Reducing gastrointestinal irritation in infant nutrition

Food products for nutrition of infants which contain no more than subirritant amounts of free long-chain (C.sub.16 -C.sub.22) fatty acids and triglycerides thereof are disclosed. Providing esters, such as ethyl esters, of such fatty acids in infant food products essentially eliminates the tendency of the fatty acid to damage the infant intestinal epithelium, but permits absorption and processing of the fatty acid moiety.

BACKGROUND OF THE INVENTION 
The present invention relates to infant nutrition, and more particularly to 
the nutrition of premature infants and infants of low birthweight. Such 
infants are at risk of irritation and damage to the intestinal lining 
(epithelium), which can lead to serious disorders such as necrotizing 
enterocolitis. Such disorders can lead to chronic gastrointestinal 
distress, and can even be fatal to the infant. 
It would obviously be desirable to be able to reduce or even to prevent 
occurrence of intestinal damage in the infant. It is all the more 
desirable to be able to achieve this objective by enteral administration 
of an appropriate agent in the infant's food. However, while the etiology 
of intestinal damage leading to necrotizing enterocolitis is not known, it 
appears that infants fed exclusively with infant formulas are at higher 
risk for incurring such damage than are infants fed with breast milk. 
Confronting this condition through any modification directed at the fatty 
acid content of formula has not been considered because heretofore it has 
been presumed that lipids are benign to the infant. 
SUMMARY OF THE INVENTION 
Surprisingly, it has now been discovered that digestion products of 
triglycerides, in particular free long-chain fatty acids and 
monoglycerides, can damage the intestinal epithelium of infants and thus 
could mediate the onset of disorders such as necrotizing enterocolitis. 
One aspect of the present invention is thus food products for infant 
nutrition which contain no free fatty acids and no triglycerides which can 
be broken down into such free fatty acids, or contain such substances only 
in amounts thereof which are insufficient to cause such damage. Another 
aspect of the present invention is the discovery that on enteral 
administration to infants of monoesters of long-chain fatty acids, 
including especially esters of linolenic acid and of linoleic acid, the 
esters can be absorbed and transported through the intestinal epithelium, 
thus enabling the infant to process the fatty acid moiety, without causing 
damage to the intestinal epithelium. The esters can be provided to the 
infant as a supplement to the infant's formula, or can be provided in an 
infant formula which also provides subirritant amounts of long-chain free 
fatty acids.

DETAILED DESCRIPTION OF THE INVENTION 
As indicated, one significant aspect of the present invention is food 
products, for providing sources of nutrition to the infant, that contain 
no free long-chain fatty acids or triglycerides which upon digestion are 
converted into free long-chain fatty acids. Another aspect is food 
products that provide long-chain fatty acids (per se and/or as 
triglycerides thereof which upon digestion yield long-chain fatty acids), 
but only in amounts insufficient to damage the intestinal epithelium of 
the infant. Preferably, the food products are essentially free of free 
fatty acids and of triglycerides thereof. 
The tendency of a given fatty acid to damage the intestinal epithelium, and 
the amount thereof that can be administered without causing damage to the 
intestinal epithelium, can be assessed with reference to the model 
described below in Example 1 which detects such damage. Preferably, the 
maximum amount of a given free long-chain fatty acid or triglyceride 
should be "subirritant", by which is meant zero up to an amount less than 
the amount of free fatty acid causing detectable damage. In general, the 
maximum tolerable content of aggregate free long-chain fatty acids 
provided in the food product will be less than about 5 mM, preferably less 
than about 1 mM. 
By "long-chain fatty acids" is meant physiologically acceptable compounds 
of the formula R-COOH wherein R is a straight or branched alkyl group, or 
an unsaturated or polyunsaturated alkylene chain containing up to 6 
carbon-carbon double bonds, and wherein R contains 15 to 21 carbon atoms 
(so that the fatty acids contain 16 to 22 carbon atoms including the 
carboxyl group). Examples of such long-chain fatty acids include palmitic, 
palmitoleic, stearic, oleic, linoleic, linolenic, eleostearic, gadoleic, 
arachidonic, behenic, cetoleic, 6,9,12,15-octadecatetraenoic, 
8,11,14,17-eicosatetraenoic, 5,8,11,14,17-eicosapentaenoic, 
7,10,13,16,19-docosapentaenoic, and 4,7,10,13,16,19docosahexaenoic acids. 
