iment of Rice tor3 (e.g., riboflavin) in t,he original brown rice arid of high milling losses for others, Tvhite riciz is a poor source of these nutrients. The magnitude of these deficiencies for the rice eater is illustrat,eti by the comI-,srison in Table I1 of t , ’ n c a content of a 500-gram ration of white rice with the minimum and reconirnended daily requirrments advocated by government authorities. Means of improving the nutritional quality of white rice may be classified in t,wo groups: older methods based on the prcscrvation of the natural nutrients of whole grain rice, and newer methods for enriching polished white rice with synthetic vitamins and miricrals. The first, d e g o r y iricludes the processes known as untlerndlirig, parboiling, conversion, and malekizing (9). Although tho products obtained by these proceses have greater iiiitritional values than M-hite rictci, they have received limited acceptance because of inlieretit, disadvantages which may be hriefly summarized as follows: 1.
Pilot Plant for Rice Premix
Although the causal relalion betweer1 twritwri and ‘i tl.et (ori.i*tiiig largelj of white rice has been recognized €or a long time, the disea*e is still rampant among the rice-eating people9 of the Orient. Becauw of the popular preference for white rice, it has been felt that the suitable enrichment of white rice with thiamine (and possiblj other vitamins) would result in a more fruitful approach to the “rice problem”. Suitable enrichment entails that the product shall not suffer any appreciable loss of the added nutrients in washing, cooking, and storing, and that the cost of enrichment shall be low. These results have been achieled b.r a n e w proress clesrribed in this report.
l ~ r i d i ~ m ~ i l liipn difficult g to royiilatc.
at
itii
s involving some type of parboiling rcquire installat~ions of special equipment. If a nat,ionwide riw vitaminization program T W R ~ adopted, the entire rice crop would have to pass through such a process, and large investment for new installations in the rice mills a.oultl be required. 3. These products are soniewhat different from ordinary white rice in appearance and flavor. Except, where prevailing custom and eating habits have already achieved this end, scccptance by rice-consuming peoples could be brought about, only by a long range program of re-education, if a t all. The situation is analogous to that in regard to enriched white bread us. whole 7%-heatbread. These objections ~ J ’ c ?largely overcome 1s , t lie ~ukverniethotl of enriching with synthetic vitamins. The basir proceduw consists of producing a fort,ified premix on a rice base, and diluting the premix n-ith ordinary whit,e rice by simple mixing. The method entails low cost, of insta,lla,tionnntl operation, as exemplified by t,he
rK and Williams (9)recently stated t h a t “tlie association of beriberi with white rice consumption is a fact familiar t o t,he reading public .Beriberi is caused by a pronounced deficiency of the antineuritic vitamin, vitamiri B1or thiamine. Up t o so~. of t,his vitamin is removed during the process of nlilling t)roTvn rice t o white, polished rice and the tiiwase is fount1 principally in those regions where people live on dirts containing large, amounts of polished rice. Beriberi ocrurs abundantly in south China, southeastcrn India, ,Japan, Kvthwlands Indies, Philippines, Burma, Siain. Indo-china and scatteringly elsewhere throughout the world.” Aykroyd’s survey ( 2 ) of t h t ~“poor riceC:Lt(3r’s’’ diet in India and its nutritional impiio:i!ioris illustrat(’h c:onditions which are prevalent throughout the Orifant. Khile t8he loss of thiamine is the outstanding deficiency incurred in the milling of rice, other nutrients suffer similar fate. Table I, compiled from &t,a in recent literature, summarizes availitbie infomiation on the fate of B-complex vitamins and iron during m~lling Con.iderable losses are incurred in all thew nutrients whcn hi o\\ n ricr 1, processed t o finished head rice, the familiar polished n hitc. I I ( i I t rommcrce. As a consequence of thv lox tontent of certniri fac486
Figure 1 .
(:oinparison of Roche Enriched with Orcliiiary 1 nenriched White Rire
noti, aamplea h a w the same appearance: t h e premix kernels in the crtrl< hed rice cannot he identified hy inspection.
with Synthetic Vitamins and Iron J
M.F. FURTER, W. M. LAUTER, Hoffmann-La Roche, Inc., . \ ~ t l e y . .V. J .
E. DE RITTER, A4NDS. H. RUBIN pilot plant process described here. The appearance (Figure I), flavor, and keeping qualities of the product are those of white rice. The number and concentration of the added nutrients can be varied considerably in accordance with any sound nutritional formula. Although white rice is generally the premix base, it is feasible to use other types, such as converted, undermilled, etc., i f desired.
d I,R
l'l
I
EVKICHED RICE
After mashing and cooking, an enriched rice should contain a t least the minimum levels of thiamine, niacin, and iron fixed by the United States Food and Drug Administration for enrirhed flour (Figure 4). Riboflavin has been excluded because of its yellow color. The rice premix is therefore made to contain the following: 1.h
Mg./Kg
500" 3200
1100" 7040 5720
\Ig.,
Thiamine Niacin Iron (added as iron pyrophosphatt? a Includes a 25% excess
:woo
Since the washing of white rice, a universal custom, removes a large part of its natural vitamin and iron content (Table IV and Figure 6), t,he natural nutrients are ignored in fixing enrichment levels. The 25% excess of Bl is added to provide a margin of safety against losses which may occur during long storage periods a t high temperatures or during prolonged cooking in. water of high pH.
.
,
-*
Figure 2.
