Beetroot Discs and the Integrity of the Cell Membrane
A Class XI student cuts six beetroot discs of identical size (1 cm diameter, 2 mm thick) and rinses them in distilled water until no colour leaks out. She places one disc in each of six test tubes containing 10 mL of liquid: (A) distilled water at 30 °C; (B) distilled water at 60 °C; (C) 5% NaCl at 30 °C; (D) 5% NaCl at 60 °C; (E) distilled water at 30 °C with two drops of detergent; (F) 10% ethanol at 30 °C. After 20 minutes the intensity of red colour in each tube is recorded: A faint, B very deep, C faint, D very deep, E deep, F moderate. She is asked to explain the pattern in terms of membrane structure, to state which tube best models a cell in a hypertonic medium, and to decide whether the red pigment is a protein or a lipid. All tubes were kept at the stated temperature for the whole 20 minutes.
1. Arrange the tubes in increasing order of pigment leakage and explain that order using the fluid mosaic model of the membrane.
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Increasing leakage: A ≈ C < F < E < B ≈ D.
The reasoning rests on where the pigment sits and what protects it. The red pigment is dissolved in the vacuolar sap, so it can escape only if both the tonoplast and the plasma membrane lose integrity. At 30 °C in distilled water (A) the bilipid layer is in its normal fluid state and the membrane proteins are folded correctly, so almost nothing leaks - the faint colour is background from the cut surface.
At 60 °C (B) two things happen at once: the lipids gain kinetic energy and the bilayer becomes excessively fluid and leaky, and membrane proteins and glycoproteins are denatured. The membrane loses selective permeability, so B is the deepest. Tube D shows that heat overrides the osmotic effect of NaCl.
Detergent (E) is amphipathic; it inserts into the bilayer, forms micelles and physically solubilises membrane lipids, producing deep leakage without heat. Ethanol (F) dissolves lipids and denatures proteins, but at 10% its effect is partial, hence moderate. Tube C is faint because 5% NaCl is hypertonic: water leaves the cell, the protoplast shrinks, the membrane is not disrupted, and leakage stays low.
2. Which tube best models a hypertonic medium, and what happens to a plant cell in it compared with a human red blood cell in the same solution?
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Tube C (5% NaCl at 30 °C) is the correct model, since the temperature is physiological and only the solute concentration differs from the control.
In tube C the external solution has a lower water potential (higher solute concentration) than the cell sap. Water therefore moves out of the cell by osmosis down the water-potential gradient, across both the tonoplast and the plasma membrane.
In a plant cell the rigid cellulose wall does not shrink. The protoplast pulls away from the wall - plasmolysis - and the cell becomes flaccid, but the wall prevents collapse and the cell does not burst or disintegrate. If such a cell is returned to water it can deplasmolyse, provided the loss was not prolonged.
A red blood cell has no wall. In the same hypertonic solution it loses water, shrinks and develops a spiky outline - crenation. In a hypotonic medium the opposite happens: the plant cell becomes turgid and the wall resists further entry of water, whereas the animal cell swells and undergoes haemolysis because it has no wall to oppose the pressure. The beetroot result is faint rather than zero because some cells at the cut edge are already damaged before the experiment begins.
3. Is the red pigment a protein or a lipid? What do the results in tubes A and F suggest about where the pigment is stored and about what the experiment actually measures?
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The pigment is not a lipid. Beetroot red is betacyanin, a water-soluble nitrogenous pigment (a betalain) that dissolves in the aqueous vacuolar sap. If it were a lipid it would sit inside the bilayer itself and would not diffuse out freely into distilled water, and it would not colour the water in tube A at all.
It is also not a protein, or at least not a membrane protein: heating to 60 °C denatured the membrane proteins and caused maximum leakage, yet the colour in tube B is still red, not a coagulated brown precipitate. The pigment survives the temperature that destroys the proteins.
Tube A shows that when the membranes are intact almost no pigment escapes, which means the pigment must be enclosed by a membrane, not free in the cytoplasm or the wall. Tube F shows that a lipid-dissolving agent releases the pigment without heat, confirming that the barrier being breached is the lipid bilayer and not the cellulose wall.
The experiment therefore measures membrane integrity, not pigment chemistry: every treatment that damages the bilayer or the tonoplast raises the absorbance, and the pigment is only the reporter.