Uploaded by mrphysh on 11.07.2011

Transcript:

The consultants at the Wal-Mart site send in a sample for 8270

they get a result back that tells them that they have 70 parts per billion 2-methylnaphthalene

where does that number come from?

one way to approach it is this: we take a liter .... this is like a big graduated cylinder

the liter mark is right here.

we fill it up... we put in 200 mg of 2-methylnaphthalene

200 mg in a liter is two hundred parts per million

this implies that there's 1,000,000 mg in a liter, which in fact, there is

200 mg is 2/10 of gram which is spatula full... it's not difficult to weigh.

our sample comes in, in a small vial .... we subject a microliter of the sample

from the site to our chromatographic analysis

and we get a response factor of 237

we subject a microliter of our standard to our chromatographic analysis

and get response factor of 1239

the way I think of it this is:

I say

"okay, start with the 200 multiply by little number and divide by the big number"

and we find out that our sample is 70.5 ppm 2-methylnaphthalene.

The 8270 list has 70 compounds

it's more than just 2-methylnaphthalene so the way to.... using the same approach

would be to simply put 200 mg of phenol, 200 mg of indene, 200 mg of dinitrophenol,

200 mg of the 2-hexylethylphthalate, 200mg of di-n-octylphthalate

200 mg of benezidine 200 mg of straight naphthalene etc. etc. etc.

and then we would end up with in the same approach.. will now quantity for all 70 compounds

a kind of important little..observation is that, as we add 200 mg of this

200 mg this 200 mg this our volume is going to go up over a liter

we don't want the volume over the over a liter

we want the volume to stay at one liter

which means that in actual practice we add

200 mg 200 mg 200 mg and then bring it up to volume

at one liter so it stays in one liter

we have our standard at 200

we want to make a calibration curve with this 200

the calibration curve is at 20, 50, 80,120 and 160

so we simply dilute 200 down to these five levels

it's pretty straightforward

we need to pick a volume

let's take a volume that will fit in these jars.

it's arbitrary... let's say 25 mls

so we want to dilute our 200 standard down to 160

and we want 25 mls total

so we go to our concentration times volume equals concentration times volume

and 25 mls total

so we do the calculations and we find that

we want to put five mls of this plus 20 mils of the solvent (methylene chloride)

for the 160

for the 120 we want 10 plus 15 ....for the 80 we want 15 plus 10

for the 50 6.5+18.75 mils

for the 20 we want... this is pretty diluted now

2.5 mils this and 22.5 mils of the solvent

and this will give us our standard

so ..now to run the initial calibration

we simply run an injection of this , an injection of this , an

injection of this and a injection of this

I know when I....I wasn't a very smart kid and I would be confused by this big jar these little jars

I would go

"but wait a minute.. it this is a big jar and this is a small jar""

it's one microliter of each no matter what you do............ so the size of the container doesn't matter

what you do

Like I say, I wasn't a very smart kid

this is an approach which will work.

I see no scientific, particular reason why this approach wouldn't work

there are problems with it. It is never actually done this way.

the problems are related to the chemicals themselves

the 8270 is a list of hazardous waste.

The compounds on the 8270 list are carcinogenic and toxic

these are like bad guys

you don't go out and buy a container of 2-methylnaphthalene.

you don't go buying benezidine

let's buy some 2-nitrophenol

no.... you don't buy this stuff

anything that comes in to the lab is eventually

going to be toxic waste

any... any hazardous compound that's produced

ends up as hazardous waste

so you don't want this stuff hanging around your lab

and then another factor is that a container of this at 200ppb

would last a laboratory about 100 years

you use a microliter at a time

and another point, and this is on a technical , scientific side

is that the standards are unstable when they're mixed.

most of the 8270 compounds are very stable chemically but some of them are not

and when they're mixed together

some of the compounds will combine with other compounds

and the result of this is that the standards

are not stable when they are mixed

you mix them and they're good

the next day they're good, the third day ...they're probably still good

but a standard that is a this week old ....they start to break down and you really can't use them when they get old

you have to be constantly making them up fresh

the way this works in a real laboratory is the standards are purchased at 2000 ppm

they come in the mix so that they come purchased and diluted 2000 ppm

but there separated such that they will be stable

in the mix

they are not stable when they're mixed together but they're separated and put in groups that will be stable together like for example the

benezidine is separated all by itself

the phenols will all be together

the PAHs will all be together

the mix consists of five very small break off vials. The five vials together form the 8270 calibration curve

the way this starts out is that the analyst gets the five break off vials and carefully labels five autosampler vials

the autosampler vials ....the most important label

is the 'primary control number' to go into the log for the EPA ..

