Domain: Natural Science — Biochemistry & Molecular Biology
Target concept: Alkaline phosphatase converts pNPP into yellow p-nitrophenol in solution.

Video generation prompt: A clear cuvette containing colorless pNPP is placed in a benchtop spectrophotometer; alkaline phosphatase is added and mixed while the visible light beam passes through the reaction system.

Evaluation guide:
(i) Target concept: Enzyme-catalyzed chromogenic reaction visualized in real time within an optical measurement device.
Minimum mechanism: pNPP must be chemically stable and colorless in alkaline buffer prior to enzyme addition; alkaline phosphatase must be active and soluble; hydrolysis must yield p-nitrophenol as the *sole* colored product; p-nitrophenol must be soluble and stable in the reaction medium to produce homogeneous yellow color.
(ii)Storyline: Frame 0–2: Static shot. Cuvette centered in spectrophotometer chamber, filled with colorless, transparent pNPP solution; visible light beam aligned and passing straight through cuvette. Frame 3–4: Addition. Droplet(s) of alkaline phosphatase enter cuvette from above, contacting surface without splashing. Frame 5–7: Mixing onset. Fluid motion begins (gentle swirl or inversion), solution remains colorless. Frame 8–9: Color emergence. Uniform yellow hue appears progressively and homogeneously *within* the moving liquid volume, co-occurring with mixing motion. Perspective: Fixed frontal view, slight downward angle to show cuvette interior and beam path; no hand/actor intrusion; full visibility of solution color and beam interaction throughout.

Manual:
Experimental Procedure
1. Place a clear cuvette containing colorless pNPP solution into the sample chamber of a benchtop spectrophotometer.
2. Add alkaline phosphatase solution directly into the cuvette.
3. Mix gently while the visible light beam passes continuously through the cuvette.
Observation of Experimental Phenomena
The initially colorless pNPP solution becomes uniformly transparent yellow during mixing, with no precipitation or cloudiness.
Summary of Experimental Principles
Alkaline phosphatase catalyzes hydrolysis of pNPP, producing yellow p-nitrophenol—a quantifiable chromogenic product in biochemical assays.

Prompt Grounding rubric:
level_1: No cuvette is visible anywhere in frame; no benchtop spectrophotometer is shown; no pNPP solution appears; the alkaline phosphatase addition step never happens; no mixing action occurs; the visible light beam and its path through the sample are never shown.
level_2: The cuvette and the spectrophotometer both appear on screen, but they are not shown together correctly (e.g., the cuvette sits outside the sample chamber, or the beam path has no visible relationship to the cuvette); the pNPP solution shown is not colorless; alkaline phosphatase is shown somewhere in the shot but is never added into the cuvette; the mixing motion is missing, or is applied to a different container than the cuvette holding the pNPP.
level_3: The cuvette sits inside the benchtop spectrophotometer's sample chamber; the pNPP solution inside it is colorless; alkaline phosphatase is added into the cuvette with a visible liquid-transfer action; the contents are then mixed (swirl or gentle inversion); the visible light beam can be seen passing through the cuvette (via a lit path, glow, or shadow cast beyond it).
level_4: Everything in score 3 holds, and additionally the geometry is precise: the cuvette sits centered in the chamber; the beam runs straight through the middle of the cuvette; the alkaline phosphatase is delivered directly into the cuvette opening with no spill onto the chamber or bench; the mixing stays confined to the cuvette's contents, without the cuvette itself being lifted out, tipped over, or displaced; the pNPP solution remains colorless right up until the enzyme is added.
level_5: Everything in score 4 holds, and additionally the sequence of setup events is strictly ordered: (1) the cuvette holding colorless pNPP sits still in the chamber before any other action begins; (2) the alkaline phosphatase addition starts only once that still placement is established; (3) mixing begins only after the addition is finished; (4) the light beam remains visible without interruption for the entire shot; (5) no hand, tool, or unrelated object ever enters or blocks the frame.

