane.The most effective antioxidants were hydroxy- and amino-substituted diphenylamines, p-substituted phenylenediamines, and derivatives of 2,2,4-trimethyl-1,2-dihydroquinoline.

Stability of carotene at elevated and room temperatures

SummaryAn attempt has been made to evaluate the elevated temperature test used for carotene stability studies in this laboratory. The stability of carotene in edible oils containing various added antioxidants was determined by storage at 25° C. and in some cases 40° C. and compared with stability as determined by the accelerated test at 75° C. In general, the results obtained at 75° C. were in agreement with those obtained at 40° and 25° C. With most of the better antioxidants the protection found at the lower temperature was better than that indicated by the rapid test.Solutions of carotene in edible oils, which ordinarily might lose a significant amount of carotene in a week or less at 25° C., can be protected by addition of antioxidants, and protection can be afforded for periods well over a year without such special precautions as refrigeration or storage under nitrogen or in evacuated containers. The effectiveness of nordihydroguaiaretic acid as an antioxidant for carotene has been confirmed by room-temperature storage tests.

Stabilization of carotene with nordihydroguaiaretic acid and other antioxidants

SummaryRelative values for the carotene-stabilizing effects in edible oil solutions of a number of antioxidants, alone and in various combinations, have been determined and are presented. One of the more promising for this purpose is nordihydroguaiaretic acid. When this antioxidant is used together with phospholipid or citric acid, its effectiveness is enhanced. Addition of alpha-tocopherol to oils containing only traces of this antioxidant causes a significant increase in carotene stability, and its sparing effect on carotenein vivo (8) would further add to the desirability of its use in such oil solutions of carotene.In refined cottonseed oil, carotene destruction occurred concurrently with the accumulation of peroxides. In the more saturated oils, carotene destruction occurred before much peroxide was detected in the oils.

Identification of 7,4′-dihydroxyflavone in forages

Abstract 7,4′-Dihydroxyflavone, a previously unrecognized constituent of plants, has been found in alfalfa ( Medicago saliva ) and in ladino clover ( Trifolium repens ).