Effects of Boiling and Processing Treatments on Protein Concentration of Broccoli

Emily Bow, *information deleted for confidentiality*

Abstract
The purpose of this experiment was to determine if cooking and processing broccoli have an effect on its protein concentration, and if so, decide the difference in protein concentration between the samples. To do this, we used Bradfords assay for protein concentration (quantitative). By performing serial dilutions of the different broccoli samples, we were able to determine the amount of protein concentration by running the samples through a spectrophotometer. As a qualitative aspect, we denoted the color changes of each broccoli sample. After determining the best fit dilution as the 75% broccoli solution, we ran more trials through the spectrophotometer. We ran a standard curve which we then compared to our 75% broccoli solution absorbances. From this, we were able to conclude that there are 1.102 x 10-5 g of protein per 1 g frozen broccoli; 9.266 x 10-5 g of protein per 1 g raw broccoli; and 3.750 x 10-5 g of protein per 1 g boiled broccoli. This concluded a strong support for our hypothesis - raw broccoli had the highest protein concentration, and the processes of boiling and cooking decreased protein concentration.
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Section 30, *information deleted* BS 171: Fall 2012 Michigan State University

Results continued

Table 1. Mean protein concentration, standard error, concentration per one gram of broccoli, and color change for frozen, raw, and boiled samples.

Results
Absorbance vs. Protein Concentra ons y = 0.1351x - 0.0031
R = 0.97095 0.25 0.2

Conclusions
During our trial runs, we denoted the color changes for frozen, boiled, and raw broccoli (see Table 1). From these colors, we inferred that raw contained the most protein concentration, followed by boiled broccoli and frozen broccoli. From the recorded absorbances of our trials, we concluded that as the percentage of broccoli increases, the absorbance (protein concentration) also increases. Also, protein concentration decreases from raw to boiled to frozen. Once we completed our trials, we decided to use the 75% broccoli solutions. We calculated the means (absorbances) of the 75% solutions to be the following: 0.067 (frozen), 0.563 (raw), and 0.228 (boiled). We compared these means to our standard curve (Figure 1) to determine the amount of protein concentration per 45 g of broccoli. Our results are the following: 0.4959 mg/ml (frozen), 4.1697 mg/ml (raw), 1.6876 (boiled). This is shown in Table 1. From these results, we can conclude the grams of protein per one gram of broccoli: 1.102 x 10-5 g (frozen), 9.266 x 10-5 g (raw), 3.750 x 10-5 g (boiled). This is shown in Table 1. Figure 2 contains the standard errors for the 75% frozen, raw, and boiled broccoli solutions. None of the three error bars overlap with one another. From this, we can conclude that raw broccoli has significantly more protein than boiled and frozen; and boiled broccoli has significantly more protein than frozen. Like stated in our hypothesis, raw in fact had the highest protein concentration compared to frozen and boiled; and boiling and freezing (processing) the broccoli ultimately decreased protein concentration. This experiment has opened up other questions. We propose the following question: Does this trend exist with other nutrients such as carbohydrates, fat, fiber, etc. in other foods other than broccoli?

Introduction
Broccoli contains protein, a biomolecule necessary for all life. This experiment focuses on determining whether cooking and/or processing (freezing) broccoli have an effect on the amount of protein present. The three forms of broccoli we used are the following: 45 g of raw broccoli 45 g of boiled broccoli (cooked for 8 minutes) 45 g of Freshlike Broccoli Cuts (processed/frozen)
Absorbance

0.15 0.1 0.05 0

We hypothesize that raw broccoli will contain the highest protein concentration compared to the processed and boiled broccoli. We also believe that both cooking and processing the broccoli will cause a decrease in protein concentration.

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Protein Concentra on (mg/ml)

Figure 1. Standard curve of protein concentration and absorbance (Bradfords assay).

Methods
We prepared our broccoli samples (raw, boiled, and processed). For each broccoli sample dilution, we started out with a 75% broccoli solution made from 45 g of broccoli and 60 ml of distilled water. We performed three serial dilutions which resulted in the following percent broccoli solutions: 75%, 38%, 19%, 9.3%, 4.7%, 2.3%, and 1.2%. After transferring our dilutions to designated cuvettes and blanking the spectrophotometer (595 nm), we ran the dilutions and determined their absorbances. We decided the 75% solutions gave the best absorbance reading. We then ran three 75% broccoli solutions for each broccoli samples (raw, frozen, and boiled) through the spectrophotometer. With the average absorbances, we were able to conclude the amount of protein in the broccoli samples using our standard curve of protein concentration.

Protein Concentra ons of 75% Broccoli Solu ons

4.5 4 3.5 3 2.5 2 1.5 1 0.5 0 FROZEN RAW BOILED

References
Bian et al, (2007) Journal of Experimental Botany. Vol. 48. No. 308. Gaiser et al, (2003) Plant Physiology. Vol. 103. Pages 413-419. Islek et al, (2009) Journal of Science. Vol. 23. Pages 1-6. Kimberly Yowell and William H. Flurkey. (2006) Journal of Food Science. Vol. 51. Issue 2. Urbance, John. (2012). BS 171 Lab Manual. Pages 6-10.

Figure 2. Mean protein concentration for a 75% broccoli solution for frozen, raw, and boiled varieties. S.E. shown.