Diagram of Pilot Plant for Production of Rice Premix
of the protective solution is applied and dried as before. The finished premix is removed from the trumbol through discharge slots and screened to remove "agglomerates" which form to the ' extent of about 1%of the rice during processing. Enriched rice is prepared by mixing premix and white rice in the ratio of 1 t o 200, either in the trumbol or in any other suitable mixer. '
VITAMIN AND IRON CONTENTS
A number of varieties of rice, including both short-grained and
long-grained types, have been enriched by the Roche process on a Pilot Plant scale. Table 111gives typical data on the vitamin and iron contents of premix and enriched rice. Each value for enriched rice content was determined on a 200-gram sample; premix 'Ontents were determined On '-gram which are equivalent to the amount of premix in 200 grams of enriched rice. Triplicate assays have been run on both premix and enriched rice to check the homogeneity of the premix and the uniformity of distribution of the Premix in the enriched rice. Since these batches were experimental, the level of enrichment is not necessarily the probable commercial enrichment level. However, it is well to reiterate that one of the chief advantages of synthetic enrichment is that the level can be adjusted to meet any practical nutritional requirements, and white, converted, or any other type of rice can be used as substrate. Table 111 indtcates that a satisfactory homogeneity of premix , The t,rumbolis 8. rotary drum UP& in the rice mill for or mixil,g has been achieved for each nutrient, the variations in content rice after pnliahing. being less than *37& from the average. The uniform distribution of premix throughout the batches of enriched rice is TABLE I. EFFECT OF A ~ I L L I N ON Q VITAMIN AND IRON CONTENTS OF RICE shown by the good agreement Mg. per Lb. (463.6 Grams).of Dry Weight-----3lilled Thiamine Riboflavin Nicotinic Pantothenic PyriBiotin among the values derived from Fraction (6) (7,sa) arid (8,85) acid ($8) doxine ($8) ($8) Irona Brown rice 1.1- 2.3 0.21-0.34 18-29 7.2-8.4 4.3-5.1 0.052-0.057 7.0-8.0 random 200-gram samples. 1st break rice 0.4- 0.8 0.09-0.22 10-15 3.7-4.8 2.0-4.0 0,028-0.037 ..... This distribution was further 2nd break rice 0.3- 0 . 7 0.09-d.22 9-14 3.4-3.6 1.2-3.5 0.023-0.027 . . . . . checked by an experiment in Brushed rice 0.2- 0 . 5 0.08-0.18 6-10 3.2-4.4 1.2-3.3 0,022-0.024 ..... Finished head rice 0.2- 0 . 5 0.08-0.14 5- 9 2.9-3.0 0.9-2.8' 0.015-0.023 3 . 0 4.o which the premix coating was Bran 5.4-15.0 0 . 7 -1.5 117-221 30-37 11-18 0.17 -0.25 Polish ' 6.8-13.0 0 . 5 -1 4 04--196 . 33-50 13-15 0.24 -0.38 ,,, . colored blue. After manufaca Dat,a of .Roche Nutrition Laboratory. ture of a batch of 'enriched rice, twenty samples of 200
ENRICH~~ENT PROCESS.Experimental batches of rice premix have been manufactured in the pilot plant shown in Figure 2 and the PhotograPh on Page 4%. m7hite rice R is introduced into t.he motor-driven trumbol', A , through a hopper. The trumbol is rotated at a fixed rate while the vitamin solution from kettle B is allowed to flow through measuring cylinder D and into the trumbol via distributor pipe E which slowly sprays the solution over the rotating rice grains through perforated section F. After the final portion of the vitamin solution is blown out of the air from G, the blower is started sprinkler pipe with to draw preheated air through the rotating trumbol until the rice is thoroughly dry. Half the coating solution i n kettle C is rlieasured into the system through D and sprayed Over the 1-0rating rice through F. After thoroughmixing has been achieved and the major part of the solvent evaporated, the iron pyrophosphate mixture is added by a scoop which slides on F. After the iron coating has been thoroughly dist,ribut#ed,a second coating
,
487
INDUSTRIAL AND ENGINEERING CHEMISTRY
488
30
NIACIN
TABLE 111. UKIFORMITY OF ENRICHMENT Eampling in Trumbol Front Ariddie Rear Average
MG/LE.
MG/L
0.5t
Comparison of \'i tamin and Iron Contents of Enriched and Uneiiriched Rice \c it11 Jliiiimunl Enrichment Le\els for Flour
Figure 3.
-
-----
-
R Rorhr eririchrd. R = b r o w n , C oonveriwl, U = onpnlishrd, W = white rice. h l i n i m u m flour e n r i r h m e n t lereln, U. S. Food and Drug Adtninistraiiotl
grams each were taker1 at random, and the colored grains were counted and weighed in each sample. The greatest variation was found to be =!=Z grains per sample or * 4 % of tlie theoretical 50 grains. Figure 3 compares the vitamin and iron contents of natural and enriched rice. The contents of the natural and converted rice are averages for each type. The enrichment levels of B,, niacin, and iron in the enriched rice satisfy the minimum Aour enrichment requirements after the usual household TTashing and cooking. Figure 3 illustrates again the nutritional deficiencies of white rice.
