Effect Of Storage Time And Type Of Fruits On Mass Percentage Of Citric Acid Available

How does the mass percentage of citric acid available from citrus fruits (lemon, lime, oranges) depends on the storage time and the types of citrus fruits and fresh lime taken?

Citrus fruits are a regular part of my diet and during this Covid-19 a lot of health experts had advised to consume more of the citrus fruits as they used to build our immunity. Therefore, I had made it a habit to prepare these juices and store it in the refrigerator so I could drink it whenever I want. However, I observed that the flavor of the juice is changing over time which made me curious, “Why does the flavour of the juice change over time ?” and “What are the sources for citric acid ?” . I wanted to understand why this change in flavour took place; is this also affecting the nutritional value. I also investigate whether this change in nutritional value is the same for all citrus fruits or varies among fruits. This would help me understand what is the best citrus fruit which is  safest to store for longer periods of time and which has least change in flavour, proving the best fruit juice to be consumed. As I had so many questions about this topic I decided to do some research to help me understand this, I came across an article which discusses the key ingredient in citrus fruits, which is citric acid itself and the article explained how there are different amounts of citric acid in the different fruits. I made the connections to the concept learned during Chemistry class like the concept of titration as an analytical procedure for quantitative estimations of acids and bases. I realized I wanted to connect something I learned from class and to understand how to apply it to real life applications and planned to use it in an experiment.

Citric acid

Citric acid is a weak organic acid that has the molecular formula C6H8O7. It occurs naturally in citrus fruits. It belongs to the functional group carboxylic acid group. It is naturally found in citrus fruits like lemon, lime, orange, grapefruits, pomelos and is used widely in industry as an acidifier, as a flavoring and chelating agent. Citric is soluble in Water and dissolves in absolute (anhydrous) ethanol. Citric acid is a weak organic acid present naturally in living cells. Citric acid is an alpha-hydroxy acid with a three carbon skeleton, which has three carboxylic acid groups (COOH), and one hydroxyl group (OH). It is used as a cleaning agent which is used on a daily basis in the kitchen. It is used to add a sour taste to soft drinks and other food items, in shampoos, food coloring. It is a natural preservative. It is used to remove the chalky deposit from evaporators, kettles, boilers.

Word Equation: Citric Acid Sodium Hydroxide → Sodium Citrate Water

Chemical Equation: C₆H₈O₇ (aq) + 3NaOH (aq) →  C₆H₅O₇Na₃ (aq)+ 3H₂O₄ (aq)

When you look at the structure of citric acid, you can see that there are three carboxylic acid groups, which mean that there need to three NaOH for each of the atoms to react with each one of them

Titration is a qualitative analysis technique that can be used to calculate the concentration of an unknown solution with the use of a solution with a known concentration. The process is usually carried out by gradually adding a standard solution ( solution of known concentration) of titration titrant with a  burette essentially a long, graduated measuring tube with a stopcock and a delivery tube at its lower end until the color change at the end point. The change in color occurs due to use of indicator, usual phenolphthalein³

The citric acid percentage content of each fruit (lemon lime, orange will does not depend on storage time.

The citric acid percentage content of each fruit (lemon lime, orange will does not depend on storage time.

-Chemicals were used in a manner that avoids wastage.

-All chemicals were disposed off into a chemical waste bin to avoid interaction with the environment.

-Unused chemicals were returned to the laboratory, safely.

Minimal amount of chemicals were used to conduct trials, keeping in mind the economic concerns and environmental effects.

Preparing 100 cm³ of 0.1 mol.dm^-3 NaOH solution

Molar The solution made has to be a volume of 100 cm³ and concentration 0.1 mol dm-³  -3.

Thus, the number of moles can be calculated from the formula:

Number of moles (n) = molar concentration ( C)  X     volume in dm³ (V ) =\((0.010 X\frac{100}{1000})\)= 0.001

Mass of NaOH to be used

= number of moles X Molar Mass= 0.001 X 39.997

= 0.039997 g

= 0.04 g (rounded of two decimal places)

-For lemon, squeeze it to remove the juice into a beaker. For lime and lemon use a machine to remove the juice and pour into a beaker.

-Filtrate the juice using a funnel with filter pare, empty the filtrate in another beaker.