Other long-chain fatty acids known to be toxic, notably ricinoleic and 
erucic acids, would of course not be utilized. 
Triglycerides of long-chain fatty acids, as that term is used herein, are 
defined by the formula R.sub.1 -OCH.sub.2 -CH(OR.sub.2)-CH.sub.2 OR.sub.3 
wherein R.sub.1, R.sub.2, and R.sub.3 are the same or different and are 
each the acyl moiety R of C.sub.16 -C.sub.22 fatty acids as defined in the 
preceding paragraph. 
The present invention encompasses any food product adapted for providing, 
in whole or in part, nutritive value to an infant human or animal. Such 
food products thus include a protein source; or a carbohydrate source; or 
a vitamin source; or a source of medium-chain fatty acids, by which is 
meant fatty acids of the formula R.sup.1 -COOH wherein R.sup.1 is a 
straight or branched alkyl group, or an unsaturated or polyunsaturated 
alkylene group, containing 8 to 12 carbon atoms, e.g. capric acid; or a 
mineral source; or a combination of any two or more of such sources. The 
food product may also include one or more of the esters described more 
fully hereinbelow, alone or in combination with one, two or more of the 
protein, carbohydrate, vitamin, medium-chain fatty acid, and/or mineral 
sources. Preferably, the food product is a formula containing a protein 
source, a carbohydrate source, a vitamin source, a mineral source, a 
source of medium-chain fatty acids, and one or more of the esters 
described hereinbelow. 
The product may be a ready-to-feed liquid water-based preparation, or in 
the form of a powder or concentrated liquid adapted to provide a 
ready-to-feed form by the addition of water and stirring. The product 
preferably contains (when such is present) about 1.2 to 3.0 g, 
advantageously about 1.5 g, of protein; about 0.25 to about 2.0 g of 
medium-chain triglycerides; and about 6 to 9 g carbohydrate per 100 ml of 
the ready-to-feed liquid formula supplying preferably 60-75 kcal per 100 
ml. 
Suitable protein sources include casein, salts of casein (e.g., potassium 
caseinate), whey protein concentrate, soybean protein isolate, cow's milk 
protein, or hydrolyzed whey or casein protein. Cow's milk protein differs 
from that of human milk in the proportions present as casein and whey 
protein. Cow's milk has about 80% casein and 20% whey proteins, whereas 
human milk has about 40% casein and about 60% whey proteins. Accordingly, 
the protein used may be adapted to simulate that of human milk by 
supplementing cow's milk protein with an appropriate amount of whey 
protein. Because whey contains a very high proportion of the minerals of 
milk, the whey is subjected to demineralization, in particular by 
electrodialysis or ultrafiltration, to prepare whey protein. Where a 
milk-free diet for infants who are intolerant of cow's milk protein is 
desired, the protein source may be isolate soy protein or hydrolyzed 
casein or whey proteins. The proteins may be used in combination. 
As a carbohydrate source lactose is generally preferred in formulas of 
normal healthy infants. However, lactose would be contraindicated for 
infants suffering from galactosemia, lactose intolerance, or cow's milk 
protein intolerance. (In the latter case, the lactose may contain cow's 
milk protein.) Where a milk-free diet is desired, the carbohydrate source 
may be sucrose, corn syrup solids (glucose polymers), or a combination of 
corn syrup solids with sucrose. The carbohydrates may also be used in 
combination. 
As a source of medium-chain fatty acid, one may employ such fatty acids per 
se or triglycerides of three such fatty acids. 
Additionally, the food product or infant formula can be, or can contain, a 
nutritionally acceptable quantity of any one or more of the following 
minerals and vitamins: calcium, phosphorus, potassium, sodium, chloride, 
magnesium, iron, copper, zinc, manganese, iodine, and selenium; and 
vitamin A, vitamin D, vitamin E, vitamin K.sub.1, vitamin B.sub.1, vitamin 
B.sub.2, vitamin B.sub.6, vitamin B.sub.12, vitamin C, pantothenic acid, 
niacin, folic acid, biotin, choline and inositol. The food product could 
contain other nutritional factors, such as taurine, carnitine, or 
nucleotides. 
