558

THE

JOURNAL

OF T I t E

AMERICAN

agents are the 1,2-diols. T h e h y d r o x y l content of the reduced lanolin is at least equal to if not greater t h a n that of the saponification alcohols. About 5% of the alcohol fraction are diols (4, 15). H y d r o x y acids have been reported (5) to comprise about 30% of the acid fraction, and these compounds upon reduction would yield 1,2-diols. I t is logical to assume that the emulsif y i n g properties Of wool wax alcohols would be enhaneed by the addition of the glycols formed by the reduction of the h y d r o x y acids of wool grease. I t was found, on testing the reduced lanolin for its emulsifying properties (13) according to a procedure b y Schulman and Cockbain, that they were equal to or better than those of wool wax alcohols obtained b y saponification.

Summary The sodium reduction technique has been modified for application to various grades of lanolin and wool grease. The i m p r o v e d process gives good yields of alcohols with low ester and acid numbers. The sterols present in the grease are not affected by the reduction. A recovery procedure is described which avoids the difficulties with extremely stable emulsions. The essential features of this procedure are the elimination of emulsion-stabilizing sodium soaps by precipitation with b a r i u m chloride p r i o r to the washing of

OIL

CHEMISTS'

SOCIETY

VOL. 32

the reduction mixture and acetone extraction of the alcohols f r o m the insoluble barium soaps.

Acknowledgment The authors wish to t h a n k W i l f r e d R. Noble for the analytical data reported herein. I~EFEI~ENCES 1. Barnes, C. S., Curtis, P~. G., a n d Hart, It. IK., Austl'alian J. App]. Sci., 3, 8 8 - 9 9 (1952). 2. F a y a u d , A., a n d Rivera, S., I n d u s t r i e p a r f u m . , 8, 3 7 9 - 8 0 (1953). 3. Hansley, V. L , Ind. E n g . Chem., 39, 5 5 - 6 2 ( 1 9 4 7 ) . 4. H o r n , n . It. S., a n d Houg'en, F. W., Chelnistry a n d I n d u s t r y , 1951, 670; J. Chem. Soc., 1953, 3 5 3 3 - 8 . 5. lIorn, n . I-I. S., Houg'en, F. W., yon Rudloff, E., and Sutton, D. A., J. Chem. Soc., 1954, 1 7 7 - 8 0 . 6. Kawai, S., Japanese P a t e n t 128,286 ( J a n . 13, 1 9 3 9 ) ; C.A., 34, 8185 ( 1 9 4 0 ) . 7. Knol, H . W., J. Am. Oil Chemists' Soc., 8I, 5 9 - 6 2 (1954). 8. Luddy, F. E., Turner, A r t h u r Jr., a n d Seanlan, J o h n T., Anal. Chem., 25, 1 4 9 7 - 9 ( 1 9 5 3 ) ; ibid. Z6, 491 ( 1 9 5 4 ) . 9. Mug'isima, K., J a p a n e s e P a t e n t 128,431 ( J a n . 20, 1 9 3 9 ) ; C.A., 34, 8318 ( 1 9 4 0 ) . 10. M u r r a y , K. E., a n d Sehoenfeld, R., J. Am. Oil Chemists' Soc., 29, 4 1 6 - 2 0 ( 1 9 5 2 ) . 11. Peel, N. S., Soap, P e r f u m e r y and Cosmetics, 25, 1 2 6 9 - 7 5 (1952). 12. Radell, J., Eisner, Abner, a n d Donahue, E. T., J. Am. Chem. Soc., 76, 4 1 8 8 - 9 ( 1 9 5 4 ) . 13. Schulman, J. H., and Cockbain, E. CT., Trans. F a r a d a y Soc., 36, 651-61 (1940). 14. Tiedt, J'., a n d Truter, E. V., J. Applied Chem. (London), 2, 633-8 (1952). 15. Tiedt, J., a n d Truter, E. V., Chemistry and Industry, 1951, 911-2. 16. Weitkamp, A. W., J. Am. Chem. Soc., 67, 4 4 7 - 5 4 (1945).

