Question 1(a)(i): Performing a Serial Dilution

You need to perform a serial dilution by half, starting with a 1.00% reducing sugar stock solution (R), to prepare four additional, progressively weaker concentrations (0.50%, 0.25%, 0.125%, 0.0625%). The method involves transferring 10 cm³ from the previous concentration and adding 10 cm³ of distilled water (W) at each step. Which step below describes an incorrect action within this specific procedure?

• Label four clean beakers for the concentrations: 0.50%, 0.25%, 0.125%, 0.0625%.
• To prepare the 0.50% solution: Measure 10 cm³ of 1.00% R, transfer to the ‘0.50%’ beaker, add 10 cm³ of W, and mix.
• To prepare the 0.25% solution: Measure 10 cm³ of W, transfer to the ‘0.25%’ beaker, add 10 cm³ from the 0.50% solution, and mix.
• To prepare the 0.125% solution: Measure 10 cm³ from the 0.25% solution, transfer to the ‘0.125%’ beaker, add 10 cm³ of W, and mix.
• Ensure thorough mixing after adding water at each dilution step.

Question 1(a)(ii): Recording Benedict’s Test Results

You have performed Benedict’s tests on your serially diluted reducing sugar solutions (1.00% down to 0.0625%) and timed how long it takes for the first colour change to appear upon heating. How should you correctly record these results in a table? Which step describes an incorrect way to present or record this data?

• Create a table with columns clearly labelled for ‘Concentration of reducing sugar (%)’ (independent variable) and ‘Time for first colour change / s’ (dependent variable).
• Record the time taken for each concentration tested, ensuring all times are recorded consistently, for example, to the nearest whole second (e.g., 45 s, not 45.3 s if that precision wasn’t achievable).
• If no colour change occurs within the maximum observation time limit (e.g., 120 seconds), record this clearly using appropriate notation (e.g., ‘> 120 s’ or ‘No change’).
• Ensure the recorded data shows a trend where higher concentrations of reducing sugar take a longer time to cause a colour change.
• Include rows in the table for all the concentrations prepared and tested in the serial dilution (1.00%, 0.50%, 0.25%, 0.125%, 0.0625%).

Question 1(a)(iii): Identifying the Dependent Variable

In the investigation where you measured the time taken for Benedict’s reagent to change colour when heated with different concentrations of reducing sugar solution, what is the dependent variable? Which option below incorrectly identifies the dependent variable?

• The variable that is measured by the investigator in response to changes in another variable.
• The time measured in seconds from the start of heating until the first visible colour change.
• The different concentrations of reducing sugar solution that were prepared (1.00%, 0.50%, etc.).
• The factor that potentially ‘depends on’ the concentration of the reducing sugar.
• The outcome measured to see the effect of the independent variable.

Question 1(a)(v) & (vi): Estimating Unknown Concentrations

Imagine you tested extracts from two different seeds, G1 and H1, using the same Benedict’s test procedure. G1 took 50 seconds to change colour, and H1 took 95 seconds. Your results for known standards were: 1.00% (20s), 0.50% (35s), 0.25% (60s), 0.125% (100s), 0.0625% (>120s). You need to estimate the concentrations of G1 and H1… Which step describes an incorrect approach to making these estimates?

• Compare the time for G1 (50s) with the times for the known standards.
• Note that 50s falls between the time for 0.50% (35s) and 0.25% (60s).
• Estimate the concentration of G1 to be between 0.25% and 0.50%, likely closer to 0.25% because 50s is closer to 60s (time for 0.25%) than 35s (time for 0.50%).
• Compare the time for H1 (95s); note it falls between 0.25% (60s) and 0.125% (100s). Estimate H1 concentration between 0.125% and 0.25%, closer to 0.125%.
• Estimate the concentration of G1 precisely by calculating the average of the concentrations whose times bracket 50s: (0.50% + 0.25%) / 2 = 0.375%.

