Biology 121 Lab

CELLULAR RESPIRATION

What is CELLULAR respiration?

chemical E (glucose) + O2 → “biochemical currency” (ATP)

C6H12O6 + 6 O2 → 6 CO2 + 6 H2O + E

  Oxygen (O2) is ESSENTIAL for AEROBIC respiration…

  4 main steps…1 is common to both aerobic AND anaerobic respiratory pathways…

Aerobic vs. Anaerobic Respiration

  Aerobic   requires O2   4 main steps   yields up to 38 ATP glucose-1   obligate aerobes, facultative anaerobes

  Anaerobic   NO O2 required   1 main step   yields 2 ATP glucose-1   obligate anaerobes

The Mitochondrion

•  Glucose is broken down in the cytoplasm •  Kreb’s Cycle occurs in the matrix •  Electron transport occurs in/on the cristae

(envelope)

Aerobic Respiration

C6H12O6 + 6 O2 → 6 CO2 + 6 H2O + E

Step 1: Glycolysis (“glyco” “lysis”; cytoplasm)*

glucose → 2 pyruvate + 2 ATP + 2 NADH2 (6C) (3C)

cytoplasm

glycolysis

Step 2: Pyruvate Oxidation (mito matrix)

2 pyruvate → 2 Acetyl CoA + 2 CO2 + 2 NADH2 (3C) (2C)

pyruvate oxidation

matrix

Step 3: Kreb’s Cycle (aka TCA Cycle or Citric Acid Cycle; mito matrix)

2 Acetyl CoA + 2 Oxaloacetic Acid (2C) (4C)

2 Citric Acid + 4 CO2 + 2 ATP + 6 NADH2 + 2 FADH2 (6C)

Kreb’s Cycle

matrix

Step 4: Oxidative Phosphorylation (aka e- transport; mito cristae)

  NADH2 + FADH2 are involved with e – transport

  donate e- to carriers in the transport chain   pumping of H+ ions → [ ] gradient   generation of ATP

  O2 is the final e- acceptor oxidative phosphorylation

cristae

out

in

mito matrix

intermembrane space

NADH2 & FADH2

ATP synthase

Summary   3 ATP per NADH2 (x 10) (= 30; steps 1-3)   2 ATP per FADH2 (x 2) (= 4; step 3)   4 “substrate-level” ATP (= 4; steps 1 & 3)

38 TOTAL glucose-1

  Theoretical maximum = 38 ATP…no system is perfect!

∴ this number is rarely [if ever] achieved…

Anaerobic Respiration

glucose → 2 pyruvate + 2 ATP + 2 NADH2 (6C) (3C)

2 lactate 2 ethanol + 2 CO2 (= fermentation) (3C) (2C)

– animals – plants – microbes – microbes

demand > O2 release of metabolic poison

5.1 Respiration in Peas Protocol 5.1: Germinating vs. Non-germinating

⇒  Atmospheric/background CO2 level = 350-400 ppm

1.  Obtain 25 germinating peas & blot dry 2.  Place the peas in the respiration chamber 3.  Place the CO2 sensor in the chamber 4.  Wait 1 minute → begin collecting data for 5 minutes 5.  Measure & record the weight (g) of the peas 6.  Place the germinating peas in a beaker and place on ice for 5 minutes 7.  Follow the instructions on pg. 4

  determine the rate of respiration (slope, m = rate; ppm CO2 min-1)   store the data for comparison with other measurements

15.  Rinse and dry chamber 16.  Place the CO2 sensor in the chamber with non-germinating peas 17.  Wait 1 minute → begin collecting data for 5 minutes 18.  Follow the instructions on pg. 4

  Use a notebook to “fan” (i.e., clear) the sensor for 1 minute, returning the CO2 level to 300-400 ppm between EACH measurement!

5.2 Respiration in Peas Protocol 5.2: Room vs. Cold Temperature

1.  Empty the chamber by PUTTING THE NON-

GERMINATING PEAS BACK ON THE SIDE BENCH, and “clear” it…

2. Rinse and dry chamber

3. Repeat steps 1-7 (Protocol 5.1) using COLD germinating peas

5.3 Respiration in Crickets Protocol 5.3: Room vs. Cold Temperature

1.  Obtain 5-8 crickets & place in the respiration chamber 2.  Place the CO2 sensor in the chamber 3.  Wait 1 minute → begin collecting data for 5 minutes 4.  Measure & record the weight (g) of the crickets 5.  Place the crickets in the chamber on ice for 5 minutes (or until static) 6.  Follow the instructions on pg. 4

  determine the rate of respiration (slope, m = rate; ppm CO2 min-1)   store the data for comparison with other measurements

7.  Repeat steps 1-6 using COLD crickets 8.  Rinse & dry the respiration chamber when finished

  Use a notebook to “fan” (i.e., clear) the sensor for 1 minute, returning the CO2 level to 300-400 ppm between EACH measurement!

Do the results support your predictions? Peas   Germinating vs. Non-Germinating

  germinating > non-germinating   WHY?

  Room vs. Cold Temperature   room > cold   WHY?

Germinating

Germinating/COLD

Non-germinating

Do the results support your predictions?

Crickets   Room vs. Cold Temperature

  room > cold   WHY?

  What about PEAS vs. CRICKETS??   Why is it important to “normalize” by some

biological parameter (= fresh weight) for comparison?