-ThiEnriched Rice, Mg./Lb. ...__
amine 2.9 3.1 3.1 3.0
Niacin 10 7
Iron 12.5 13.1 12.3
18.6
19.0 19.1
12.6
tation. Since household washing practices vary, any tttteinpt a t quantitative estimation of washing losses must be based on the adoption of an arbit,rary washing method. Two general procedures have been adopted in this laboratory for determining washing losses-namely, the South Caroli ria method and a mechanical procedure designated as the Roche method. LOSSESBY SOUTH C.4ROLISA TEST. This method is based on the official South Carolina washing test for enriched grits (15). L'Customnryrinsing" is defined to bc equivalcnt to that obtained with the following procedure: "Place one-half pint of grits in a pan containing one quart of Tvater a t 25" C., Stir and rotate for one minute, allow to settle one minute, drain off arid discard +rater, add a second quart of fresh mater, stir and rot,ate for one minute, allow to sattle one minute, drain off and discard water." This procedure has been applied to rice with the added stipulation that the stirring be a t a constant rate of two revolutions per second in order to achicve maximum reproducibility. Table IV givcs washing losses of vitamin I3,, niacin, and iron by this procedure for a series of rice prcmixes and for white rice. The protection of the added nutrients in the pwrnixes against washing is practically complete in each case. White rice, however, suffers extremely high losses of vibamins and iron, xvhich are apparently concentrated for the most part close to the sudace, perhaps as a result of the milling operations. These high washing losses for white rice are confirmed by the observation of Swaminathan (16, 17) that 60T0 of t,he vitamin B, and niacin in raw milled rice are lost in washing with cold water.
TABLE11. COMPARISON
O F k h N I M U M AND RECOMMENDED REQUIRERIEXTS UITH CONTENT OF ~OO-GRAM PORTIONS OF RICE
Iron 10
12
75 18 65
15
Food, Drug, and Cosmetic Act, regulations under Section 403 (J), Federal Register h o v 22 1941. b Natl. R h r c h Cbuncil, Reprint Circ. Series 115 (Jan., 1943).
7 - S . Carolina Washing Losa, yoVitamin BI Kiauin Iron
Rice premix Blue Rose, A Blue Rose, B California Rexoro
0.1 2.8 0.3
0.6 0 1 1.2
1.3
0.6 0.6 83
1.1 63
Average White rice, av.
0.1 0.2 0.0 0.1 0.1
65
To determine tlie effect of the wash water temperature on the washing loss from the premix, the Rexoro premix of Table I V was also washed with water a t 25" to 55" C. with the following results:
.
0
Rice Premix, bIg./Lb. Thiamine Niacin Iron 480 2220 1810 510 2310 1940 500 2300 1770 500 2310 1840
TABLE IV. WASHISGLOSSESIX SOUTH CAROLINA TEST
RETENTION OF KUTRIENTS IN WASAIh-G
The custom of washing rice thoroughly before cooking is universally prevalent in spite of t,he fact that appreciablc loss of nutritional value is involved. Consequently any method of Bynthetic enrichment of white rice must afford protection of the added nutrients against washing. In the Roche process this is accomplished by a water-resistant film which protects the enrichment premix. Thus, any washing losses which are incurred are due almost entirely to the usual losses from the untreated white rice grains. The percentage retention of nutrients in washing any type of rice depends on a number of factors, including length of washing period, volume and temperature of wash water, and vigor of agi-
(Based on the average content of washed cooked rice) Thiamine Riboflavin Nicotinic Acid Daily a d d t requirement, rng. 1.0 2.0 Not established Miiiiniuni" Recorniriendedb 1.8 2.7 18 yo of minimum requircment 158 13 ... Brown rice 15 5 ... White rice yo of recommended requirement 88 10 123 Brown rice White rice 8 3 10
Vol. 38, No. 5
Tcmpcrature, ' C. 25 35
BI Loss, % 1 3 2 9
Tempcraturc, ' C. 45 :5
Bi
1,088,
% '
9 1 12 7
The protedtive coating provides almost complete protection at temperatures which are most commonly used for n7ashing (up t o 35" C.). At 55" C., an uncommon washing temperature, the loss is still relatively small. However, the added nutrients are physiologically available from the rice after normal cooking (Table XI). The washing loss of any nutrient from the enriched rice may be considered as a summated loss from the premix and from the untreated white rice. T h t t portion of the loss due to the white rice is unavoidable and constant for any given rice variety; the loss due t o the premix is a controllable factor dependent upon the premix content and washing loss. Table V gives average vitamin and iron losses for a number of experimental batches of enriched
.
INDUSTRIAL AND ENGINEER1N.G CHEMISTRY
May, 1946
2.0
washing. As in t h e S . Carolina washing, the losses of vitamin B1, niacin, and iron are extremely high for white rice, even for relatively short washing periods. For Roche enriched rice the washing losses me due largely t o losses from the white rice, especially in short washing periods during which the premixes suffer practically no loss. Table VI shows a breakdown of the enriched rice prashing losses into fractional losses from premix and white rice. Since the-white rice loses a considerable part of its vitamin and iron content while the premix loses practically none, it is again evident that an enrichment program should be based only on the amounts of these nutrients added in the premix. It must be emphasized that even a 10-minute wash in the rotating jar is much longer than the average household rinsing. As an example of losses by a procedure which is more typical of household washing, Table VI1 gives data of Kik and Williams (5) on Roche and Fieger enriched rice. E. A. Fieger, of Louisiana State University, developed a method of enrichment with synthetic vitamins in which the premix is prepared by impregnating polished rice with an aqueous solution of thiamine, niacin, and a highly soluble salt (primary sodium phosphate) ; the premix is coated with a thin collodion membrane (9). The experiments were carried out in Kik's laboratory (5). His washing technique resulted in considerably smaller losses than those found in our laboratory by both the S. Carolina and Roche washing procedures; the latter must therefore be considered as rigorous washing tests; Kik's results show distinctly lower washing and cooking losses from Roche rice than from Fieger's rice.
