Since the variation vras considered smaJJ., subsequent values for 

 cholesterol in salmon eggs were detemdned in duplicate by the colori- 

 metric procedure. Due to the higher cholesterol content of salmon 

 eggs as compared to the samples of egg white contaminated with small 

 amounts of egg yolk for which the colorimetric method was originally 

 adapted, it was not necessary to use moi^ than two or three grams of 

 ground salmon eggs for each analysis. With this small sample it was 

 possible to saponify the eggs directly by the addition of 30 milli- 

 liters of 95 percent ethanol and 3 milliliters of 50 percent KOH fol- 

 lowed by a period of refluxing on a steam bath for 30 minutes. The 

 combined ether extractions of the unsaponifiable fraction were washed 

 with distilled water untU. the washings were neutral to phenolphtha- 

 lein. The extract was then made up to a volume of 100 milliliters 

 with ethyl ether. Five milliliter aliquots were placed in dry test 

 tubee for color development j the ether was raaoved by immersion in 

 a water bath maintained at 60°C. and five milliliters of C.P. chloro- 

 form were added when the ether had evaporated. The color was develop- 

 ed at 1S°C. for 25 minutes in accordance with the Cook-Mehlenbacher 

 technique using acetic anhydride-suLfuidc, acid mix. However, the per- 

 iod of color development was not critical as it was found that a per- 

 iod of from 20 to 30 minutes gave reasonably good agreement on repli- 

 cates. Transmittance values were determined at 640 millicrons with 

 the Becknan spectrophotometer using 1 centimeter corex cells. Con- 

 version values were obtained from a standard transmittance-concentra- 

 tion curve with ccxicent rat ions of 0.04 to 0.12 milligrams cholesterol 

 per milliliter chloroform in the final dilution. Blank determina- 

 tions showed no absorption caused by impurities in the reagents used. 



Fat Determination 



The fat content represented by the total ether soluble fraction 

 of the raw eggs was determined with a modified Mojonnier method using 

 an initial acid hydrolysis. This treatment v/as necessary to break up 

 lipid-probein complexes which are relatively insoluble in an oixiinary 

 ethyl ether extraction of raw eggs. The following procedure when ap- 

 plied to raw salmon eggs gave close agreanent on duplicate samples. 



A representative sample of 250-300 grams of ground salmon eggs was 

 homogenized for 30 seconds in a Waring blendor. Two-to five-gram sam- 

 ples were removed immediately after mixing and weighed into 50-milli- 

 liter beakers. A small glass rod and 0.5 gram of purified sand were 

 used to mix and distribute 5 milliliters of concentrated hydrochloric 

 acid throughout the sample. A digestion jjeriod of 10 minutes on a 

 warm hot plate was usually adequate to hydro lyze the eggs sufficiaitly 

 so that a fluid mixture free of lumps was produced. This was transfer- 

 red with the aid of 7-10 milliliters of 95 percent ethanol to a liojon- 

 nier tube. Fifty milliliterd of ethyl ether were added. The tube was 



70 