this EPA mandated and everything has to be documented

So the first thing you do is document the standards of the EPA

they are given an ascension number on the jar and

then an ascension number in a book

and so it would have ascension number and then maybe a label for what it is or not

and in the description goes into the book

so the vials are broken off, transferred into the appropriate vial.... the first one's broken

off transferred

the second one is broken off.... transferred

third one's broken off.... transferred

fourth one's broken off..... transferred

fifth one's broken offf... transferred

it has to be done such that they don't get contaminated

we have these five standards

we want to dilute them down to make a calibration curve

and we want them to have half a milliliter total

remember we picked the 25 mils

we said "well let's make 25 mils"

this is like.... has to be a half a ml.

so we want to have a half a ml of these guys mixed together for 20

we want to have half a ml of these guys mixed together for a 50

a half a ml of these guys mixed together for an 80

a half a ml these guys mixed together for 120

and you have to have half a ml mixed together for 160

so how are we going to do this?

let's start with our 20

we want to make a 20 standard and

we want a half a ml total and want to put it in one of these

okay we want 20 standard

we want 20 standard and we want it made.....

these guys diluted all appropriately such that it comes out as 20 part per million these are at 2000 ppm

we want to dilute these five guys down equally

so that when we are done we will have 20 ppm and a half a mill total

okay this is how works

concentration times volume equals concentration times volume

we want 0.5 mls as the final volume

we want 20 ppm as the final concentration

we are starting out with 2000 ppm and the volume is our variable

so we do the calculations and we find the volume to be .005 milliliters.

"0.005 milliliters!"

.005 milliliters is 5 microns... that's not a problem

we have this syringe that will do 5 µL

so we put 5 µL of this one in it

5 µL of this one in it

5 µL of this one in it

5 µL of this one in it and we're done with that one.

set that one aside

okay we need one that's 50 ppm

so we want a vial that's 50 ppm of each of these 50 ppm and

we want .5 mL when we are done

so let's take a look at this

concentration times volume equals concentration times volume our final volume is .5 mils

our final concentration is 50 ppm

our initial concentration is 2000 ppm

how much do we need to add

well... we end up with 0.0125 mL that's 12.5 µL ....not a problem

we take this guy,,, put 12.5 µL from this one ,

12.5 µL from this one

12.5 µL of this

12.5 µL from this one in here and we're done

moving forward... we want to make an 80

take a vial,...label it..... this also needs to go in your EPA standards control log

incidentally this needs to be logged in also

label it as an 80 opportunity how much do we need...well same thing

concentration times volume equals concentration times volume

80 x .5 ml concentration times volume 2000 ppm what's our volume going to be? Our volume is going to be 0.020 which is 20ul

so we put in 20 µL of this

20 µL of this

20 µL of this

20 µL of this

20 µL of this and we're done .....so go on to the next one

120 which is 30 µL so 30 ul of each one

and our 160 is 40 µL each one

okay labeled and everything

and we end up with their five points

we've added each of the five to each of our five standards so we're done with these ...cap these off real good

put them away somewhere where they're safe... they're expensive

and we have our 20, our 50, our 80, our 120, and our 160

these are all nicely labeled

they go in the our control log for the EPA

but we are not quite done with them are we.

we need to have a half a ml for each one so

so 160 ....we have added 200 we need to add 300 to dilute it up to 500 ul

in theory you could bring it up to volume at a half a ml

but in actual fact you really can't do that

it you need to calculate and add it

so we add 300ml to the "160"

so we add 350 mils to the "120"

we add 400 mils to the "80"

we add 437 mils to the "50" and we add 475 mls to the "20"

and then we are done

let me give you a quick walk-through on how to actually do this

The way this is actually done is..... I think this is kind of important

it's a trick but it's not anything that's completely insurmountable

so.... starting from scratch let me give it to you

as a lab tech... as a veteran lab tech

let me give you the technique you use

you get a solid displacement pipette

start with the ... 20...

solid displacement pipette

turn it so that it will give you 5 µL (microliters)

we take the first one

draw 5 µL ....put it in the first vial, labeled of course,

go through a series of rinses

the rinses are dirty, not so dirty clean clean super clean

so you go through it (noise, noise etc) and you're done with the first one

take out the second one...take out 5 µL ...add it ...

