A comparison of soy milk and rice milk as a substitute for cow's milk

by testing the carbohydrate, protein, amino acid, and lipid content.

By: Liz Vanderlaan

Stephen Peretin

Lindsay Wieber

Kyle Kutsche

 

 

 

Abstract

By: Stephen Peretin, Revised by Lindsay Wieber and Liz Vanderlaan

Many medical altercations such as lactose intolerance and other milk allergies call for the use of a nutritionally sufficient milk substitute. In order to determine the best substitute for natural cowâs milk an array of tests were used to compare the carbohydrate, protein, amino acid, and lipid content of cowâs milk, rice milk, and soy milk. It was hypothesized that soy milk will have the closest results to that of cowâs milk, making it the best milk substitute.ÊÊ

The first portion of these tests to determine whether rice or soy milk is the best alternative involved examining the carbohydrate content. Barfoedâs test was used to distinguish monosaccharides from disaccharides and polysaccharides. Selivanoffâs test was used to differentiate between ketoses and aldoses. The Iodine test was used to distinguish starch from other sugars. A Bradford assay was used to determine the amount of protein, in μg, present in each milk sample. A standard curve was made using BSA to determine the amount of protein in each milk sample. The method of finding fat or lipid content in each milk sample using a spectrophotometer was employed. The test used to determine amino acid content was a TLC test or Thin Layer Chromatography. In regards to Barfoedâs test the rice milk was the only sample found to contain monosaccharides. Selivanoffâs test showed that there were ketoses present in soy and rice milk, but aldoses present in cowâs milk, which confirms that, neither soy or rice milk contains lactose, but cowâs milk does.Ê The Iodine test for starch indicated that soy was the only milk containing starch. The Bradford assay confirmed the assumption that soy milk contained the largest amount of protein of all the three types of milk tested followed by cowâs milk and then rice milk. It was confirmed that soy milk has the closest amount of fat to that of cowâs milk, but the amount was significantly higher than expected, considering soy beans are 20% fat (Murano, 2004).Ê Cowâs milk was predicted to show the most positive result for amino acids because it contains all eight essential amino acids followed by soy milk and then rice milk, but due to poor experimental design produced weak results in this portion.Ê They indicated that soy milk contained the most amino acids, and cowâs milk only contained three amino acids, one of which was essential.Ê The hypothesis that soy milk would have the closest results to that of cowâs milk, in terms of carbohydrates, protein, amino acids, and lipids was confirmed by these four tests, making it the most qualified substitute for those with lactose intolerance and other cowâs milk allergies.

 

 

Discussion

By Lindsay Wieber, Liz Vanderlaan, and Kyle Kutchse, Revised by Steve Peretin

Lactose intolerance is a widespread issue as it affects 30 to 50 million people nationwide.Ê A deficiency in the lactase enzyme causes an inability to break down lactose into its simpler forms of glucose and galactose (Vonk and Priebe, 2003).Ê This lactase shortage creates numerous digestive problems, including cramps, bloating, nausea, and diarrhea about 1-2 hours after consuming significant amounts of lactose.Ê Lactose is the most prominent sugar in milk, posing a dilemma for millions of people.Ê Milk is one of the main components of the dairy group, which the food guide pyramid recommends 2-3 cups per day (Horvath, 2005).Ê It provides essential nutrients to the body, and is particularly known for its calcium, vitamin A and D, and protein.Ê Simply eliminating milk from lactose intolerant individualsâ diet would sacrifice a healthy, balanced diet.Ê To avoid this consequence many people are led to alternative milk sources which may have questionable nutritional values.Ê Milk allergies, like lactose intolerance are also prevalent in infants, forcing them to consume an alternative formula.Ê Seeing that infants donât consume solid foods until they are 4-7 months old it is vital that their milk formula has adequate amounts of nutrients (Dowshen and Rutherford, 2001).Ê