20
MGJLB.
10 .
IO
R
B
C
W
.
R
B
C
W
R =ROCHE ENRICHED RlCE B =BROWN RICE C 5 CONVERTED RICE W: WHITE RICE = F.O.A. MINIMUM FLOUR LEVEL
---
=
ORIGINAL RlCE
=
S.CAROLINA WASHED RICE
=
S.CAROLINA WASHEO AND COOKED RICE
( NO COOKING WATER DISCARDED)
Comparative Retention of Vitamins and Iron on Washing and Cooking Various Rices
Figure 4.
rice. The experimentally determined losses are compared with values calculated as the sum of the predetermined losses from the premix and from the white rice. As would be expected, good agreement is obtained for both vitamins and iron between the experimental and calculated values. The breakdown shows t h a t the losses incurred in washing enriched rice are due almost entirely to losses from the untreated white rice rather than from the premix. Thus, the added nutrients are practically unaffected by washing, whereas the natural nutrients are removed t o a large extent. Figure 4 compares the retention of vitamin B1, niacin, and iron i n the S. Carolina washing test for Roche enriched, brown, converted, and white rice. The enriched rice after washing contains more than the minimum flour enrichment levels of the two vitamins and iron. Brown and converted rice are above the flour level for niacin, but below for vitamin Bi and iron. White rice, which is deficient in all three nutrients even before washing, is considerably poorer after washing. LOSSESBY ROCHETEST. Seventy-five grams of rice plus 375 ml. of water are rotated in the screw cap jar (Figure 5) a t a constant speed of about 3 r.p.m. t o ensure uniform and reproducible washing. The jar rotates on rubber rollers, one of which is driven by the small induction motor geared t o slow speed. B y this method the washing may be continued for any desired period. Figure 6 shows comparative washing losses for washing periods up to 20 minutes, which is much longer than the usual household
Vitamin Niacin Iron
489
RETENTION OF NUTRIENTS IN COOKING
I n considering nutrient losses which occur during the cooking of rice, it must be emphasized t h a t such losses can vary considerably depending on the cooking method or recipe used. Methods involving cooking with a large excess of water, followed by discharge of the cooking water, or methods involving steaming with subsequent discard of the condensed water, lead t o high losses regardless of the kind of rice used. Unnecessarily long cooking also causes increased losses of vitamins. From a nutritional viewpoint the most sensible method of cooking rice is t o add just sufficient water to produce a palatable but not too soft rice by the time the cooking water is all absorbed. The use of a double boiler is desirable t o avoid high temperatures at the bottom surface of the pot which tend t o accelerate vitamin losses. However, if a reasonable cooking time is allowed, vitamin losses are not excessive even with direct heating. Vitamin B1and niacin cooking losses have been determined with the following cooking procedure: 750 ml. of water containing about 10 grams of salt are brought t o a boil in a n aluminum pan; 200 grams of rice are added and cooked over direct heat with OCCBsional stirring for a t least 30 minutes. Heating is continued until the water is completely absorbed. This is the preferred method of the U. S. Army (18). Figure 4 compares retention for Roche enriched, brown, converted, and white rice. The cooking losses of vitamin B1 were 7-10oJo WASHINGLOSSES TABLE V, AVERAGESOUTHCAROLINA and of niacin 2-4'%, regardless of the kind of rice Premix White Rice Washing L,oss of cooked. I n these experiments the p H of the cooking ConWashConWashEnriched Rice, % water was approximately 6.5 initially and 6-6.5 after ing tent ing -Calcd. from.tent loss, mg.i loss, White Exptl. mg.) completion of cooking. Iron is not lost in this pro% lb. % Premix rice Total total lb. cedure since none of the cooking water is discarded. 472 1.1 0.4 63 0.9 9 10 11 BI 2260 2170
0.6
0.1
7.3 3.3
83 65
0.3 0.1
33 15
.
33
35
15
17
TABLE VI.
BREAKDOWN OF WASHING LOSSESIN ROCHEMETHOD IN PERCENTOF CONTENT IN ENRICHED RICE" -1-Min. Premix
WashWhite
Enriched
-5-Min. Premix ' 2 1
WashWhite
6 3.8 4 0.2 Vitamin Bi 18 24 0.1 17.9 Niacin 9.8 6 6 0.2 0 Iron a .Loss from premix plus loss from white rice equals loss from enriched rice.
Enriched
8 25 10
-10-Min. Premix
5 3 2
WashWhite
Enriched
8 26 9
13 29 11
-20-Min. Premix 9 6
4
WashWhite
10 29 8
Enriched I9
34 12
INDUSTRIAL AND ENGINEERING CHEMISTRY
490
Figure 5 .