[Received April 27, I955]

Some Effects of Semolina Lipoxidase Activity on Macaroni Quality I G. N. IRVINE, Grain Research Laboratory, Winnipeg, Canada PROBLEM O f assessing the m a c a r o n i - m a k i n g quality of a sample of d u r u m wheat or d u r u m semolina is largely one of predicting the color of the macaroni which the salnple will produce. The most reliable w a y of doing this is to process macaroni f r o m the sample and to measure the color of the product, either visually or by using some type of instrument. This requires too long a time for some commercial applications and is not convenient in dealing with new varieties at early stages of developnient because of the small amount of wheat that can be spared for testing. Accordingly it is necessary to seek useful indirect methods of predicting macaroni color. The desirable color for macaroni is a clear bright yellow. This results f r o m the presence in the semolina of certain yellow carotenoid pigments. The problem, then, should be simply one of determining the amount of pigment in the semolina f r o m which the macaroni is made. However when quality testing of d u r m n wheats was first begun about 20 years ago, it was noted (1) that certain semolinas a p p e a r e d to bleach badly during processing while others retained their color. F o r this reason no useful correlation between semolina pigment and macaroni color was found. I n seeking an answer to this problem some years ago, a study was made (5) of the rate at which semolina pigment was destroyed d u r i n g mixing of macaroni doughs. As a result of this work it was postulated that d u r u m wheats contained the enzyme lipoxidase and that it was the presence of this en-

T

IlE

Presented at the fall meeting of the American Oil Chemists' Society, Minneapolis, Minn., October 11-13, 1954. P a p e r No. 145 of the G r a i n Research Laboratory, B o a r d of G r a i n Commissioners for Canada, Winnipeg', ~J[anitoba, Canada.

zyme, in v a r y i n g amounts, that gave rise to the variations in pigment loss during macaroni processing. I n this brief review I will discuss the application of this hypothesis to the development of a simple prediction test which allows one to estimate the macaroni color of a sample of wheat or semolina of about 20 g. The data in F i g u r e I are taken f r o m some of our earlier work and show t h a t the rate of pigment oxiclarion d u r i n g mixing for two samples of semolina which contain the same amount of pigment initially but which v a r y widely in macaroni-making quality; The methods have previously been described (5). The u p p e r curve, showing r a p i d oxidation of pigment, is for a semolina yielding a pale m a c a r o n i ; the lower curve, showing slow oxidation of pigment, is for a semolina yielding a yellow macaroni. Following this investigation we began working with a W a r b u r g a p p a r a t u s to find out if crude aqueous extracts of d u r u m wheat or semolina showed a n y lipoxidase activity. E n z y m e extracts were p r e p a r e d by grinding the material with sand and water, followed by centrifuging ( 3 ) ; p r e p a r a t i o n of a suitable linoleie acid emulsion was eventually solved by using a non-ionic surface-active agent, Triton X-100 supplied by Rohm and H a a s (3). The p H o p t i m u m for this enzymesubstrate system was f o u n d to be 6.5 in phosphate buffer. W e found t h a t these wheats did contain signifieant amounts of this enzyme and obtained the sort of results shown in F i g u r e 2. These curves show the rate of oxygen uptake for aqueous extracts of semolinas, using the linoleic acid emulsion as substrate, at p H 6.5; they represent the same t w o types of semo]ina as those shown in F i g u r e 1. The close eorre-

Nov. 1955

IRVINE: SOME E F F E C T S

with environment t h a n with variety. Because of the limited effect of environment, it is possible to distinguish good varieties f r o m poor varieties of d u r u m wheat, in a general way, by lipoxidase measurement alone even though samples m a y come f r o m different areas, where growing conditions m a y v a r y widely. I t is even possible to distinguish between good and poor varieties when the wheat is so shrunken as to preclude identification in other ways. This technique is proving very useful for the badly rusted crops which are c u r r e n t l y being harvested.

