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Cindy Chen

November 20, 2014


Thursday AM
AA
SK
Lab 5- Inactivation of Enzymes By Blanching
I.

PURPOSE/OBJECTIVE:

The purpose is to understand protein denaturation and the concept of blanching. It is also important to
understand why blanching is important in the food industry. Another purpose is to understand blanching
efficiency, which is the rate of denaturation of peroxidase. Lastly, it is important to understand and to be
able to apply rate laws to determine the kinetics of blanching.

II.

INTRODUCTION:

The importance of this experiment is to understand how to enzymes can be inactivated by blanching and
the effects of blanching. It is also important to know how to calculate the blanching efficiency because it
is important in the food industry. In a clinical nutritionist's point of view, blanching can retain the
nutrition inside the food and be able to keep fresh foods for a longer period of time. In a food scientist's
point of view, it is important to understand that microbial growth is a big problem in food and blanching
helps reduce the possibility for that to occur. Also, enzymes in vegetables help them grow and mature
which contributes to the sensory aspect.

III.

PROCEDURE:

The procedure followed for the experiment is found in Principles of Food Composition Laboratory
Manual (2013) Experiment 5, Inactivation of Enzymes, pages 50-56. Modifications include: 1. adding
125 microliters of turnip extract instead of 100 microliters, 2. absorbance readings for every 5 seconds for
the first minute and then 90 and 120 seconds instead of readings at every 30 seconds, and 3. assume that
absorbance at time 0 is 0.

IV.

DATA:

Table 1: The absorbances (a.u.) and times (min) (at which they were recorded) for each of the blanched
samples (0, 15, 30, 60, 90, 120 sec).
Time (min)

Abs (120 s

Abs (90s

Abs (60 s

Abs (30 s

Abs (15 s

Abs (0 s

blanch time)

blanch time)

blanch time)

blanch time)

blanch time)

blanch time)

0.000

0.000

0.000

0.000

0.000

0 0.000

0.083 0.011

0.014

0.015

0.139

0.220

0.280

0.17 0.011

0.014

0.015

0.199

0.312

0.390

0.25 0.011

0.013

0.015

0.250

0.392

0.501

0.33 0.011

0.014

0.015

0.300

0.472

0.592

0.42 0.010

0.013

0.016

0.352

0.552

0.694

0.5 0.010

0.013

0.015

0.396

0.616

0.756

0.58 0.010

0.013

0.015

0.450

0.676

0.806

0.67 0.010

0.013

0.016

0.488

0.716

0.840

0.75 0.010

0.013

0.016

0.568

none

0.855

0.83 0.010

0.013

0.017

0.596

0.770

0.875

0.92 0.010

0.013

0.017

0.620

0.790

0.885

1 0.010

0.013

0.019

none

0.798

0.895

1.5 0.010

0.012

0.029

0.680

0.825

0.925

2 0.016

0.013

0.044

0.692

0.850

0.945

Table 2: The slopes (a.u./min) from each line on figure 1. Also included the enzyme activities (EAU) and
the ln of the EAU's.

Blanch Time

Slope

Enzyme

(sec)

(au/min)

activity

ln (EAU)

(EAU)
120

0.0089

8.9

2.186

90

0.0121

12.1

2.493

60

0.0155

15.5

2.741

30

0.7091

709.1

6.564

15

1.0776

1077.6

6.982

1.2950

1295.0

7.166

Fig. 1a. Absorbance vs Times for All Blanched Samples


1
0.9
A
b
s
o
r
b
a
n
c
e

Abs (120 s blanch


time)
Abs (90s blanch
time)
Abs (60 s blanch
time)
Abs (30 s blanch
time)
Abs (15 s blanch
time)
Abs (0 s blanch
time)

0.8
0.7
0.6
0.5
0.4
0.3
0.2
0.1
0
0

0.5

1.5

2.5

Time (min)

Figure 1a: Graphs of the absorbances versus times for the data from Table 1 for each of the blanched
samples, without trend line.

Fig. 1b. Absorbance vs Times for All Blanched Samples

1
0.9

y = 1.295x + 0.1247
R = 0.9418

0.8
A
b
s
o
r
b
a
n
c
e

Abs (120 s
blanch time)
Abs (90s
blanch time)

0.7
0.6

Abs (60 s
blanch time)

y = 1.0776x + 0.0907
R = 0.9579

0.5

Abs (30 s
blanch time)

0.4
y = 0.7091x + 0.054
R = 0.9663

0.3
0.2

Abs (15 s
blanch time)

y = 0.0155x + 0.0087
y = 0.0121x + 0.0082
R = 0.3474
R = 0.2651
y = 0.0089x + 0.0066
R = 0.2315

0.1
0
0

0.1

0.2

0.3
0.4
Time (min)

0.5

0.6

Abs (0 s
blanch time)

0.7

Figure 1b: Graphs of the absorbances versus times for the data from Table 1 for each of the blanched
samples, with trend line.