ÊÊÊÊÊÊÊÊÊÊÊ This experiment focused on two popular milk alternatives; soy milk and rice milk.Ê These milks were analyzed based on their carbohydrate, protein, amino acid, and lipid content and compared to 2% cowâs milk.Ê The best milk substitute was determined by the milk that compared the closest to the cowâs milk.Ê It was hypothesized that soy milk would produce the most similar results to those of cowâs milk.Ê More specifically, the sugar tests were predicted to confirm that cowâs milk contains lactose, a disaccharide composed of glucose and galactose.Ê Soy milk was predicted to contain more complex sugars while rice milk was expected to contain simple sugars, but neither were thought to have lactose.Ê In particular soy milk was expected to test positively for sucrose, as it is a sugar in certain soy milks that has a negative effect on teeth (Davison and Simmer, 1997).Ê It was also expected that soy milk would contain the highest amount of protein because the soy bean is 40 % protein (Murano, 2004).Ê Typically, foods that are quality sources of protein can attribute this benefit their amino acid content, leading to the prediction that soy milk would have more essential amino acids than rice milk (Anonymous-4, 2004).Ê For the lipid test, it was predicted that soy milk would be much closer to the fat content of cows milk because the soy bean is 20 % fat (Murano, 2004).Ê One of the popular aspects of rice milk is its low fat content, leading to the prediction that it will not even compare to the amount of fat in cows milk.Ê The four aspects; carbohydrate, protein, amino acids, and lipids, gave a general idea of which alternative source of milk is the best alternative. It was predicted that soy milk would produce the most similar results to cowâs milk, therefore making it the best option.Ê After many experiments this hypothesis was confirmed as soy milk had very comparable results to cowâs milk and rice milk was distinctive and not similar to cowâs milk.Ê

The main sugar in cowâs milk is lactose, also known as the ãmilk sugarä.Ê It is a disaccharide characterized by the linkage of two aldoses, glucose and galactose (Freeman 2005).Ê The carbohydrate portion of this research confirmed the presence of lactose in cowâs milk.Ê Barfoedâs test determined that it was a di- or polysaccharide and Selivanoffs test determined that it contained aldoses.Ê This is agreeable with the structure of lactose, as both glucose and galactose are aldoses.

It was predicted that soy milk would contain complex sugars, especially sucrose, and that there would be an absence of lactose.Ê Previous research suggests that certain soy milk formulas consist of maltodextrins, corn starch (isomaltose) or sucrose (Davison and Simmer, 1998).Ê The results of this experiment, however, did not support this prediction.Ê Sucrose is a disaccharide characterized by the glycosidic linkage of one molecule of glucose and one molecule of fructose.Ê Disaccharides containing fructose should react intermediately between that of fructose alone and one of the aldoses in Selivanoffâs test (Khra 2006).Ê The soy milk samples in this experiment showed evidence of di- or polysaccharides, the presence of an aldose and the presence of a starch.Ê The existence of a di- or polysaccharide was consistent with the sugars described in previous experiments, and consistent with the indication of starch, a coiled polymer of glucose.Ê

It was predicted that rice milk would contain simple sugars which provide the body with a short surge of energy but then the sugar turns to fat (Hart, unknown).Ê Simple sugars, or monosaccharides, include glucose, fructose, and galactose.Ê The series of sugar tests performed on rice milk generated results that indicated the presence of a monosaccharide as well as an aldose.Ê It did not, however, show evidence of starch.Ê These results lead to the fact that soy milk and cowâs milk provide the body with complex carbohydrates which take longer to digest.Ê Complex carbohydrates are useful in supplying the body with long term energy, while simple carbohydrates will not maintain sufficient energy levels.Ê It is advised to avoid getting the bulk of your carbohydrates from refined foods, which contain simple carbohydrates because they are typically low in nutrients and energy (Grieger, unknown).Ê This leads to the fact that soy milk is not only the closest to cowâs milk in terms of sugars, but it is providing the body with the most efficient and more recommended sugar.Ê

Another crucial aspect of nutrition to consider is the protein content.Ê Proteins are vital in maintaining the structure of red blood cells, enzyme regulation, proper function of antibodies, and most commonly known for their aiding in growth and repair of body tissues (Anonymous-8, unknown).Ê It was predicted that soy milk would contain more protein than cowâs milk or rice milk.Ê Soy milkâs main economic and nutritional advantage is the soy beanâs rich protein content of 40 percent (Murano, 2004).Ê It is important to understand, however, that soy protein contains only one-third of available nitrogen as essential or semi-essential amino acids, and therefore has a lower biological value than milk protein (Davison and Simmer, 1997).Ê In general, animal proteins are considered higher quality than plant proteins.Ê The results of this experiment confirmed the predictions, indicating that soy milk does in fact contain the highest amount of protein at 124.218 mg/ml.Ê It was followed closely by 2% milk with 109.731 mg/ml.Ê Rice milk was found to contain dramatically lower amounts of protein than both soy and cowâs milk with a value of 6.059 mg/ml.Ê It should be noted, however, that previous research describes rice as relatively rich in energy and a good source of protein, with the highest protein digestibility among staple grains (Fiocchi 2003).Ê Though soy milk was found to have a greater amount of proteins than both cowâs milk and rice milk, the protein content of cowâs milk is considered to be ideal due to the essential amino acids responsible for the composition of proteins.Ê