4.00 o

o

3.00

ti3 N X

O

559

2.00

OLLOWING upon this p r e l i m i n a r y work, We studied

F the kinetics of the lipoxidase system as it occurs in wheat (2) and developed suitable manometric as-

tz

{L LO0

"

o

I I

I 3

i 5

TIME, MINUTES

FZG. 1. tlate of p i g m e n t oxidation d u r i n g mixing of m a c a r o n i

doughs for two varieties of durum wheat. spondence of the sets of curves in these two figures suggest that our working hypothesis was correct, and this has been verified in other ways which need not be discussed here. While both of these experiments were made with semolinas milled f r o m the same varieties, Golden Ball and Mindum, different samples of each v a r i e t y were used in the two experiments. The similarity in behavior of different samples of the same v a r i e t y has been noted in m a n y other experiments, and it has been established (3) that the amount of this enzyme present in wheat is a varietal characteristic. Envir o n m e n t a l variation in lipoxidase is v e r y small as compared with varietal variation whereas other characteristics of wheat, such as yield per acre, bushel weight, or protein eon~ent, m a y v a r y more widely

say methods for the enzyme in wheat and in semolina which are described in detail in two subsequent papers (3, 4). W i t h this tool the next step was to work with a large n u m b e r of d u r u m wheat samples, including both plant breeders' material and commercial carlots, to see if we could improve the prediction of the a m o u n t of pigment in macaroni by measuring not only the pigment content of the semolina but also its lipoxidase activity. P i g m e n t content was determined by an overnight extraction of 8 g. of material with 40-ml. of water-saturated n-butyl alcohol. The ext r a c t is filtered, and its transmission is measured in an E v e l y n colorimeter, using a 440 m~ filter. Pigment coneentrations are reported as p.p.m, calculated as fl carotene. The relation of the pigment content of macaroni to that of the semolina f r o m which it was made, for semolinas milled f r o m 227 different wheats, is shown in F i g u r e 3. No useful prediction can be obtained

201 J o" 2401.

l~Io. 3. Relation between pigment content of semolina and pigment content of tJ~e eorresponding macaroni. 160

80

iO

~0

6' 0

8' 0

TIME, MINUTES

Fie. 2. l~ate of oxygen uptake by lipoxidase extracts of two varieties of semolina, using linoleic acid as substrate.

f r o m these data. However, by utilizing as well the data for semolina lipoxidase, a multiple regression can be calculated, relating macaroni pigment to both semolina pigment and semolina lipoxidase ( 3 ) ; and this yields an equation from which macaroni pigment can be predicted f r o m the other two measurements. The result of plotting these predicted values against actual macaroni pigment values is shown in F i g u r e 4; this techniq~le improves the prediction considerably. The prediction equation is P~ ~ 0.669 ~- 0.726 P~ - - 0.042 L, where Pm is predicted macaroni pigment, p.p.m. ; P, is semolina pigment, p.p.m. ; and L~ is semo-

560

T H E JOURNAL OF THE AMERICAN O I L CI-IEMISTS' SOCIETY

VOL. 32

lina lipoxidase activity calculated as ~1 of oxygen per minute, per g r a m of semolina. The s t a n d a r d error of prediction of macaroni pigment f r o m these measurements is 0.38 parts per million. This t y p e of prediction test should be v e r y useful to mills, for control of semolina quality, and to purchasers of semolina in products control. The same procedure can be followed for wheat itself rather t h a n semolina (4) although one would not expect quite the same degree of precision since the semolina stage is being bypassed. F i g u r e 5 shows the relation between wheat pigment and the pigment content of macaroni processed f r o m it for 137 wheat samples. A p p l y i n g the same multiple regression tech-

9 9 6.00 d. o.

2s o_

9

.,~

4.00 ".~

0.

.',

~ z

o

.."

.'~..

..

9 " .'" "I " " 9 ::"

.'.

:.:."

2.00

9

, %, 9

6.00

..~ 9

Q. ;~

9

9. ~.::'.." 9

9.

2.00

9

9

.

SEMOLINA

..:.2..-.~. 9

. ." ,: .:..,.'...

...a0 .: "0

Loo PREDICTED

41oo MACARONI

PIGMENT

6'.00 p,p.m.