Iiofiniann-La Roche Initial Washed 3 times6 Cooked in double boilere Cooked in open vesseld Fieger Initial V a s h e d 3 tirnesb Cooked in double boilerc C o o k e d in open vesseld
Rice Tashiiig Apparatus
2 43
2.2i
2.351 2.12 2.14 1.83 2.00 1 09
Vol. 38, No. 5
8 3 14
ii
G
10
20.0 18.2 10.7 18,5
1%
20.0 17.0 20.0 13 3
36
!I
1
I4 4
T h e writers are indebted t o 11. C.IGk for permission t o rcpro(iur:e thir ahle. 6 Onc cupful of rice is covered witli one cupful of wawr ani1 stirred: t t c supernatant liquid discarded. This is repezited twice. C One cupful of rice and three half-cups of boiliilg water are placed in t i i t top of a double boiler. All the water is absorbed in tho cooked rice, which i. not rinsed afterward. d One cupful of rice is placed in a n ope11 vcssel with ten cupfuls of boilini: water, cooked, placed i n a colander, and d r a i n d . Q
i
Table VI11 illustrates the cooking loss in the method described above as contrasted Tvith methods which involve draining off more or less of the cooking xr-ater. I t is apparent t,hat the losses of thiamine are roughly proportional to the volume of water drained off after rooking, nhich bears out the contention that the proper cooking of rice should involve no such drainage. I t is noteworthy that the relative losses, expressed :is a percentage of the original content of thiamine, are similar for both enriched and converted rice. The fact of this agreement provides evidence that the vitamin is dispersed uniformly throughout the cooked mass and hence is lost in proportion to the volume of water drained off. The absolute amount of thiamine is considerably greater in all three cases in the enriched rice, which illustrates the greater flexibility of this product, in providing greater nutritive values under a variety of circumstances.
periods up to 12 months at conqtant room temperature (23" G.'i and the Lend-Lease accelerated aging test which involves storagc a t 45" C. for 21 days. St,ability data are presented in Table IX.: it is evident that the vitamin B: loss is small and thc niacin 1oss negligibIe2. Although the lms of B, during st,orage of unenriched white rice has been found to be appreciably higher (as much as 15% after 9 months a t room temperature), the loss for enriched rice is not greatly affected, becaiise the unenriched rice contrihLites only a small fraction of the tot,nl B, content. Since the average period of commercial storage of rice in the United States may be taken as &out five moiit>hs(Q),it is apparent that vitamiri losses diiring normal storage i-iill not, be significant.
STABILITY O F ESRICI-IEU KICE
Both premixes and enriched rice have been stored in paper. burlap, tin, wood, and glass containers for periods up to 12 months under various climatic conditions up to 40" C., including high humidity. The premixes did not deteriorate physically, and the addition of premix to n-hite rice did not affect the keeping qualities of the white rice. The question of separation of the premix in enriched rice during packing, transportation, and storage was investigated by t'he colored premix technique previously described. Samples T-ierc stored, unpacked several times, and shipped by truck and railroad in various containers. No separation of the premix grains was found under any of these conditions. The stability during storage of the vit,amins in prcmixes and enriched rice was determined by two procedures: storage for
y I r r R I ' r i o u I, GXPERIMENTS
Physiological responses t,oenriched rice have been st,udicciiii t w o types of experiments. The one tests the tolerance of rats for large doses of the premix (toxicity tests), and the other measures t,he physiological availability of the vitamins t o rats and to h u mans. This experiment was designed to study growth TOLERANCE:. Lmd reproduction in successive generations of rats fed t . 1 1 ~riw 1)rernix. The original group consisted of forty weai!liiig rats irhicli were separated into four groups containing equal numbcr!: b?th sexcs. These n-ere fcd a Ftock diet of ground Purina d o g r!lo~vt o n-hich were added gr:~ded amount,s of ground, uncooketl rice premix in tlic. proportions shon-n in Figurcp 7 and Table S. The growth curves in Figure 7 sl~owthat thc iiigestion of rice premix in amounts ritnging up t,c 10% of the ration did not retard the growth of the original group of rats. The female rats in the original group were mated according to the program and with the results given in Table X. The ingestion of large amounts of premix had no delet,crionL effects on reproduction. A full-grown rat, consumes 10-15 grams of food daily. On this basis the animals in group 4 (10% premix) had LZ daily iritakc of 1-1.5 grams of premix, an amount which would be consumed by :i human eating 200-300 grams of enrichcd ricc daily. From the data in Figure 7 and Table X, it i.q evident that the consumption of this large amount (for a rat) of premix had no discerniblc c f f r ~ t (1;
60 0 0
5 cn
40
20
5
IO
15
20
5
IO
15 20
WASHING PERIOD - MINUTES
Figure 6.
Comparative Washing Losses by Roche Method
ER = Roche enriohed, B = brown, C = converted, W = white, RP = rice premix
2 After the above was written, d a t a hecaine available o r three premises, made with Blue Rose, Cnlifornia, s n d hexorc rice, which were kept a t 4 5 O C . for G months. These shaweii BL losses of 6 , 13, a n d 1070, and S. Carolina wnshin:. losses of 1, 2, and 2%, respectively. These losscs are qnili, small for such an extended period of acceleratad acinF.