Question 1(a)(vii): Improving Estimation Accuracy

After getting preliminary estimates… you want to modify the procedure to get more accurate estimates. Which of the following modifications would be incorrect or least helpful for improving accuracy?

• Prepare a new set of standard reducing sugar solutions with concentrations at much smaller intervals (e.g., every 0.05%) within the ranges suggested by the preliminary estimates for G1 and H1.
• Plot a calibration curve (graph) of the time taken for colour change (y-axis) against the known reducing sugar concentrations (x-axis) using the standard solutions, and read the unknown concentrations from the curve.
• Use a thermostatically controlled water bath to ensure all Benedict’s tests (standards and unknowns) are performed at exactly the same, constant temperature.
• Instead of timing the first colour change, perform the tests only once for each concentration but observe them very carefully for subtle differences.
• Use a colorimeter to obtain quantitative measurements of the final colour intensity (absorbance) after a fixed heating time, instead of relying on subjective timing of the first change.

Question 1(a)(viii): Effect of High Temperature

Imagine the experiment comparing enzyme activity… was repeated at 80°C… the estimated reducing sugar concentration was very low and similar for both seeds… Which statement provides an incorrect explanation?

• The high temperature (80°C) likely caused the enzymes involved in producing reducing sugars (e.g., amylase breaking down starch) to denature.
• Denaturation involves a significant change in the enzyme’s three-dimensional tertiary structure, particularly affecting the shape of the active site.
• Once the active site shape is irreversibly altered, the enzyme can no longer bind effectively to its substrate (e.g., starch).
• At 80°C, the enzymes work extremely rapidly, consuming all available stored carbohydrate substrate very quickly, thus leading to low final concentrations of reducing sugars.
• If the key enzymes responsible for hydrolyzing storage carbohydrates into reducing sugars are denatured and lose their function, then very few or no reducing sugars will be produced from the stores.

Question 1(b)(i): Plotting Graph of Amylase Activity

You are given data showing the activity of amylase (in arbitrary units, au) at different times after seed germination (in hours). You need to plot a graph of this data. Which step below describes an incorrect procedure for plotting this graph?

• Orient the axes correctly: Place the independent variable (‘Time after germination / hours’) on the horizontal x-axis and the dependent variable (‘Activity of amylase / au’) on the vertical y-axis.
• Choose linear scales for both axes that allow the plotted data points to occupy less than one-quarter of the available graph paper area, in order to conserve paper.
• Label both axes fully, including the quantity measured and its units (e.g., ‘Time / hours’, ‘Amylase Activity / au’), with clearly marked intervals along the scales.
• Plot each data point precisely at the intersection of its corresponding x and y values, using a small, neat symbol like a cross (x) or a dot within a circle (⊙).
• Draw an appropriate line to illustrate the trend shown by the points – this could be a smooth curve drawn by hand that best fits the overall pattern, or ruled straight lines connecting consecutive points if specified.

Question 2(a)(i): Drawing a Plan Diagram of Leaf Section P1

Imagine you are viewing a prepared slide (P1) showing a transverse section of a plant leaf under low power. You need to draw a plan diagram of a representative part of this section. Which instruction below is incorrect for producing a good plan diagram?

• Make the drawing large enough to see the different tissue layers clearly, utilising a significant portion of the space provided.
• Use clear, single, continuous lines to delineate the boundaries between the main tissue regions (e.g., epidermis, palisade mesophyll, spongy mesophyll, vascular bundle).
• Include drawings of numerous individual cells within the palisade and spongy mesophyll layers to show their shape and packing.
• Ensure at least one vascular bundle (vein) is included in the diagram, showing its correct position relative to the other tissues.
• Add label lines drawn with a ruler, touching the structure being identified, and provide accurate labels for key tissues like ‘Upper epidermis’ or ‘Xylem’.

Question 2(a)(ii): High Power Drawing of Epidermal Cells

Now, using the same leaf slide P1 under high power, you need to draw a line of four adjacent epidermal cells. Which instruction below describes an incorrect technique or convention for this type of high-power biological drawing?