Fro. 4. Relation between macaroni p i g m e n t as predicted f r o m data on semolina and experimental m a c a r o n i p i g m e n t values.

nique, but using wheat data in place of semolina data, and again plotting predicted values against actual values for macaroni pigment, the results shown in F i g u r e 6 are obtained. Again there is a very significant improvement in the regression as a result of adding in the lipoxidase factor. The prediction equation for wheat is P m = 0.807 P~ - - 0.0105 Lw - - 0.383 where P~ again is predicted macaroni pigment, p.p.m.; Pw is wheat pigment, p.p.m.; and L~ is wheat lipoxidase activity ~l Oz/min./g. The s t a n d a r d error of prediction with this equation is 0.48 p.p.m. This prediction test should prove valuable to plant breeders; the test requires only 20 g. of wheat and, by scaling down, could be done on wheat samples as small as 10 g. The wheat prediction test should also be useful to millers for wheat selection. OW will the new methods of v a c u u m processing affect the usefulness of the lipoxidase measurement? The answer seems to be t h a t they will not affect it. We recently investigated a series of 12 semolinas of widely v a r y i n g quality and found t h a t the macaroni pigment was identical for normal and v a c u u m mixed samples. This confirms similar findings f r o m earlier work on the rate of pigment oxidation in macaroni doughs. The pronounced i m p r o v e m e n t in macaroni color that results f r o m v a c u u m processing seems to be entirely due to a much smoother maca-

H

'

9.00

p.p.m.

I

9 9 .t.,~.

0

T!oo PIGMENT

FIG. 5. Relation between pigment content of wheat and pigment content of the corresponding macaroni.

9 ~ a.'.'15 "" -

r.

5'.00 WHEAT

4.00

9 :'%'~I

o

,t t 3'.00

~176

4 ~ 99 0 ~ ~:

roni surface and to the increased translucency which results f r o m absence of air bubbles in the macaroni. These prediction tests are concerned with estimating macaroni pigment content while the ultimate goal is estimating macaroni color. H o w good is macaroni pigment as a measure of macaroni color? The answer seems to be that it is the best single p a r a m e t e r measure that we have. On the other hand, no single p a r a m e t e r will suffice to measure all possible macaroni colors encountered in commerce. F o r t u n a t e l y the great m a j o r i t y of commercial samples can be characterized very largely by a single parameter, yellowness. An additional p a r a m e t e r is necessary to deal with brownness. W o r k i n g with a reflectance spectrophotometer, we have caleul'~ted values of dominant wavelength, purity, and brightness in the C.I.E. system for about 100 semolinas and the corresponding

6,00 ci

b-

t9

E

~176 , 9176149

w

400

0,,t 909 .~~ 9 9 _%

z

0

9

~

9

~

~

~ ~

9 .,# 9 0~

0 :E

2.00

.

v , .-'~ :;'..'~:.

. ,.:', 9 :, ," .

2:oo PREDICTED

FIG. 6.

WHEAT

."

.

41oo MACARONI

6'.oo PIGMENT

p.p.m.

Relation between macaroni pigment as predicted from wheat data and experimental macaroni pigment values.

Nov. 1955

IRVINE: SOME EFFECTS

macaronis. Over the entire range of colors encountered, dominant wavelength was r e m a r k a b l y constant, and the p u r i t y and brightness values were sufficient to establish the macaroni colors observed. Of these two, brightness varied significantly only for samples which were brownish in color; otherwise p u r i t y alone would suffice to establish the color. P u r i t y values for macaroni are v e r y closely correlated with macaroni pigment values, and either will serve equally well as the color index for prediction tests. We have worked out the prediction test to yield macaroni p u r i t y values; but, except u n d e r special circumstances, we feel that macaroni pigment values are more readily understood and meaningful. I t is also possible, with semolina, to use a single reflectance measurement in place of the semolina pigment determination, along with the semolina ]ipoxidase measurement, to predict either p u r i t y of macaroni or macaroni pigment.

561

Summary I t has been established that d u r u m wheats contain lipoxidase and that the presence of this enzyme is responsible for the low correlation between the pignient content of semolina and the color of macaroni. B y taking account of the lipoxidase factor and using a multiple regression equation, it is possible to predict macaroni pigment or percentage of p u r i t y of macaroni f r o m measurement of semolina reflectance, semolina pigment, or wheat pigment. REFERENCES 1. Fifield, C. C., Smith, G. S., a n d Hayes, J. F., Cereal Chem., 16, 661-673 (1937). 2. Irvine, G. N., a n d Anderson, J. A., Cereal Chem., 80, 2 4 7 - 2 5 5 (1953). 3. Irvine, G. N., a n d Anderson, J. A., Cereal Chem., 80, 3 3 4 - 3 4 2 (1953). 4. Irvine, G. N., a n d Anderson, J. A., Cereal Chem., 3~, 88 ( 1 9 5 5 ) . 5. Irvine, G. N., a n d Winkler, C. A., Cereal Chem., 27, 2 0 5 - 2 1 8 (1950). [ R e c e i v e d A p r i l 18, 1 9 5 5 ]