Suitable vitamins and minerals, any or all of which may be included in the 
infant food products of the present invention, and preferred amounts 
thereof to include (per 100 ml of infant food product), include the 
following 
______________________________________ 
Substance Amount 
______________________________________ 
Vitamin A 80-160 micrograms 
Vitamin C 20-60 milligrams 
Vitamin D 6-12 micrograms 
Vitamin E 8-16 milligrams 
Vitamin K 5-10 micrograms 
Vitamin B.sub.1 
80-200 micrograms 
Vitamin B.sub.2 
160-300 micrograms 
Vitamin B.sub.6 
85-160 micrograms 
Vitamin B.sub.12 
0.1-0.7 micrograms 
Niacinamide 0.8-2.0 milligrams 
Folic Acid 40-150 micrograms 
Pantothenic Acid 
400-800 micrograms 
Biotin 1-4 micrograms 
Sodium 20-60 milligrams 
Chloride 40-85 milligrams 
Calcium 50-80 milligrams 
Phosphorus 25-40 milligrams 
Zinc 820-1200 micrograms 
Iron 100-800 micrograms 
Copper 85-160 micrograms 
Taurine 2.5-10 milligrams 
Carnitine 1-5 milligrams 
______________________________________ 
An infant food product meeting the foregoing description can readily be 
prepared by standard measuring and mixing techniques well within the 
abilities of those familiar with the preparation of source products. It 
will be recognized that the amount of the food product that is 
administered to the infant at any one time will depend on the size, 
health, appetite and needs of the infant, and will generally comprise up 
to about 3 fluid ounces per feeding. 
Another significant aspect of the present invention is the discovery of a 
means by which fatty acids (as defined herein) can be provided in 
physiologically useful form to the infant, even though free long-chain 
fatty acids and triglycerides thereof are minimized or excluded from the 
infant's food as described herein. This discovery is significant since 
certain long-chain fatty acids, such as linoleic acid and linolenic acid, 
are believed to be dietary components essential to the proper development 
of the infant. 
More specifically, the fatty acid is provided enterally to the infant as an 
ester component which comprises a physiologically acceptable ester of the 
general formula RC(O)OE, wherein the ester component does not damage the 
intestinal epithelium of the infant. By "physiologically acceptable" is 
meant that the ester and the esterifying moiety E which cleaves from the 
ester do not adversely affect the health and development of the infant. 
Preferred "physiologically acceptable" esters will be those in which the 
esterifying moiety E is inert to the infant. It has been determined that 
fatty acid esters meeting this description can be absorbed by and do not 
damage the intestinal epithelium of the infant, yet provide the fatty acid 
in free form following absorption of the ester. The tendency, or lack of 
tendency, of any given ester to damage epithelium can be assessed by 
carrying out the procedure described in Example 1. 
The fatty acids whose esters are administered in accordance with this 
aspect of the invention are those wherein R is as defined hereinabove, and 
more preferably are those of the foregoing definition which are saturated, 
or are unsaturated or polyunsaturated omega-3, omega-6 or omega-9 fatty 
acids. (The terminology such as "omega-3" refers to the position of a 
carbon-carbon double bond with respect to the end of the molecule farthest 
from the --COOH group. For instance, linolenic acid is an omega-3 acid 
which contains 18 carbon atoms and is unsaturated at the 9, 12, and 15 
sites; the 15-site is by convention 3 atoms from the omega end of the 
molecule, hence, "omega-3"). Most preferred are linoleic, linolenic, and 
arachidonic acids. Other commonly found dietary fatty acids include oleic, 
palmitic and stearic acids. 
The esterifying moieties E include lower alkyl, by which is meant straight 
or branched alkyl containing 2 to 6 carbon atoms, the most preferred 
embodiment of which is ethyl. Indeed, ethyl esters of any fatty acid 
desired to be administered are most preferred. Also contemplated are 
glycerol phospholipid esters of inositol, ethanolamine, or choline, by 
which is meant compounds of the formula R.sub.4 OCH.sub.2 
-CH(OR.sub.5)-CH.sub.2 Op(O)-(X.sup.1) wherein X.sup.1 is inositol, 
ethanolamine or choline, and R.sub.4 and R.sub.5 are the same or different 
and each is the acyl moiety of a fatty acid as defined above for the 
moiety R. 
