Protein, Carbohydrate & Pigment Tests of Orange Juice Show Higher Nutritional Value in Natural vs. Unnatural Juice

By:

“The Spartans”

 Kelli Bigoness
Curtis Hunt
Chad Mcalvey
Berk Tanal

    The purpose of our research was to determine if the nutritional value of orange juice would decrease with regard to carbohydrate and protein content as two variables changed:  one, if the juice received more pulp filtration, and secondly, if it contained chemical and flavor additives.  It was determined that as pulp was removed from the juice, there was no effect on sugars present and protein concentration increased.  Data also indicated that accumulation of chemical and flavor additives decreased protein concentration.
                Three types of orange juice, home produced, Tropicana Pulpless, and Sunny Delight, were analyzed.  Each of the juices was subjected to four carbohydrate tests; Benedict’s, Barfoed’s, Selivanoff’s, and Bial’s, which indicated that homemade and Tropicana orange juice contained complex sugars and furanose rings.  Sunny D indicated the presence of many commercial sugar additives, including high fructose corn syrup.
Utilizing paper chromatography, we determined that none of the juices contained any of the four photosynthetic pigments. Through the Bradford Assay, we verified that Tropicana juice contained the highest concentration of protein at 2.13ug/ul.  Home produced juice contained less, and the Sunny D had the smallest level of protein at 0.035ug/ul.
                It was concluded that Tropicana juice had superior nutritional content, because it contained complex sugars, furanose rings, and had the highest protein content.  This is a result of this juice being 100% natural.  Since we found no photosynthetic pigments in any of the juices, no juice held a nutritional advantage over another with regard to pigments.

Tables

Table 4.  Absorbance reading and Protein Concentration in Three Orange Juice Samples.  Absorbance data was collected from each of the three orange juice samples.  That data was then compared to the BSA standard curve to determine the concentration of protein in each of our orange juices.

Discussion:

In our experiment, we set out to analyze the differences in carbohydrates, photosynthetic pigments, and protein concentration present in three different types of orange juice: home produced, Tropicana Pure Premium 100% pure Florida orange juice, and Sunny Delight.  From our research on nutrition, we concluded that the most nutritious energy providing carbohydrates were complex sugars.  Also, the higher the protein level in the orange juice, the more nutritious it would be (Spaaij and Pijls, 2004).  Our research determined if the nutritional value of orange juice would decrease with regard to carbohydrate and protein content as two variables changed.  One, if the juice received more pulp filtration, and secondly, if chemical and flavor additives increased.  Our hypothesis was that the hand-squeezed orange juice would be most nutritious, thus containing the most complex sugars and highest protein level.  We predicted that our carbohydrate tests would reveal the hand-squeezed orange juice to contain the most complex sugars and furanose rings, and that Sunny Delight would contain the most monosaccharides and few complex sugars.  In our pigmentation tests, we predicted that the home produced juice would have the greatest number of photosynthetic pigments, the Tropicana would contain fewer, and the Sunny Delight would contain no pigments, due to its lack of natural materials.  For our protein testing, we predicted that hand-squeezed orange juice would contain the highest protein level, followed by the Tropicana, while Sunny Delight would have the least.

 Carbohydrate Analysis:

            The first test we performed for carbohydrates was the Benedict’s test, which indicates the presence of reducing sugars.  All three of our samples tested positive with an orange/red precipitate.  This allowed us to conclude that there were free aldehyde and ketone groups in all three juices.  This data supported our original hypothesis, as it indicates that there may be mono, di, or polysaccharides present in the juices. Further test were needed to isolate which specific sugars were present.  No conclusions about the nutritional and energy value of the juices can be drawn form this test, because all three juices tested similarly.
            In Barfoed’s test, which indicates the presence of monosaccharide sugars, we found that all three tested positive with a precipitate.  Notably, the three test tubes containing Sunny D contained a larger amount of precipitate.  This may indicate a larger presence of monosaccharides in the Sunny D relative to the other juices. This data allowed us to conclude that all three juices contained monosaccharide sugars, and that Sunny D most likely contained a larger quantity than the home produced and Tropicana orange juices.  Our original hypothesis predicted that we would only find monosaccharides in the Sunny Delight.  The test data partially supports our hypothesis, as we expected to see monosaccharides in the Sunny D, but rejects the portion that predicted the absence of monosaccharides in the home produced and Tropicana juices.  From this test alone, Sunny D has an energy advantage over the other juices due to its higher sugar content, although to it is not possible to determine to what extent from this test.
            Selivanoff’s test allowed us to further isolate the types of sugars present in our juices.  All three juices changed color in less than one minute, leading us to conclude that they all contained monosaccharide ketoses.  Our original hypothesis predicted that these sugars would only be present in the Sunny D.  One reason that they all may have tested positive is that all three juices contain some level of natural juice, thus creating commonalities between the three.  No nutritional differences are indicated from this test, as they all tested positive for the same types of sugars.
            The last carbohydrate test, Bial’s, allowed us to determine what types of furanose rings were present in the juices.  Both the home produced and Tropicana juices produced a muddy-brown precipitate, indicating the presence of hexose-furanose rings.  The Sunny Delight also tested positive, but it was significantly darker and therefore contained a generally larger amount of hexose-furanose rings. The difference between the Sunny Delight and the others is most likely attributed to a sweetener added to some artificial juices called High Fructose Corn Syrup, which is a hexose-furanose.  This data rejects our original hypothesis, as it predicted that only the home produced and Tropicana juices would contain complex sugars and rings.  In actuality, the Sunny D contained a generally larger amount of high-energy complex sugars and rings.
            After all of the carbohydrate tests were completed, no solid conclusions about how pulp filtration and the addition of artificial flavors affect the nutrition and energy value of the juices could be made.   All four tests produced the same general results for all trials and juices.  We were able to conclude that the Sunny D had a generally larger amount of monosaccharide sugars, and hexose-furanose rings due to the larger amounts of precipitate formed in the tests.  This is most likely due to the addition of commercial sweeteners and artificial flavoring.  However, since these tests are qualitative, no definite conclusions about the quantity of sugar present could be made.  These results can be attributed to all three of the juices containing at least a small portion of natural juice, causing the qualitative carbohydrate analysis to show similar results.