Ethanolamines and Other Amino- and Hydroxyl-Containing Compounds in the Refining of Rice Oil 1 E. R. COUSINS, R. PRACHANKADEE, ~ and S. BHODHIPRASART, ~ Southern Regional Research Laboratory, ~ New Orleans, Louisiana C O M M E R C I A L R E F I N I N G O f crude rice oil with sodium hydroxide in the usual m a n n e r (3) generally results in losses that are much greater than would be expected f r o m the characteristics of the oils. While rice oils processed entirely u n d e r carefully controlled laboratory or pilot plant conditions have given refining losses similar to those of cottonseed oils with comparable free f a t t y acid content (6, 7, 13), the usual commercial losses are of such a magnitude as to make it v e r y difficult, if not impossible, to refine rice oil economically. Considerable effort has been expended in a t t e m p t s to find a practical method of reducing these losses. The Japanese, in particular, have done a great amount of work on the refining of rice oil (8, 9, 10, 11, 12). I n this country attempts have been made to refine rice oil, using special techniques (4, 13) and the m a n y methods developed for decreasing the refining losses of other vegetable oils. However, as f a r as is known, no practical commercial method applicable to rice oil has been reported. When normal refining procedures are employed, the foots formed f r o m crude rice oil are unusual in their inability to cohere and settle out of the oil cleanly. The soapstock is usually composed of small individual grains. These grains settle v e r y slowly, occlude considerable oil, and are almost as fluid as the oil itself. Without the use of special techniques for the separation of the soapstock, the refining loss on a comparatively good rice oil (content of free f a t t y acids, 5.5%) could be in excess of 50.0%. Centrifugation of the mixture or refining in solvents increases the yield of refined oil but still results in losses of about 25 to 30%. Similar reductions in refining loss m a y be obtained in the l a b o r a t o r y b y simply filter-

T

HE

1 Presented at the 46th a n n u a l meeting of the American 0il Chemists' Society, New Orleans, La., April 1 8 - 2 0 , 1955. D e p a r t m e n t of Science, Ministry of I n d u s t r y , Bangkok, Thailand. One of the laboratories of the Southern Utilization Research Branch, A g r i c u l t u r a l Research Service, U. S. D e p a r t m e n t of Agriculture.

ing the mixture through a 100-mesh stainless steel screen (5). The peculiar behavior of rice oil loots has been attributed to some unknown material in the crude oil that tends to emulsify the oil u n d e r the conditions of refining. Light non-settling foots are thus formed. I n the past year the m a j o r objective of the research on rice oil in this laboratory has been to isolate and to identify this unknown material. The results of the initial phase of thi~ work will be the subject of a forthcoming report. Recently, in the p r e p a r a t i o n of liquid floor polishes containing high concentrations of rice wax in the solid portion, it was noted that the emulsions would invariably grain out as though no emulsifying agent were present whenever a triethanolamine soap was used as the emulsifying agent. The use of morpholine instead of triethanolamine, or carnauba wax instead of rice wax, resulted in stable emulsions. A p p a r e n t l y the triethanolamine was effectively neutralizing the emulsifying properties of the mixture. Therefore an investigation of the effect of triethanotamine and related compounds in the refining of crude rice oil was undertaken. As a result of this investigation enough information has been gathered to provide the basis for f u t u r e development of t h e problem. No a t t e m p t has been made to present a complete commercial process for the refining of rice oil.

Materials and Methods Three samples of clarified crude rice oil and one of unclarified crude rice oil were obtained f r o m commercial processors. The clarified oils, used in most of the work, were filtered through diatomaceous earth to insure u n i f o r m i t y of successive samples of each oil a n d to remove the last traces of solid wax. The unclarified rice oil was used as received. The clarified crude oil having a free f a t t y acid content of 5.5% was used to establish the p r e f e r r e d refining condi-