A third key aspect that was analyzed directly relates to the protein content.Ê Amino acids are the building blocks of protein, meaning that a milk source high in protein should contain many amino acids.Ê After calculating the Rf values for each of the amino acid bands, the values were compared to the corresponding amino acid.Ê It was difficult to determine the amino acids present because each trial for each sample varied slightly.Ê It was determined that 2% milk contained cystine, lysine, and histidine.Ê Soy milk contained arginine, alanine, valine, and tyrosine.Ê Rice milk was found to contain alanine and tyrosine.ÊÊ There were other amino acids found that occurred in only a single trial that were not included due inconsistency between the trials.Ê This could mean that the amino acid was in fact present in the milk, but in very low concentrations, making its presence on the TLC vary.Ê It was also difficult to distinguish between the many bands on the TLC strip and it could have been very close to another band, causing them to blend together.Ê

ÊÊÊÊÊÊÊÊÊÊÊ Despite the problems with the results of this test, they did support the hypothesis that soy milk is most similar to cowâs milk.Ê This was determined from several conclusions.Ê First, soy and 2% milks each showed signs of essential amino acids.Ê Essential amino acids were those that were needed in the body but could not be synthesized by the body.Ê Therefore, essential amino acids must be included in the diet.Ê There are eight essential amino acids for humans (Anonymous-8, 2006).Ê According to this test, soy milk had one of these, valine.Ê Two percent milk also had one essential amino acid, lysine.Ê Histidine, another amino acid in 2% milk, is essential for children only.Ê Rice milk produced no essential amino acids, meaning that all 8 essential amino acids would have to be obtained from another food source because the body cannot produce them.Ê

ÊÊÊÊÊÊÊÊÊÊÊ More evidence that soy milk was closer nutritionally to cowâs milk came from previous research.Ê The results collected through this lab were not complete, yet they compared well to the research of others.Ê In a 2002 article, Tom Overton discussed research that compared the effects of amino acids on protein content in dairy milk.Ê This study concluded that a balance of essential amino acids in a cowâs diet increased protein content in cow milk (Overton, 2002).Ê This linkage of amino acids and proteins made sense in that proteins are composed of amino acids.Ê When comparing the amino acid content from the TLC to the protein content from the Bradford assay, it was seen that the protein is highest in soy milk as is the amino acid content.Ê These results from the amino acid TLC were inconclusive in part, yet they confirmed the hypothesis that soy milk is higher in amino acid content as cowâs milk should be.Ê

The lipid content of each milk was analyzed in a similar fashion to the protein assay.Ê A standard curve was constructed, but the absorbances of the milk samples were too low to fit the curve, therefore the lipid content of each milk sample could not be quantified.Ê Instead, the absorbance values were compared to each other to give a comparative analysis of the lipid content.Ê This was a relatively good indicator of the lipid content because the standard curve followed a linear trend, indicating that the amount of fat is directly proportional to absorbance.ÊÊ Soy milk was found to have the highest average fat content, with an absorbance of 1.021.Ê The average absorbance for cowâs 2 percent milk was reported as 0.377, which is significantly lower than soy milk.Ê As expected, the skim milk registered the lowest at 0.188.ÊÊ Rice milk reported an average absorbance of 0.279, which is concordant with the suggestion that rice milk is low in fat compared to soy and cowâs milk.Ê Previous experiments report that rice milk contains only one fourth as much fat as cowâs milk (Gilbert, unknown).Ê The results found in this experiment do not indicate it to be one fourth the absorbance of cowâs milk; however, they do show that a much lower absorbance value was found.Ê Another recent study in which the fatty acid composition of cowâs milk substitutes were compared to breast milk reports that, although the substitute formulas were found to be highly variable, the lipid values acquired for soy milk were much lower than that of human milk (Kaila 1999).Ê The absorbance for soy milk was significantly higher than that of cowâs milk, when in reality they should have had nearly the same absorbance.Ê This is likely attributed to some reaction that occurred with the soy milk and ethanol to produce a slightly darker, less transparent solution.Ê

ÊÊÊÊÊÊÊÊÊÊÊ The four experimental processes all involved a slight bit of error, which was mostly accounted for when analyzing the results.Ê The sugar tests ran smoothly with the exception of the difficulty of distinguishing between an intermediate, positive, or negative result.Ê This was mostly due to the translucent milk, which made the solutions a slightly different color than the controls which were transparent.Ê The Bradford protein analysis was carried out without much trouble, but there was potential for error when using the spectrophotometer.Ê Fingerprints on the cuvette or dirty cuvettes could have caused absorbance values to be inflated, creating either an inaccurate dilution curve or inaccurate values for the quantification of protein.Ê Again, this test ran smoothly and the R-squared value for the line was near 1, indicating that the Beer-Lambert plot is very precise.Ê