May, 1946
INDUSTRIAL AND ENGINEERING CHEMISTRY
491
which was fed the basal diet EXPERIMENTAL only, manifested a weight plateau a t about 100 grams. D Bi Content PHYSIOLOGICAL .AVAIL' after.Cooking, Total BI ir ABILITY TO HUMANS.The Drained and Bi .Lost in Q Washed Rice Drain physiological availability of the thiamine and niacin in the Army methoda enriched rice was studied in .. *. .. .. .. 88 Roche enrichedb 3.1 2 7 1.17 1.05 90 .. .. .. .. .. Convertede six normal male subjects Ex tl. No lp' (chemists) according t o the 2 0 65 2 8 90 1130 24 0 72 23 0.75 3 1 floche enrichedb 98 1140 0 77 66 1 14 30 32 0 35 1 17 0.37 Converted" bioassay technique which has Exptl. No. 2' 1 00 32 2 8 90 239 52 57 1 69 3 1 1 77 Roche enricchedb been elaborated recently by Converted 1.17 0.61 52 0.58 49 0 46 39 1 07 91 345 Oser and eo-workers ( I O , If). a Rice boiled in minimum of water until all water is absorbed ( 1 8 ) . In normal human subjects the b Blue Rose white rice enriched with premix C-1-88B. C Converted rice, purchased in Nutley, N. J. urinary excretion of the waterd Rice boiled in excess water, drained, and washed. 8 Rice boiled in minimum of water, drained, and washed. soluble vitamins, as such or as their derivatives, is directly proportional t o the quantity consumed, provided the subjects are subsisting on an adequate diet at the time of the on either growth or reproductioii. Such variations as did occur test. To calculate the degree of vitamin availability from a test among the various groups are within the limits of random variamaterial, the extraurinary excretion after taking the test dose (of tion for this type of experiment. enriched rice) is compared with that following the oral administraPHYSIOLOGICAL AVAILABILITY TO RATS. The availability to tion of an aqueous solution of the vitamins-i.e., the form rerats of the thiamine in the uncooked rice premix was tested in a garded as most readily available for absorption. prophylactic trial. The basal diet was the U.S.P. B1-deficient To achieve suitable precision in the conduct of this bioassay, ration (19). The ground premix was fed at a level providing 4 it was found necessary to feed the rice in more concentrated form micrograms of thiamine per rat per day, which approximates than usual; the test dose consisted of 100 grams of cooked rice the growth requirement ( b l ) , and was compared with a control containing 10 grams of premix, sufficient t o provide an intake of group which were fed 4 micrograms of pure thiamine per day. about 10 mg. of thiamine and 50 mg. of niacin. Under these conThese two groups showed identical growth over a 6-week period ditions, both vitamins were found to be completely available, the (Figure 8), an indication that the thiamine in the rice premix was average of two runs being 95 * 13% availability for thiamine fully as available to the rats as the crystalline thiamine. It is and 97 * 22% for niacin. Table XEgives a protocol of a typical also noteworthy t h a t the total growth (to a weight of about 170 thiamine experiment. These results demonstrate that, even when grams) was essentially the same as that, observed in rats on the stock diet for 6 weeks (Figure 7). The negative control group, present in amounts which are twenty times greater than usual, the components of the protective coating do not interfere with the physiological availability of the vitamins in the enriched rice. TABLEVIII.
VITAMIN B1LOSSES IN COOKING RICEB Y ARMY METHOD AND METHODS OF QUARTERMASTER DEPOT
BY
I .
270)-
TEST PROCEDURES
VITAMINBI. The assay method is the thiochrome procedure essentially as published by Hennessy (6). It includes extraction,
enzyme digestion, purification with Decalso (synthetic zeolite), and oxidation t o thiochrome which is measured fluorometrically. For assay of the premix, the enzyme and Decalso treatments are unnecessary because of the high potency. NIACIN.Assays have been carried out chiefly by a chemical method with spot checks by microbiological assay. The chemical
210-
-
TABLEIX.
9
k
Storage" at Room Temp., (24' == 4' C.) Bi loss, Niacin Myths %b loss, % b 12 6 9 3 ,; 9 7 0 9 5 3 9 5 -2 9 2 -1 9 6 -2 5 3 -4 4 4 0 4 3 8 2 0 2 2 1 -6 7 4 0
a +
u)
150Y 0
I-
5
B*
n
P
0
Figure 7.
STABILITY OF VITAMINS I N PREMIX AND ENRICHED RICE
WEEKS
e
12
Effekt of Feeding Graded Doses of Rice Premix to Weanling Rats
Lend-Lease Test (21 Days at 45* C.) Niacin Bi loss, %b loss, % b
Rice premixC Sample A 3 2 Sample B 1 4 Sample C 9 -2 Sample D 5 1 Sample E 6 -5 Sample F 6 3 Sample G 8 -3 Sample H 7 4 Sample I 7 -6 Sample J 3 0 Sample K 6 2 Sample L 8 6 Average 6 1 Enriched rice Blue Rose 8 7 2 9 7 8 7 6 4 -5 California Rexoro 8 3 -3 7 4 Average 8 6 2 7 2 a Tbe rice premixes were stored in screw-cap glass jars, and the enriched rice in carpenter drums lined with glazed paper. b The random variations of the individual assay values are due to experimental ermrs inherent in the chemical assay methods and hence are not significant. In addition, two premixes stored a t 30" C . for 8.5 and 9 months showed BI losses of 3 and 1 %, respectively.
492
INDUSTRIAL AND ENGINEERING CHEMISTRY
Vol. 38, No. 5
3
I70
2
I50
E
-
130-
W
v
v)
a
110-
LL
0 * I -
I
Figure 9. Rapid Iron Test for Roche Enriched Rice
a 3
90-
*0
Premix is represented by the dark (red) kernels.