-First rinse the pipette with some distilled  water.

-Take 20.00 ±0.05 cm³ of the juice from the filtrate using a pipette into a 100cm³ flask

-Juice should be diluted by adding distilled water until the mark reaches 100 cm³, start it using a glass rod.

-Pouring the solution in a fresh beaker and use a pipette to take 20.00 ±0.05 cm³ again

-Pour this into a conical flask, then add a few drops of phenolphthalein indicator

-Take initial reading of NaOH  from the burette (two decimal places)

-Drop NaOH into the juice slowly, drop by drop until the color changes pink

-Check and the record the reading on the burette

-Calculate amount of NaOH (initial reading -  final reading)

-Calculate the moles of citric acid reading to the NaOH, use this information to calculate the mass of citric acid.

-Using this information calculate the percentage of mass

-The data was collected in 3 trials and the same procedure was repeated for lemon juice, orange juice.

-All steps were repeated for 5 days, three trials and the same procedure for all three fruits.

Sample calculation

For Day 1, Trial-1

Initial burette reading = 0.00 ± 0.05 cm³

Final burette reading = 7.20 ± 0.05 cm³

Difference in burette reading = Final – Initial = (7.20 ± 0.05 cm³) – (0.00 ± 0.05 cm³) = 7.20 ± 0.10 cm³

As the data collected is of acid base titration, the reading that is repeated (concordant or precise reading) has been chosen as an average instead of calculating arithmetic average to procure more accurate data.

For example, for Day-1, the three trial values for difference in burette readings are 7.20 ± 0.05 cm³, 7.00 ± 0.05 cm³ and 7.00 ± 0.05 cm³ respectively. So, the reading 7.00 ± 0.05 cm³ repeats itself twice and is thus the precise or concordant reading. Thus, the average reading is taken as 7.00 ± 0.05 cm³

For Day-1, Standard deviation (SD)=\(\frac{(7.20-7.00 )^2+(7.00-7.00)^2 +(7.00-7.00)^2}{3}\)=0.12

Sample calculation

Citric acid reacts with NaOH according to the equation

HOCH2CH2OH(COOH)CH₂COOH + 3 NaOH →  NaOCCH₂CH₂OH(COONa)CH₂COONa + 3H₂O

Moles of citric acid ( n citric acid ) =\(\frac{moles\ of \ NaOH}{3}\)\((as\ they \ citric \ acid \ and\ NaOH\ reacts\ in\ the \ ratio\ 1:3)\)

=\(\frac{molar\ concentration \ of \ NaOH\ used× \frac{Average\ volume \ of \ NaOH \ taken \ in\ cm^3}{1000} }{3}\)=\(\frac{0.0×\frac{7.00}{1000}}{3}\)=2.33 × 10^-4

Molar concentration of citric acid

\(=\frac{number\ of\ moles\ of \ citric\ acid }{\frac{Volume\ of \ juice\ used\ in \ cm^3}{1000}}\)=\(=\frac{2.33× 10^{-4} }{\frac{20}{1000}}\)\(=\frac{2.33× 10^{-4} }{{20 × 10^{-3}}}\)=0.1165 × 10^-1

=11.65 × 10^-3 mol dm^-3

Moles of citric acid present within 1 dm³ or 1 L  of juice = 11.65 × 10^-3 mol

Mass of citric acid present within 1 dm³ of juice

= moles × molar mass=11.65 × 10^-3 ×192.12

=2238.19 × 10-³=2.238 g ≅2.24 g

Concentration of citric acid = 2.24 g L^-1

For Day-1 of lime juice

Average volume of NaOH = Precise reading among the three trial values of difference in burette reading = 7.00 ± 0.10 cm³

Concentration of NaOH solution used (c)=\(\frac{moles }{Volume}\)=\(\frac{mass (m) }{\frac{molar \ mass}{Volume(V)}}\)

\(\frac{∆ \ c}{c}=\frac{∆ \ m }{ m}+\frac{∆\ V }{V}\frac{± 0.01 }{0.40}+\frac{±0.60 }{100.00}±0.03\)

Moles of NaOH (n) = concentration (c) × Volume of NaOH consumed (V)

