Estimation of free and total acidity in a gastric juice sample

Presented by Dr. Nada Soliman

Content • Biological fluid: gastric juice

pH Types of acid

• Quantitative method: titration Principle of titration Type of titration used in free and total acidity

estimation Titrant and indicators used End points (X1 and X2) procedure and calculation of free and total acidity.

• Fractional test meal procedure Normal curve Abnormal curves: 3

Gastric juice the most acidic secretion of the body fluid, with a pH=

1.0 (in adult) Types of acid in gastric juice:1. HCL: secreted by parietal cells of the gastric mucosa

Free HCL Combined HCL (combined with protein)

2. Organic acids: lactic, acetic acid…… Free HCL+ combined HCL+ organic acids: responsible

for total acidity Free HCL: responsible for free acidity of the gastric

juice When the free HCL is neutralized the pH of gastric

juice become 3

• Alkaline tide:Means the presence of an alkaline urine that

often follows the ingestion of a meal due to the presence of excess bicarbonate in the blood during HCL secretion

Biological substance of unknown concentration andof known volume (sample)

To determine

its concA standard solution: titrant

Knowing the volume of the titrant needed to react completely with the analyte, helps to determine its concentration

The titrant is added drop by drop until the reaction is complete

According to type of the reaction, we have 3 types of titration:1. Acid-base: HCL with

NaOH2. Precipitation: silver

nitrate with barium chloride

3. Oxidation-reduction: uric acid with K permenganate

1. Another substance2. Known concentration3. React with the analyte4. Volume: measured

during the experiment

Definition of titrant: the solution of known concentration added during titration

How we know that the reaction is complete?By observing physical change. the end pointCould be:

changes in color Appearance or disappearance of ppt Change in electrical properties

Definition of end point:Point in titration where physical change manifests itself

For color changes, we use an Indicator:Supplementary chemical compound that changes its color with changes in the medium at the end point

Equipments • Burettes• holders.• Pipettes• beakers

Types of titration: acid-base titration

Titrant: 0.1 N NaOH End point (X1 &X2): color

change Indicators used:• Topfer’s:

Pink color pH <3Yellow pH>3Salmon pink pH=3 (X1)

• Phenolphthalein:Colorless at pH <8Pink at pH= or >8 (X2)

procedure1. Fill a burette with 0.1 N-

NaOH solution2. Pipette 5 ml of the gastric

juice into a beaker (using a 5 ml pipette).

3. Add to the contents of the beaker one drop of phenolphthalein indicator colorless

4. Add 2-3 drops of topfer’s indicator pink

5. Titrate against 0.1 N-NaOH until the color changes from pink to salmon pink (orange yellow).

6. Stop the titration and take the reading of the burette which represents the end point (= X1) at which all free HCl, present in the 5 ml gastric juice, is neutralized by 0.1N NaOH. Now the pH is 3

7. continue titration until the color changes to pink. Stop the titration and take the reading of the burette from the start which represents the end point(= X2) at which all the total acidity, present in the 5 ml gastric juice, is neutralized by 0.1N NaOH. Now the pH is 8.

5ml GJ + phenolphthale

incolorless

Topfer’s

pink

NaOH

Salmon pink

NaOH

pink

X1

X2

X1=number of ml of 0.1 N-NaOH required to neutralize the free HCl in 5 ml GJ

Yellowish

gradually

X2=number of ml of 0.1 N-NaOH required to neutralize total acids in 5 ml GJ

Calculation of free and total acidity

• From the previous procedure, X1 and X2 were determined in only 5 ml of the gastric juice sample.

• Free acidity is expressed as the number of milliliters of 0.1 N-NaOH required to neutralize the free HCl in 100 ml of gastric juice.

• Total acidity is expressed as the number of milliliters of 0.1 N-NaOH required to neutralize the total acids present in 100 ml of gastric juice.

• So:Free acidity = X1 x 100/5 0.1 N-NaOH %

Total acidity =X2 x 100/5 0.1 N-NaOH %

Fractional Test Meal1. The patient comes fasting and then a stomach

tube (Ryle tube) is introduced in the mouth.2. a fasting sample is aspirated and titrated for

determination of free and total acidity.3. Then the patient takes a standard meal of 1-2

pieces of toast + 2 cups of tea (with sugar but no milk).

4. Samples of 10 ml are then aspirated every 15 or 30 minutes for about 2.5 hours.

5. After all samples are titrated, the amount of free and total acidity can be plotted in a curve against time.

The curve of normal people shows

the following characters:

The fasting acidity is

neutralized by 20-30 ml 0.1

N-NaOH / 100 ml GJ.

15-30 minutes after the meal, the

acidity falls to a lower

level.

Then the acidity rises again and

reaches its maximal value after about

90 minutes.

After 120 minutes,

acidity starts to decline

gradually. the fasting level is

reached at about 150 minutes.

• AT 15-30 minutes, This decline is due to:1. Dilution of gastric juice with saliva and tea.2. Neutralization of gastric juice by NaHCO3 of saliva,

protein of the meal and mucin inside the stomach.• At 90 minutes, the acidity rises again and

reaches its maximal value:o due to stimulation of the gastric glands by the

meal.• at the end of about 150 minutes, This

decline is due to:1. Gastric secretion gets diminished.2. Gastric contents are evacuated to the duodenum.3. Gastric contents are neutralized by duodenal

regurgitation.

Abnormal curves:

• Hyperacidity (hyperchlorhydria): Acidity is more than normal e.g. in cases of duodenal ulcers and gall bladder diseases.

• Hypoacidity (hypochlorhydria): Acidity is below normal e.g. in gastric carcinoma.

• Anacidity (achlorhydria): There is complete absence of hydrochloric acid in this condition. The curve obtained is flat. Achlorhydria is compatible with pernicious anemia.