many chemistry and biochemistry procedures would require you to perform
reaction at a specific P each to accomplish this task you must understand
how to both make a buffer
and how to use a pH meter this video but demonstrate
two methods but for preparation as well as proper use
a pH meter
a buffer is an eye and a compound that resists changes to its ph1 strong acids
or bases are added to it
the amount that's John acid a strong base that can be neutralized by a
particular but very
is dictated by the papers capacity but is determined by the concentration a
weak acid and weak base
in the buffer a higher concentration a buffering reagents
translates into greater buffering capacity in the first part of this video
we will walk through two methods a buffer preparation in the first example
you will need to prepare one leader other 50 millimolar buffer
for protein that is stable at pH 7.3
the first step is choosing an appropriate buffer system
this buffer system should produce a stable pH in the range needed
and should have a capacity large enough to accommodate your solutions
the week acid used but the system must have a PK that is plus or minus one unit
the desired pH the closer the better from this abbreviated this dove weak
acids
the fast I and with the PKF 6.82
is clearly the best choice for a buffer with the desired pH
up 7.3 therefore the week acid used for this system
its h2 PR for minus and the kind you get base
its HP 04 two-minus which correspond to the south
sodium phosphate monobasic and sodium phosphate dibasic
after determining an appropriate buffer system
the Henderson Hasselback equation is used to calculate the ratio
acid too can you get base needed at your desired pH
if your desired pH up 7.3
and the PK at the fast a tie and are substituted into this equation
than the ratio of conjugate base to acid is calculated to be 1.6
therefore here but we must contain 1.6 times the amount of sodium phosphate
dibasic
to sodium phosphate monobasic after determining the appropriate ratio of
conjugate base to acid
exact amount each reagent is calculated in the following way:
because you're planning to make one leader other 50 millimolar fast it back
for
at pH 7.3 50 millimolar is
equal to the concentration acid as the concentration a conjugate base
in addition you already know that the ratio a base to acid is 1.6
so the concentration a basis people to 1.6 times the concentration
acid by substituting equation 2 into equation 1
you can see that the concentration acid press one point six times the
concentration
acid is equal to 50 million dollar through manipulation at this equation
the concentration up acted needed for this particular but very
is nineteen point two million dollar and the concentration a base
is thirty-point eight million dollar which corresponds to 2.30
and 4.37 grams respectively for a one meter solution
to make the buffer simply dissolve 2.30 grams of sodium phosphate monobasic
and 4.37 grams of sodium phosphate dibasic
in one leader distilled water an alternative method a buffer preparation
is simple
accurate in requires much less Mac to demonstrate the second method
we will prepare one leader a one-point all millimolar chest
after at pH 7.6 0 the first step involves the determination
up the amount a weak acid or in this example week
base needed in the system to make one leader
a one-point il mio amore their chess batter 1.0 million miles
or 121 milligrams chess are required
at approximately eight hundred milliliters
distilled water as well as a stir by to the BK then
weigh 121 milligrams actress and transfer it to a one meter BK
ensure that I love the material has been transferred
by rinsing the way boat and transferring the Prince's into the BK
while staring monitor the pH of this solution
at a sodium hydroxide solution dropwise until the pH
is exactly 7.60 the concentration at the sodium hydroxide solution doesn't really
matter
however a solution that is to Delhi may require an excessive valium to be added
to the batter
while a solution that is too concentrated may cause you to overshoot
the desired pH
if you do happen to exceed the desired pH
than at a hydrochloric acid solution to bring the pH back down
transfer this solution to a one liter value magic flask
watch the beaker with distilled water several times and add these washings to
the value my check flask
David the buffer to the mic and that I am a chick flask
and mix the solution by inverting the flask several times
for the second method of preparation you needed the pH meter
the final part in this video what discussed the Packer you serve this
equipment
a pH meter allows the determination up hydrogen I an activity
by potentiometric measurements using a glass electrode
collects rejoined to a sober sober chloride reference electrode
when the probe is immersed in solution the reference electrode makes contact
with the sample
which completes the electrical contact between the reference electrode
sample and pH electrode if the unknown solution has a different pH of the
solution in the probe
and electrical potential results which is registered on the meter
this video will demonstrate the use than all Ryan essay 520
pH meter the important components have a pH meter include the display
the probe and the electrode tip the electrode tip contains a fragile glass
bulbs
be extremely careful not to hit the probe and the side of the beaker
and when using stir bars to apparate the pH meter
first make sure that the electrode is connected to the meter
then calibrate the pH meter the pH meter must be recalibrated
every ten you turn it on and after three to four hours abuse
we will perform a two-point calibration this method involves calibrating the pH
meter but
two different doctors have known pH
turn any instrument and select the pH mode as
.1 4.01
then press the I so button and verified that the pH read
7.00 Prince of the probe
with distilled water into a waste beaker and gently blot it with a kimwipe
place the probe into the first buffer and pressed the Cal button
the number one should appear in the display
to display what alternate between one and the pH value at the solution
wait until the display stabilizers and press enter
the number two should then appear on the display rents the probe with distilled
water
planet with the kimwipe and place it into the second buffer
if your solution has an acidic pH then use the 4.01 per
if it has a basic pH use the 10.01 buffer
the display will alternate between two and the pH value at the solution
wait until the display stabilizers and press Enter
the letters key age will be displayed indicating that the pH meter is now
ready to take measurements
before taking a measurement make sure that your sample buffer is at the same
temperature you'll be using in your experiments
because pH is temperature-dependent rents in black the probe and place it
into the buffer
wait for the display to stabilize a record the pH
correct the for preparation pH meter technique
are essential to ensure that your proteins and other sensitive chemicals
that long functional lives