Fractional error in moles of NaOH\(\bigg(\frac{∆ n }{n}\bigg)=\frac{∆ c }{ c }+\frac{∆ V }{ V }=±0.03+\frac{± 0.10}{7.00}=±0.04\) 

Moles of citric acid =\(\frac{moles\ of \ NaOH }{3}\)

Therefore, fractional error in moles of citric acid = fractional error in moles of NaOH

Mass of citric acid = moles of citric acid  molar mass

Concentration of citric acid in g L^-1\(=\frac{mass\ of \ citric \ acid }{Volume \ of \ juice}\)

Volume of juice used (V_juice)= 20.00 ± 0.05 cm³

Fractional error in concentration of citric acid

= fractional error in mass of citric acid + fractional error in volume of juice

\(=\frac{∆n }{n}+\frac{∆V }{V}=(± 0.04)+\frac{± 0.05}{20.00}b\)

Percentage error in citric acid concentration = fractional error\( 100 \) × 100 = ± 0.04 × 100 = 4.00

Graph-1 is a scattered plot of mass percentage of citric acid in g/L against the number of days for which the juice was stored.

• The graph displays that the mass percentage of citric acid in g/L (mass of citric acid present within 1 L of the juice) is decreasing from 15.47 g/L to 14.15 g/L for lemon juice, 8.74 g/L to 6.79 g/L for orange juice and 2.24 g/L to 1.22 g/L for lime juice. Thus, for all the three kinds of juices, it has been observed that the mass percentage of citric acid decreases with time. Thus, a negative correlation can be claimed between the mass percentage of citric acid and the storage time (measured in terms of number of days).
• For all the days, the values of mass percentage is the highest for lemon juice followed by orange juice and the least for lime juice. This allows us to claim that the citric acid content is maximum for lime followed by lemon and the least for orange juice.
• An equation of linear trend line has been displayed in the graph for each of the three types of juices and the trend lines are shown below:
  
  Lemon juice
  y=-0.322x + 15.824
  at y =0,
  x = \(\frac{15.824}{0.322}\)=49.14 ≅49
  
  
  Orange juice
  y=-0.454 x + 9.19
  at y =0,
  x = \(\frac{9.19}{0.454}\)=20.24≅20
  
  
  Lime juice
  y = -0.269x + 2.517
  at y = 0
  x = \(\frac{2.517}{0.269}\)=9.35 ≅9
   
• The above discussion reveals that theoretically, the citric acid content within the juice must diminish to 0.00 within 49 days for lemon juice, within 20 days for orange juice and within 9 days for lime juice. Thus, it can be claimed that the citric acid content lasts the longest for lemon juice followed by orange juice and the least for lime juice.
• Comparing the gradient of the trend line; -0.322 for lemon juice, -0.454 for orange juice and -0.269 for lime juice; it can be claimed that the mass percentage of citric acid in the juice is decreasing with time as the values of gradient are negative. Moreover, the values of the gradient shows that the decrease of mass percentage with time is fastest for orange juice as it has the maximum value of gradient and minimum for lime juice as it has the minimum value of gradient.

Sample calculation

Mean % of citric acid content =\(\frac{Sum \ of\ mass\% of \ all\ five\ days}{5}\)= \(\frac{2.24+1.99+1.76+1.34+1.22 }{5}\)=1.71 gL^-1

The y axes values represent the dependent variable percentage content of citric acid present in each type of fruit The x axis values represent the independent variable, which is the type of fruit. As the x axes values (independent variable values) are categorized and not numbers/ figures, it is not possible to make a scatter plot or line graph. Therefore, bar graph being most suited to present this type of data and effectively exhibiting the data collected.

The graph clearly displays that the mean content of citric acid in g/L is maximum for lemon juice (14.86 g/L) followed by orange juice (7.83 g/L) and least for lime juice (1.71 g/L). This claim is in accordance with the results from Graph-1.

Citric acid is an organic tri-carboxylic acid. It can undergo decarboxylation according to the reaction below

HOOC-CH₂-C(COOH)(OH)-CH₂(COOH) ------🡪     H₃C-CH(OH)-CH₃ + 3CO₂ (g)

As gas is liberated during the reaction, the disorderness of the system increases and thus the change of entropy is positive indicating an increase in entropy of the system. Thus, the reaction is entropically favored and spontaneous in nature. With the increase in storage time, more and more citric acid molecules undergo decarboxylation and thus the mean mass percentage of citric acid decreases. The qualitative observation that more bubbles were formed and the appearance of the juices turned hazy with the passage of time confirms the liberation of CO₂ with the passage of time.