The ester or a mixture of such esters can be administered to the infant in 
a preparation containing no other nutritive component, or can be combined 
with a protein source; or with a carbohydrate source; or with a vitamin 
source; or with a source of medium-chain fatty acids; or with a mineral 
source; or with two or more of such sources. Suitable sources and amounts 
for the protein, carbohydrate, vitamins, and minerals include those 
described above with respect to this invention. Also, as described above, 
products containing the ester and any additional nutritive sources that 
are present can be prepared as powder, liquid concentrate, or as 
ready-to-feed liquid. Any such product should contain enough of the one or 
more esters that are present to provide the esterified fatty acid(s) in 
amounts up to an amount sufficient to satisfy the dietary needs of the 
infant, without providing an excessive amount which might for any reason 
be harmful. Generally, amounts aggregating a total of up to about 4.0 g, 
preferably about 1.5 to about 3.5 g, of all such esters per 100 ml of the 
product in its ready-to-feed form will be satisfactory. 
It will thus be appreciated that products in accordance with the present 
invention are particularly useful when administered to infants afflicted 
with damage to the intestinal epithelium, or to infants susceptible to 
such damage (such as premature and low-birthweight infants). However, 
products in accordance with this invention are for that very reason useful 
when administered to any infant. 
EXAMPLE 1 
This example describes the model employed for observation of intestinal 
epithelium damage, and demonstrates the lessening of damage upon perfusion 
of fatty acid ester. Mucosal changes in newborn piglet intestine before 
and after ischemia and reperfusion ("I/R") during luminal perfusion with 
infant formulas and human milk were evaluated. 
Methods: Plasma-to-lumen clearance of .sup.51 Cr EDTA (ml/min/100g), was 
measured in the jejuno-ileum of 1-day-old piglets for 1 hour each of 
control, ischemia, and reperfusion during luminal perfusion with 
formulas/milk which were predigested and bile acid solubilized. The 
perfusates consisted of term formula (Similac, Ross Laboratories, 
Columbus, Ohio) (3.6% lipid with primarily long-chain fats (LCT)), preterm 
formula (Special Care, Ross Laboratories, Columbus, Ohio) (3.6% lipid with 
50% medium-chain triglycerides (MCT) and 50% LCT), human milk (1% lipid 
with 70% LCT), and delipidated preterm formula. Gross and histological 
changes were evaluated after reperfusion. 
Results: EDTA clearance was markedly higher for preterm formula than for 
all other perfusates during control. Following I/R, clearances for preterm 
formula. were not significantly different from term formula, but were 
higher than delipidated formula (p&lt;0.05). Grossly, all segments perfused 
with preterm formula appeared gangrenous, while those perfused with term 
formula were variably hemorrhagic. Intestinal segments perfused with human 
milk or delipidated formula appeared normal. Histologically, the intestine 
perfused with preterm formula showed severe villous necrosis and 
hemorrhage, while that perfused with delipidated formula showed 
significantly less damage. 
Conclusions: The combination of luminal perfusion with infant formulas and 
I/R results in an animal model of acute phase necrotizing enterocolitis. 
It appears that removing the lipid from formula significantly attenuates 
mucosal injury. 
EXAMPLE 2 
To investigate whether esterification of oleic acid alters its effect on 
mucosal permeability, .sup.51 Cr-EDTA clearance (ml/min/100 g) during 
luminal perfusion with oleic acid was compared to that of its ethyl ester. 
Clearance was measured in the jejunum of 1-day-(n=7), 3-day-(n=5), 
2-week(n=6) and 1-mo-(n=6) old piglets after sequential perfusion with 
saline (20 min) followed by 5 mM solutions of either oleic acid or its 
ethyl ester (20 min). Mucosal permeability was significantly higher than 
control in all age groups when oleic acid was perfused (p&lt;0.05). In 
contrast, ethyl oleate did not cause an increase in clearance compared to 
control values. There was thus a marked difference in permeability 
observed between oleic acid and its alkyl ester for all age groups 
(p&lt;0.05). 
To determine whether the ester was absorbed and transported by the 
intestine, .sup.14 C-oleic acid and .sup.14 C-ethyl oleate (.sup.14 C 
incorporated into the acyl chain) were placed for 1 hr into loops of 
jejunum of 2-wk-old piglets (n=8 loops in each gp). Uptake was 69% and 57% 
for oleic acid and its ester, respectively, and .sup.14 C-radioactivity 
was detected in mucosa, lymph, blood, and liver. These studies indicate 
that the oleic acid-induced increases in intestinal permeability can be 
abolished when the carboxylic head of the fatty acid is esterified. This 
difference is not due to lack of absorption of the ethyl ester, as both 
long-chain fats are absorbed to a similar extent by the mucosa and 
transported to the systemic circulation.