0
tion are not necessary for assays of the premix because of the high potency. Rlicrobiological niacin assays were conducted according t o U.S.P. procedure (PO). The rapid qualitative test for niacin in enriched Aour developed by the War Food Akdmiiiietration(4)has been applied t o the enriched rice. Twenty milliliters of 47, aniline in ethanol are added I I I I I to 100 grams of enriched rice in a beaker, followed by 30 ml. of 30. I 2 3 4 5 6 37, aqueous cyanogen bromide. The beaker is swirled and allowed to stand. After about 4 minutes the premix kern& deWEEKS velop a yellow color due to the reaction of niacin with cyanogen Figure 8. Availability to Rats of Thiamine in bromide and aniline. This test indicates the presence of added Rice Premix niacin in the premix by making the prcmix grains easily distinguishable from the unenriched rice. If it is desired t o count the number of premix grains in the portion of rice tested, the remethod is a modification of the Dann (3) and Perlzweig ( I d ) proagents may be rinsed off with several small portions of water cedures. It is a colorimetric determination based on the reacwithout washing the color from the premix grains. The teat tion of niacin with cyanogen bromide and an aromatic amine, p can be used as a rapid method of determining the uniformity of methylaminophenol sulfate (Metol), to form a greenish-yellow distribution of premix in a batch of enriched rice. compound. I n assaying enriched or unenriched rice, 3 N hydroIROX.The A.A.C.C. assay method ( 1 ) involves the reaction chloric acid is used for extraction and hydrolysis of niacinamide between ferrous iron and a,n'-dipviidyl to produce a red complex t o niacin. Filtered extracts are purified by adsorption on and which is measured colorimetrically. The h.,I.C.C. qualitative elution from Lloyd's reagent and by two successive treatments test ( 1 ) for iron in enriched flour has also been applicd t o Itoc.1ic of the eluate with lead nitrate. The hydrolysis and decolorizaenriched rice. Twenty-five milliliters of potawum thiocyanate solution ( I to 10 in water) and 25 ml. of 2 X hydrochloric arid itre mixcd and added a t once to 100 grams of enriched ricc in a beaker. After 10 minutes the premix grains which contain large amocmts of iron become intensely red and easily di-tinguisliable (Fiyurr 9). If it is desired to count these premiS kernels, the reagents TABLE X. REPRODUCTIVE SVCCESS IN RATSFEDG R ~ D E D may be removed by two successive, rapid rinses with water. This DOSESOF RICEPREMIX test can also be used as a rapid means of determining the uiiiformity of dibtribution of premix grains in a batch of enriched lvt, Av.of No. hTo. of No. of Size Young, rice. Females Times Pregof 22nd RIBOFL4VIN. A fluorometric assay method developed in this Group Diet Alated Alated nancies Litter Day laboratory ( I S , 14) is usrd. 1 Puiina dog chow 5 3 12 14 36 IODINE TEST. Another rapid method has been devised for de2 f O 1570piemiv 4 3 12 38 termining the distribution of premix grains in enriched rice. It 3 + 3 0% piemix 5 3 192 12 37 4 + l o % premix 5 3 14 11 37 consists simply of shaking the enriched rice with a dilute iodinepotawum iodide solution. The untreated ricc grains are blackencd by the starch-iodine reaction, 1% hcreas O F T H I A W N E IN TABLE XI. PHYSIOLOGICAL AVAILABILITY" the premix grains remain white because of the proEKRICHED RICE tective action of the coating film (Figure 10). This procedure has the advantages of being rapid and not Control Dose: Test Doseb: involving dangerous reagents such as cyanogen broEnriched Rice, 9.2 M g . B1 Ciystalline Thiamine, 9.0 Mg. Extra Extra mide used for the rapid niacin test. Consequently, Excre- excroExcre- excreit provides a safe factory control for determining the Excretion tiont o Contiol Excietion tiont o number of premix grains in a given weight of cntion on due after due tion on after Test dose basal test test dose riched rice. basal control contiol I - N E G A T I V E CONTROLS 2 - 4 M C G 81 P E R GM OF D I E T 3 - 4 M G R I C E P R E M I X PER G M DIET
diet, dosec, recovdose, dosec, diet, dose, recovmg./ ered, mg / mg./ ered, mg./ mg / mg / Subject day day day 70 day day day 70 EDR 0 31 1 27 0 96 11 0 28 1 49 1 21 13 1 37 1 09 12 1 28 14 0 28 SHR 0 24 1 52 15 0 41 2 02 EH 0 69 2 06 1 37 1 61 18 1 72 1 33 15 0 48 1 24 14 JCB 0 48 1 81 8 0 44 1 72 LD 1 00 1.67 0 67 1.28 14 0 39 1 84 1 45 16 FWJ 0 62 2 06 1 44 16 Av. 0 56 1.73 1 17 13 2 0 38 1 69 1 31 14.5 5 Availability of thiamine in rice = (14.75/13.2) X 100 = 110 * 11% (standard error of quotient). b The subjects ingested 100 grams of rice containing 10 grams of premix as t h e t e s t dose. 0 T h e excretion on t h e basal diet is subtracted from the excretion after the dose t o obtain excretion d u e t o t h e dose. This IS permissible since t h e basal diet -as exactly duplicated on the succeeding d a y when the dose of thiamine was given.