• Select four typical, adjacent epidermal cells that are clearly in focus and draw them joined together as they appear.
• Use sharp, continuous, unshaded lines for all outlines (cell walls, visible organelles if required). Avoid sketchy or thick lines.
• Represent the cell wall shared between two adjacent cells using only a single, thin ruled line.
• Draw the shapes and relative sizes of the chosen cells as accurately as possible, reflecting what is observed through the microscope.
• If labelling the cell wall, use a neat label line drawn with a ruler, touching the structure, and write the label horizontally.

Question 2(b)(i): Comparing Leaf Structures

Imagine Figure 2.1 shows a diagram of a transverse section of an aquatic plant leaf, notable for having large air spaces (aerenchyma)… and possibly stomata only on the upper surface. You compare this diagram to the actual leaf section on slide P1 (assume P1 is a typical terrestrial dicot leaf). Which statement describes an incorrect or unlikely observable difference?

• Fig 2.1 (aquatic) likely shows extensive, large air spaces within the mesophyll for buoyancy, whereas P1 (terrestrial) would have smaller, less prominent intercellular spaces primarily for gas diffusion.
• Slide P1 might exhibit epidermal hairs (trichomes) for protection or reducing water loss, which are typically absent or reduced in aquatic leaves like that in Fig 2.1.
• Both Fig 2.1 and slide P1 will show a clearly defined upper and lower epidermis tissue layer, indicating this is not a difference.
• Vascular tissue distribution might differ; the aquatic leaf (Fig 2.1) may have reduced vascular bundles compared to the more robust network needed for support and transport in the terrestrial leaf (P1).
• The cuticle on the epidermis is likely to be very thin or absent in the aquatic leaf (Fig 2.1) but noticeably thicker on the terrestrial leaf (P1) to prevent dehydration.

Question 2(b)(ii): Adaptation in Aquatic Leaf

Considering the described features of the aquatic leaf in the imaginary Figure 2.1 (e.g., large air spaces, stomata potentially on upper surface, thin cuticle)… which option below incorrectly states a visible feature and its adaptive function for life in water?

• Feature: Large internal air spaces (aerenchyma); Function: Provides buoyancy, helping the leaf float to access sunlight and air.
• Feature: Stomata present mainly or only on the upper epidermis; Function: Allows gas exchange (CO₂ uptake, O₂ release) directly with the atmosphere above the water surface.
• Feature: Thick, waxy cuticle covering the epidermis; Function: To prevent excessive water from entering and water logging the leaf tissues.
• Feature: Reduced vascular tissue (e.g., xylem); Function: Less structural support needed as water provides buoyancy, and less water transport needed as the leaf is surrounded by water.
• Feature: Finely divided or feathery leaves (if applicable); Function: Increases the surface area for absorption of dissolved gases and minerals directly from the water.

Question 2(c)(i) & (ii): Calculating Actual Size from Micrograph Measurement

Imagine Figure 2.2 is a micrograph showing several air spaces… magnification stated as x12. You measure lengths… L1=6 mm, L2=5 mm, L3=7 mm, L4=6 mm. Calculate the mean actual length… in µm. Which step below describes an incorrect calculation or procedure?

• Calculate the mean image length from the measurements: (6 + 5 + 7 + 6) mm / 4 = 24 mm / 4 = 6 mm.
• Recall the formula relating actual size (A), image size (I), and magnification (M): A = I / M.
• Calculate the mean actual size in mm using the formula: Mean Actual Size = (Mean Image Length) / Magnification = 6 mm / 12 = 0.5 mm.
• Convert the mean actual size from millimeters (mm) to micrometers (µm) by multiplying by 1000: 0.5 mm * 1000 µm/mm = 500 µm.
• Calculate the mean actual size by multiplying the mean image length by the magnification: Mean Actual Size = 6 mm * 12 = 72 mm.