Photosynthesis Analysis:

To test for photosynthetic pigments, we performed paper chromatography which separates the pigments present in a fluid.  All three of the trials for each juice showed no bands.  From these results, we determined that there are no photosynthetic pigments present in orange juice, and that variation of pulp filtration and the addition of artificial flavors had no effect on the presence of the pigments.  This is most likely due to the fact that orange juice is extracted from a part of the plant that does not undergo photosynthesis.  These findings go against our original hypothesis which predicted a reduction in photosynthetic pigments as pulp was removed and artificial flavors were added.  From this data, it is not possible to draw any conclusions regarding the nutritional and energy differences between the juices.
            During the experiment, the home produced orange juice blotted poorly onto the paper strips due to the excessive amount of pulp.  This may have caused some error in the experiment, and a more liquefied solution may have reduced this problem.

 Protein Analysis:

To conclude our analysis on orange juice, we used the Bradford Assay to test for protein concentration.  This test produced our most significant results and conclusions.  From this test, we determined that Tropicana orange juice contained the highest protein level at and average of 2.13ug/ul, followed by the home produced with a level of 1.44ug/ul, while Sunny Delight contained almost no protein at 0.035ug/ul.  This data allowed us to conclude that our original hypothesis was correct, and that the addition of artificial flavoring and other additives decreases protein concentration in orange juice.  This point is illustrated when average protein concentration dropped to near zero when Sunny D was tested as opposed to the relatively high test results for the two 100% natural juices.
            The hypothesis that as pulp filtration increased protein concentration would decrease was not supported by this data.  The test results show that protein concentration actually increased from an average of 1.44ug/ul in the home produced juice, to 2.13ug/ul in the pulp filtered Tropicana.  It is possible that a more homogeneous home produced orange juice solution would allow more protein from the pulp to be released into the solution.  We believe the solution we used may have been too heterogeneous, and that protein from the pulp didn’t react with the Bradford reagent, causing a lower protein concentration for the home produced juice.  This is the only conclusion we can come to, as past research has shown high concentrations of protein are found on the pulps of fruits (Araujo et al., 2003).
            Since our research led us to conclude that a higher protein level is most nutritious, we were able to conclude that the Tropicana orange juice offered the highest nutritional value (Spaaij and Pijls, 2004).  Our data suggests that this is due to it being 100% natural juice with no flavor additives.

 Conclusion:

            After analyzing the data to all of our tests, we determined that part two of our hypothesis was correct and supported by our experiments.  As chemical and flavor additives increased, nutritional value decreased.  While the carbohydrate and photosynthetic pigment tests were largely inconclusive, the Bradford Assay produced enlightening results.  The test showed that as chemical and flavor additives increased, protein concentration decreased dramatically.
            From a nutritional standpoint, the Tropicana Pure Premium 100% Natural Orange Juice had the superior overall nutritional content.  It contained the highest level of protein, and comparable types of simple and complex sugars.  The home produced orange juice was a close second, containing significant quantities of protein, and comparable sugar content.  While the Sunny D had a generally larger quantity of sugar, which provides the juice with a higher energy punch, this is not enough to outweigh its near total lack of protein.  The conclusion of this study is that consumers should buy 100% natural orange juice for its superior nutritional benefits.