ÊÊÊÊÊÊÊÊÊÊÊ ÊThe experiments became more complicated with the thin-layer chromatography amino acid test.Ê It took numerous attempts to get just three amino acid bands for the 2% milk.Ê This could be attributed to many factors.Ê It took a long period of time (about two and a half hours) for the solvent front to rise to a point on the strip in which the amino acids could separate enough to distinguish.Ê Had the strips developed for a longer period of time, the results may have been more accurate.Ê Another problem with this test was in the dilutions of the milk samples.Ê The 2% milk inhibited the solvent front from moving far enough up the strip.Ê Multiple dilutions were performed with little success.Ê If this part of the experiment were to be repeated the appropriate dilution for 2% milk would need to be determined in order to obtain working results.Ê

The lipid test also had much room for error as the soy milk produced values that didnât correspond with the label.Ê The soy milk concentration was significantly higher than the 2 % cowâs milk, when in reality they should have been relatively close values.Ê This is likely due to the fact that the soy milk reacted with the ethanol, causing the absorption to increase.Ê Again, with the use of the spectrophotometer, fingerprints on the cuvette or dirty cuvettes can cause skewed absorbance data.Ê A second dilemma with the lipid test occurred when analyzing the results.Ê A poor experimental design when creating the standard curve caused the dilutions of the standard curve to be too different from the dilution of the ethanol and milk samples to compare.Ê This made it impossible to quantify the amount of fat in the milks; instead the absorbance values were compared to give a general idea of which milk contains the most fat.Ê

Based on the results of the experiments performed, soy milk can be considered the best alternative to cowâs milk.Ê This conclusion isnât an indicator of how healthy soy milk, but an indicator of how it compares to cowâs milk by composition.Ê The results verify the hypothesis that soy milk is the best lactose-free alternative to cowâs milk based on its similarities in carbohydrates, proteins, amino acids and lipids.Ê It should be taken into consideration, however, that previous research has found that up to 40 percent of infants allergic to cowâs milk may also develop soy protein allergy (Davison and Simmer 1997).Ê Although soy proteins appear to be less sensitizing than cowâs milk proteins there is still a risk that an individual experiencing an allergic reaction to the proteins found in milk might experience a similar situation with soy milk as a dairy alternative (Murano 2001).Ê

This research gives a microscopic view of the components of cow milk, rice milk, and soy milk, leaving the opportunity for further studies.Ê Many other aspects of the milks could be explored to further compare them.Ê Relating to the soy protein allergy, it could be determined what quantity of protein is causing the allergic reaction in many infants, and how their substitute formulas compare to cow and soy milk.Ê A quantitative vitamin analysis could be performed to confirm the vitamin content listed on the labels.Ê Another interesting aspect pertaining to the benefits of soy milk could involve testing for isoflavones.Ê Isoflavones are in soy beans and are thought to effect estrogen levels and act as an anti-oxidant (Messina, 2002).Ê Research indicates that cowâs milk and rice milk do not contain isoflavones, but it would be interesting to see how significant their presence is in soy milk.Ê A fourth component that could be explored is the type of fat present in each milk.Ê There are many types of fat, including polyunsaturated, monounsaturated, saturated, cholesterol, triglycerides, and trans-fatty acids, all which have different effects on the body.Ê Both polyunsaturated and monounsaturated are known as ãgood fatsä as they lower LDL cholesterol and raise HDL cholesterol.Ê Saturated fat and cholesterol are common in milk, and raise LDL cholesterol, in turn increasing the risk for heart disease (Anonymous-7, unknown).ÊÊ It is recommended that 20-35% or less of daily food calories are fat, which could be compared to the amount of fat per serving and the recommended 2-3 servings of dairy per day (Horvath, 2005).Ê Another very interesting aspect that could be studied is how the processing of each type of milk impacts its nutritional benefits.Ê For instance, the soy bean could be compared to the actual soy milk product, in terms of type of fats, amount of protein, and amino acids.Ê All of these studies would only further the knowledge of the composition of cowâs, rice, and soy milk, therefore giving a good understanding of the benefits and pitfalls of using a particular alternative source of milk.Ê

 

 

 

 

Figures

 

Figure 1: Bar graph of average protein amounts found in three different types of milk using the Bradford Assay. Fifty μl of each milk sample was aliquoted into glass tubes.Ê Fifty μl of NaOH was then added to each and vortexed.Ê Three ml Bradford reagent was then added and the complete solutions were placed in the cuvettes and read at 595 nm in a spectrophotometer.Ê Using Microsoft Excel, a bar graph was created using the average value of proteins found for each milk: 2% milk with 109.731, soy with 124.218, and rice with 6.059.Ê According to the Bradford assay performed, it is obvious that soy milk contains the highest amount of protein followed by cowâs milk.Ê Rice milk contained dramatically lower amounts of protein than both soy and cows milk.

 

 

 

 

 

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