Scientific and Causal Correctness rubric:
level_1: No yellow color develops at all; or the pNPP solution is already yellow/colored before alkaline phosphatase is added; or the substance added is clearly not an enzyme solution consistent with alkaline phosphatase; or the final solution turns a hue inconsistent with p-nitrophenol (e.g., blue, green, brown, cloudy white); or any color change shown has no visible relation to the addition/mixing event.
level_2: A yellow tint appears, but the science is wrong in some way: the color shows up before the alkaline phosphatase actually contacts the pNPP, or appears without any mixing taking place, or forms only as a surface film or floating precipitate rather than dissolving through the liquid, or reads as orange, brown, or green rather than the pale yellow expected of p-nitrophenol in alkaline solution.
level_3: The pNPP solution starts out colorless; alkaline phosphatase is added and mixed in; a uniform pale-yellow color consistent with p-nitrophenol then fills the solution rather than staying at the surface; the color change follows the mixing step rather than preceding or replacing it.
level_4: Everything in score 3 holds, and additionally the cause-and-effect timing is tight: no yellow tint appears during the addition step itself: it only begins once mixing gets underway, with no long unexplained delay afterward; the yellow builds up gradually and evenly through the whole liquid column rather than in isolated patches; the solution stays clear and transparent yellow throughout, with no cloudiness, film, or floating particles.
level_5: Everything in score 4 holds, and additionally the causal sequence is complete and clean: (1) the pNPP solution is visibly colorless for at least a second before the enzyme droplet enters; (2) no yellow appears at any point during the addition itself, only after mixing begins; (3) the yellow color spreads outward from the region being stirred rather than appearing everywhere at once or from an unrelated spot; (4) the finished solution is evenly pale yellow with no leftover colorless streaks or pockets; (5) the color stays chemically stable to the end: no shift to a second hue, no fading back toward colorless, no precipitate forming as mixing concludes.

Spatiotemporal Consistency rubric:
level_1: The cuvette jumps to a different position or chamber slot between moments with no connecting motion shown; the spectrophotometer body or chamber vanishes and reappears; the light beam switches fully on and off with no visible cause; the liquid's fill level or shape changes instantly between moments with no pour or mixing motion linking them; the room or bench lighting around the setup abruptly changes (e.g., a bright frame is immediately followed by a dark one) with no cause tied to the experiment.
level_2: The cuvette visibly slides, rotates, or tilts by a large, unexplained amount partway through the mixing action, with no cause shown for the change; the beam appears to bend, break, or shift off the cuvette's centerline; the stirring or inversion motion stutters, jumping from one pose to a distant pose without the in-between motion; the highlight/reflection on the cuvette's glass switches from present to absent (or the reverse) without a matching change in the beam or the viewing angle.
level_3: The cuvette stays in the same chamber slot and orientation throughout, aside from the deliberate mixing motion; the light beam stays continuously visible and runs straight through the cuvette's center the whole time; the swirling or inversion motion of the liquid looks like real fluid behavior: it flows and settles rather than snapping between poses; any change in the cuvette's illumination or surface highlight tracks the mixing motion and viewing angle rather than happening on its own.
level_4: Everything in score 3 holds, and additionally the motion reads as physically grounded: the liquid keeps moving briefly and settles gradually after the mixing motion stops, rather than freezing instantly; the beam's width and brightness stay steady rather than flickering; the highlight and any reflection on the cuvette's glass shift smoothly as the liquid moves, without jumping to a new spot or vanishing between one moment and the next; the meniscus (liquid surface line) stays visible and continuous, never blinking out.
level_5: Everything in score 4 holds, and additionally the spatiotemporal behavior is fully coherent: (1) the cuvette's position and tilt look unchanged, moment to moment, whenever it is meant to be resting still: no perceptible shift, drift, or wobble in its position or angle; (2) the beam stays aligned with the cuvette's centerline for the entire sequence, never drifting to one side; (3) the liquid's motion speeds up and slows down gradually, with no instant starts, stops, or teleporting fluid; (4) the cuvette and the spectrophotometer body behind it move together consistently with the camera, with no floating or sliding relative to each other; (5) the moving liquid's color, clarity, and containment inside the cuvette stay clearly legible throughout, with no flashes of static or smearing that hide what is happening inside.