How does the mass percentage of citric acid available from citrus fruits (lemon, lime, oranges) depends on the storage time and the types of citrus fruits and fresh lime taken?

The mass percentage of citric acid in g/L (mass of citric acid present within1 L of the juice) is decreasing from 15.47 g/L to 14.15 g/L for lemon juice, 8.74 g/L to 6.79 g/L for orange juice and 2.24 g/L to 1.22 g/L for lime juice. Thus, for all the three kinds of juices, it has been observed that the mass percentage of citric acid decreases with time. Thus, a negative correlation can be claimed between the mass percentage of citric acid and the storage time (measured in terms of number of days).

The citric acid content within the juice must diminish to 0.00 within 49 days for lemon juice, within 20 days for orange juice and within 9 days for lime juice. Thus, it can be claimed that the citric acid content lasts the longest for lemon juice followed by orange juice and the least for lime juice.

The decrease of mass percentage with time is fastest for orange juice as it has the maximum value of gradient and minimum for lime juice as it has the minimum value of gradient.

The mean content of citric acid in g/L is maximum for lemon juice (14.86 g/L) followed by orange juice (7.83 g/L) and least for lime juice (1.71 g/L). This claim is in accordance with the results from Graph-1.

With the increase in storage time, more and more citric acid molecules undergo decarboxylation and thus the mean mass percentage of citric acid decreases. The qualitative observation that more bubbles were formed and the appearance of the juices turned hazy with the passage of time confirms the liberation of CO₂ with the passage of time.

Graph-1 shows the values of regression coefficient obtained from Excel for the equation of linear trend line depicting the correlation between the mass % of citric acid in g/L and the number of days. The values are 0.9851 for lemon juice, 0.9723 for lemon juice and 0.9794 for lime juice. This shows that there is a strong correlation between the mass percentage of citric acid in g/L and the storage time (measured in terms of number of days). Moreover, Graph-2 clearly indicates the differences in the values of mean values for the mass percentage of citric acid in juices. Thus, the null hypothesis is rejected and the alternate hypothesis has been accepted.

• Wide range of independent variables- five days of storage time and three different types of fruits has been chosen due to which, the result is more specific and accurate.
• The information found in this IA is resourceful as it explains what affects the nutritional level of the fruits and provides an idea of what food sources are rich in citric acid. I understood which one is best to have and safest to store for longer periods of time, which would least change in flavor, proving the best fruit juice to be consumed.
• The experimental procedure is very simple and can be easily followed. The experiment can be done in other laboratories as well which increases the validity of the data.

Citric acid also contains a alcohol group – (OH). This group can also react with NaOH along with the three carboxylic acids. This changes the mole ratio of the citric acid and NaOH to 1:4 from 1:3. Thus, the calculations and data processing in this investigation would become invalid. Alcohol (OH) group is acidic in nature and thus this possibility cannot be averted.

(HOOC)CH₂-C(COOH)(OH)-CH₂-(COOH) + 4 NaOH -----🡪 (NaOOC)CH₂-C(COONa)(ONa)-CH2-(COONa) + 4 H₂O

To optimize this, the concentration of citric acid can be monitored by measuring the absorbance of the solution using a UV-Visible spectroscopy at a wavelength at which the molecule shows maximum absorbance.

Apart from storage time, storage temperature is also an important parameter to decide the shelf-life of any processed food. Thus, investigating the effect of temperature on the mean citric acid content of the juices will be a significant research. To do so, we can take the same three kind of juices and heat them to different temperatures using a Bunsen burner or electric heater. After that, the mean citric acid content can be determined using the acid base titration with NaOH.

-Penniston, Kristina L., et al. “Quantitative Assessment of Citric Acid in Lemon Juice, Lime Juice, and Commercially-Available Fruit Juice Products.” Journal of Endourology, vol. 22, no. 3, Mar. 2008, pp. 567–570, www.ncbi.nlm.nih.gov/pmc/articles/PMC2637791/, 10.1089/end.2007.0304.

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