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258 - physiology > LAB EXAM LAB 2: NADH dilution > Flashcards

LAB EXAM LAB 2: NADH dilution Flashcards

1
Q

true or false: there are many physiolgy sitautions where chemical events result in production of tandem products

A

true

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2
Q

how come you can take a blood samply and determine the amount of lactate in the blood due to a bout of exercise

A

because when a person is exercsing and the substrattes in the muscle (glucose, glycogen) are being metabolised for ATP for the muscles to contract, there may be a build uo of lactate in the msucle

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3
Q

true or false: measuring lactate is useful and cheap

A

false \

it is useful but it can be lengthy and costyl

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4
Q

what is the compound that is produced in tanden with pyruvate and in eeuqal concenttrations to lactate

A

NADH

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5
Q

is lactatet concentrtataion DIRECTLY measured by mreasuring the concentration of NADH

A

indirectly

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6
Q

what is the enxyme that cahnges lactate to pyruvate

A

lactate dehydrogen

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7
Q

what is the reaction of lactate to pyruvate

A

lactaute (c3H6o3) to Pyruvate (c6h4o3) by LADH

While NAD+ is reduced to NADh

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8
Q

how come lactate and pyruvate are simulat

A

lacttae is pyruvae with a pair of hudrrened added on

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9
Q

how can lactate concentration be dtermined

A

by transmorning any lactate in the blood back to pyruvate with the help of LDH and NAD+
(if add LDH and NAD+ to blood,, lactate will go to pyruvate)

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10
Q

what is the equip that allows concentration of lactate to be indirectly measured by measuring concentration of NADh

A

spectrophotometer

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11
Q

true or fasle: epct has the ability to measure concentration of lactate with easy where as it cannot directly measure concentration of NADH

A

false opposite

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12
Q

what is the use of the spect

A

to measure the absorption of light whena. beam passes through a sample

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13
Q

NADH absorbs at what wavelentgh

A

340 nm

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14
Q

do NAD+ and absorb light at 340 nm

A

no

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15
Q

what is the waavelength that the spect is set to

A

340 nm

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16
Q

the amount of light absrobed depends directly on what

A

on the amunt of NADH in the sample (which is proportionate to the amunt of lactae in orginal sample)

17
Q

True or false: NADH and NAD both absorb at 260

A

true

18
Q

spect cant only be ised tp assess the lactate/NADH concentration

A

false

it can be used to assess substrate=by product concentration of many different sotaution

19
Q

what is NADH

A

NADH is short for Nicotinamide Adenine Dinucleotide. It is a derivative of vitamin B
(niacinamide) combined with a ribose (5 carbon sugar), a phosphate group and an adenine
=coenzyme (found in cytosol and mito)

20
Q

what is the role of NADH

A

The role of NADH in cellular metabolism is directly linked to the role of NAD+. These
compounds are the most important carriers of the H+ atoms that are released during the breakdown of energetic substrates. This breakdown occurs in order to produce ATP,

which subsequently transports the H+ to the respiratory chain of electrons located in the inner membranes of the mitochondrion. There, the H+ are relinquished to the electron acceptors of the respiratory chain. This process is a major component of aerobic metabolism

21
Q

what is the difference between FAD and NAD

A

NAD and FAD are both electron (hydrogen) carriers.

is that FAD is able to accept and carry 2 hydrogen ions, whereas NAD can only accept 1 hydrogen ion.

FAD cannot produce more ATP than NAD since NAD enters the ETC earlier
the more ATP can be produced.

every NADH molecule, 3 ATP
every FADH2 molecule, 2 (ATP

22
Q

what is the goal of lab 2

A

Using a spect, generate a NADH standard
curve to determine the concentration of an
unknown sample of NADH.

23
Q

what is a spect

A

Used to ID a solutions’ components by measuring

the amount of light absorbed by that solution.

24
Q

what are 3 examples of things a spect can measure

A

glucose levels
chorlesterol levels
enxyme activites

25
Q

ehat yype of info can the spect provide about the tthings it measures

A

Healthy/Diseased
• Overall metabolic
profile
• Active/Inactive

26
Q

expkain how to use a micropupette

A 
Select the volume.
Set the tip.
Press and hold the plunger at the first stop.
Place the tip in the liquid.
Slowly release the plunger.
Pause for a second and then move the tip.
Insert the tip into the delivery vessel.
Press the plunger to the second stop.
27
Q

what is the main task of lab exercise 2

A

to generate a standard curve from the known NADH concentration solution (stock solution) and then use the curve to determine unknown NADH concentrion in the unknown sample

28
Q

how to generate a standard curve

A

must gtaph relationship between concetnration of a compound (NADH im this case) and the absorbance of loight incurred by solutions that contrain varrying concentrations of the same comppund must be generated

29
Q

what is a dilution factor

A

is a dulition factor of 4 is chosen (Doluted 4 times)

1 PART STOCK IS ADDED TO 3 PARTS DISTILLED WATER

30
Q

how much can the cuvette hold

A

3000 microliters (3ml)

31
Q

how to calculate the volume for 1 part

A

diveide the total volume by the dilution factor
ex: 3 ml/4 parts

0.75 ml/1 part

32
Q

how to calculate concentrations

A

divide the concentration of the stock. by the dilution factor !!
(EX: 100 mM/4= 25 mM (concnetration of new solution)

33
Q

true or false: you do not need to make replicates of the concentrations

A

false

important because it controls for pipetting errors

34
Q

once the concentrations diluted are prepared what must you do before placing them in cutte

A

wrap in parafilm and vortex

35
Q

in order to minimize contamination, what must you do to the cuvette between each concentration

A

rinse with distilled water

36
Q

before measuring the degree of light absorbed by each of the samples, what must you do to the spect

A

zero it with a blank sample

37
Q

how do you zero a spect adn whats the purpose

A

place duvette full of distilled water into the readig device and reset spect (to negative absorbance incurred by cuvette and water)

38
Q

measure and note the absorbances of the samples at what wavelength

A

340

39
Q

true or false: the hattch must remain open at all times

A

false do not leave it open at any other times besides when putting samples in cuvette

258 - physiology (21 decks)

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