Fig. 2 ln (EAU) vs Blanching Time


8
7
y = -0.049x + 7.2633
R = 0.8628

6
5
ln (EAU) 4

ln (EAU) vs Blanching
Time

3
2
1
0
0

50

100

150

Blanching Time (s)

Figure 2: Graph of ln (EAU) versus blanching time (sec).

V.

CALCULATIONS:

Enzymatic Activity for 120 sec blanch time.


Enzymatic Activity = slope (au/min) x 1 EAU/0.001 (au/min)
= 0.0089 (au/min) x 1EAU/0.001 (au/min) = 8.9 EAU

The rate constant k


k is the slope of the best fit line for the ln (EAU) vs blanching time.
y = -0.049x + 7.2633
k = slope = 0.049/ sec

Half- life (t1/2)


ln [A]t = -kt + ln [A]0
kt = ln [A]0 - ln [A]t = ln([A]0/[A]t)
At the half-life [A]t = [A]0/2
kt1/2 = ln([A]0/([A]0/2)) = ln 2
t1/2 = ln (2) / k = 0.693 / k
= 0.693/ 0.049(/sec) = 14.14 sec

VI.

DISCUSSION:

(y = mx +b)

Looking at figure 1a and b, as the blanch time increases, the absorbance increases. For example, at blanch
time 120seconds, the absorbance at 120 seconds was 0.016 versus at blanch time 0seconds, the
absorbance at time 120 seconds was 0.945. The longer the blanch time, the smaller the slope. Smaller
slopes mean smaller changes in absorbance over time. Therefore, there should be less enzyme activity in
higher blanch times versus lower blanch times. It is apparent that effective blanching occurs in a range of
temperatures, around 80-100C. Temperature is important in order to avoid under-blanching (not all
enzymes are activated) and over-blanching ( too much heat will cook food and produce tissue damage).
The slope presented in figure 2 are the rate constants (-0.049). This rate constant can only be determined
experimentally. Also, rate constants are used in integrated rate laws to determine the half-life of the
reactant. If I was a food manufacturer, I would pick 90 seconds as the optimum blanch time since there is
little enzyme activity at this point, around 12.1 EAU. Different vegetables should have different optimum
blanching time due to their difference in sizes. They also have different surface to volume ratios that
should also be considered. For example, leafy greens will probably have lower optimal blanching time
compared to turnips. Enzyme activities I calculated decrease as the blanching time increases because as
blanch time increases, more enzymes will be inactivated, causing lower enzyme activity. Form this
experiment I learned how to measure blanching efficiency and the process of blanching. I also learned the
effects of blanching. The expected results was to have decreasing enzyme activities as the blanching time
increases. These expected results were obtained from lab.

VII.

CONCLUSIONS:

In this experiment, I learned the process of blanching and the importance of inactivating enzymes through
physically blanching turnips in lab. I learned what blanching efficiency is and its importance. I also
learned to apply rate laws to determine the kinetics of blanching. The method used was useful because it
helped determine the blanch efficiency and the values of half-life. The advantages of the method used was
to physically blanch and understand how the blanching process works and to be able to see the different
components that affect blanching through the method used. The disadvantage of the method used was that
the filtering process for the turnip extract made huge puddles on the lab bench. A suggestion is to have
any type of shallow containers that the beakers can lay on top of to prevent the puddles on the bench, if
possible.

VIII.

QUESTIONS:

1. The calculated half life represents the time at which half of the starting concentration is present.

2. During blanching, the enzyme was not continuously exposed to 100C since we are adding room
temperature turnips into them that causes the temperature to drop. We assumed that the water was 100 C
since that is the temperature at which water boils. I would redesign this experiment by using a
thermometer to keep track of the water's temperature, and wait until the water is 100C before adding the
next bag of turnip pieces.
3. The most blanched sample is always the first to be measured and the least blanched last because it is
easier to get used to writing down the absorbances since they will change really quickly as the blanch
time decreases. Also, the most blanched sample should have the smallest changes in absorbances
compared to the least blanched sample.