SUlIkI4RY
h method is described for the enrichment of rice with synthetic vitamins and minerals, based upon the preparation of a suitable premix. This premix provides thiamine, niacin, and iron in amounts equal to 'IUiched flour, after the enriched rice has washed and cookcd. The rice premix is resistant to washing, cooking, and storage losses. The small washing losses from the Premix deserve Particular notice, since loss in washing constitutes the chief difficulty in the retention of the nutrients in enriched rice. Un-
May, 1946
INDUSTRIAL AND ENGINEERING CHEMISTRY
493
search and Nutrition Laboratories who contributed their efforts to this work. LITERATURE CITED
(1) Am. Assoc. of Cereal Chem., Cereal Laboratory Methods, 4th ed., p. 47,Lincoln, Nebr., 1941. (2) W. R., Bull. Health Organization League Nations, 9, . . Aykroyd, . 342 (1940-41). (3) Dann, W. J., and Handler, P., J. Biol. Chem., 140,201 (1941). (4) Feinstein. L.. Science. 101,675 (1945). (5) Hennessy, D. J., IND. Eao. CHEM.,ANAL.ED.,13,216 (1941). (6) Kik, M. C., Cereal Chem., 20,103 (1943). (7) Kik, M.C.,and Van Landingham, F. B., Ibid., 20,563 (1943). (8) Ibid., 21, 154 (1944). f~, 9) Kik. M. C.. and Williams, R. R.. Natl. Research Council, BUZZ. ii2 (1945). (10) Melnick, D., Hochberg, M., and Oser, B. L., J . Nutrition, 30,
.-,
Figure 10. Rapid Iodine Test for Roche Enriched Rice White premix kernels are liot darkened by the etarch-iodine reaction because of their protective, water-repellent coating.
enriched white rice loses most of its small content of 'thiamine, niacin, and iron readily, even with gentle washing methods. The content in white rice is therefore disregarded in preparing enriched rice, reliance being placed entirely on the content in the premix. Studies of growth and reproduction in rats have demonstrated that massive amounts of the premix are tolerated without ill effect. Further studies with rats showed that the thiamine in the raw premix is readily available for growth. Both the thiamine and niacin in cooked enriched rice are completely available to humans, as demonstrated by quantitative measurements with the human bioassay technique.
A 945). -.7 I1 \_ ---,-
(11) Oser, B.L., Melnick, D., and Hochberg, M., IND. E N ~~ .H E M . , ANAL.ED., 17,405 (1945). 112) . , Perlzweia, W. A., Levy, E. D., and Sarett, A. P., J . Biol. Chem., 136,729 (1940). (13) Rubin, S. H., and De Ritter, E., Ibid., 158,639 (1945). (14)Rubin, S. H., De Ritter, E., Schuman, R. L., and Bauernfeind, J . C., IND. ENG.CHEM.,ANAL.ED.,17,136 (1945). (15) S. C.Legislature Act, H.R. Bill 347,Senate,Bill 382,approved by Governor April 14, 1943. (16) Swaminathan, M.,Indian J . Med. Research, 29,83 (1941). (17) Ibid., 30,409 (1942). (18) U.9. Army Cooking Manual TM-10-405,p. 197,April, 1942. (19) U.S. Pharmacbpeia XII, p. 625,Easton, Pa., Mack Printing Co., 1942. (20) Ibid., 1st Supplement, p. 75 (1944). (21) Williams, R.R.. and Spies, T. D., "Vitamin Bi in Medicine", p. 313,New York, Maimillan Co., 1938. (22) Williams, V. R., and Fieger, E. A., Cereal Chem., 21, 540 (1944). (23) Williams, V. R.,Knox, W. C., and Fieger, E. A., 20, 560 (1943). .
ACKNOWLEDGMENT
The authors wish t o express their gratitude t o L. Hodax for the photographs and t o the yarious members of the Applied Re-
~
r
Drying Oil and Oleoresinous Varnish Films INCREASE IN ACIDITY ON AGING VINCENT J. FRILETTE Ridbo Laboratories, Inc., Paterson, N. J .
A
method is described for accurately determining the acid number of an oil or oleoresinous varnish film. The method is similar to an ordinary acid titration, but is on a semimicro scale. The acidity of oils and oleoresinous varnish films was studied by this method. In all cases a large increase in acidity occurred with aging. Linseed oil films showed much larger increases than tung oil films,
and oleoresinous varnish films showed much smaller increases than the corresponding oil film. It was also found that while the acid number increased, the alkali resistance decreased. For several oleoresinous varnish films the alkali resistance of the films could be directly correlated with the acid number 6f the films; the age of the films was thus eliminated as a factor.
T"
which occur during the useful life of the film. Chemists have concerned themselves mainly with reactions in the first two categories. Many problems remain t o be solved, but considerable success has been achieved in correlating technical and chemical properties. The chemical changes in the third, or aging period, are by far the most important technically, for these changes determine how well the film will fulfill its purpose. Numerous practical tests have been developed t o evaluate the suitability of films for varipus applications; among them are
chemistry involved in the drying of oil and oleoresinous varnish films has been the subject of much study.' The problem is difficult 'because the chemistry is known t o be involved, and because it is difficult t o investigate these films by chemical treatment once gelation has taken place. The changes in chemical and physical properties of oleoresinous coatings may be divided into three categories: (1) those which occur in preparing the coating; (2) those which occur between the time the coating is applied and the time i t has reached the usefully "dry" stage (usually less than 48 hours); and (3) those
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