go through the series of rinses..... take the next one

add it...go to the series of rinses

take the next one

add it ...go through the series of rinses

take the last one .....add it ....go through the series of rinses

take the last one, add it

go through the series of rinses

okay so this one's done

take your 50

you want to dial the 50 to 12.5

dial your solid displacement pipette to 12.5

go 50 ( noise) go through a series of rinses

go 50 (noise) go to your series of rinses

50 go through your series of rinses

50 go through your series of rinses

50 go through your series of rinses

I like solid displacement pipettes because they're easy to clean

you can just reuse the same sleeve

just clean it which you really can't do very well

the alternative would be to use a disposable tip

And then the way to add the methylene chloride

to bring it up to volume at .5ml

is to get a pipette,, automated,, adjustable, pipette with 1 mL,

dial in the first volume you want

air out the pipette

the methylene chloride is very volatile

and it will tend to want a blow out

of the pipette and mess up your volumes

so let it come to equilibrium

the whole.... let the pipette come to equilibrium

and then do the first one

punch in a new number....do the second one

punch in a new number....do the third one

punch in a new number....do the fourth one

punch in a new number....do the last one

and throw the pipette tip away

it's all methylene chloride

so you don't need to use bunch of different tips

so we have our five points

we have our 20 50 80 120 160

we get some samples in to run.

so we line them up

we have a prep blank it has to have half of a ml

we have three samples.... they each have a half a mill in them

and we have a couple of spiked samples.... a couple of quality control samples

they each have a half a ml in them

in the half a milliliter later is extremely important

the half a milliliter is not an approximate half a milliliter

It is exactly at half a milliliter

and you should be able to look across them and see that they all have the exact same volume

exact.... exact same volume

it's not any great trick

but it's important to understand

that it is very important that the volumes be right

now the last step is.... we take a ...an internal standard

the internal standard comes at 4000 ppm

and we put 5 µL in

each one so we put 5 µL of internal standard

in each one of them

remember that the internal standard quantitation

is based upon the ratio of the internal standard to the total volume

so the volume has to be exactly .5

and the internal standard has to be exactly 5 µL

so be careful..... this is a good time to be as careful as you can

And now ... like next week.... you need make a continuing calibration standard at 50

just follow the same directions

take a vial, put in 12.5 uL of each one

add 437.5 µL to bring it up to 500

add 5 µL of internal standard ...exactly of the same internal standard

and you can hit the iCal every time

if the other variables are correct

if the GC is working properly you can hit the iCal every time

it's.....its fairly ....it's actually pretty satisfying

the initial calibration is complicated

but when it comes out... which it will... if you're careful

it's cool

it's cool

they get a result back that tells them that they have 70 parts per billion 2-methylnaphthalene

where does that number come from?

one way to approach it is this: we take a liter .... this is like a big graduated cylinder

the liter mark is right here.

we fill it up... we put in 200 mg of 2-methylnaphthalene

200 mg in a liter is two hundred parts per million

this implies that there's 1,000,000 mg in a liter, which in fact, there is

200 mg is 2/10 of gram which is spatula full... it's not difficult to weigh.

our sample comes in, in a small vial .... we subject a microliter of the sample

from the site to our chromatographic analysis

and we get a response factor of 237

we subject a microliter of our standard to our chromatographic analysis

and get response factor of 1239

the way I think of it this is:

I say

"okay, start with the 200 multiply by little number and divide by the big number"

and we find out that our sample is 70.5 ppm 2-methylnaphthalene.

The 8270 list has 70 compounds

it's more than just 2-methylnaphthalene so the way to.... using the same approach

would be to simply put 200 mg of phenol, 200 mg of indene, 200 mg of dinitrophenol,

200 mg of the 2-hexylethylphthalate, 200mg of di-n-octylphthalate

200 mg of benezidine 200 mg of straight naphthalene etc. etc. etc.

and then we would end up with in the same approach.. will now quantity for all 70 compounds

a kind of important little..observation is that, as we add 200 mg of this

200 mg this 200 mg this our volume is going to go up over a liter

we don't want the volume over the over a liter

we want the volume to stay at one liter

which means that in actual practice we add

200 mg 200 mg 200 mg and then bring it up to volume

at one liter so it stays in one liter

we have our standard at 200

we want to make a calibration curve with this 200

the calibration curve is at 20, 50, 80,120 and 160

so we simply dilute 200 down to these five levels

it's pretty straightforward

we need to pick a volume

let's take a volume that will fit in these jars.

it's arbitrary... let's say 25 mls

so we want to dilute our 200 standard down to 160

and we want 25 mls total

so we go to our concentration times volume equals concentration times volume

and 25 mls total

so we do the calculations and we find that

we want to put five mls of this plus 20 mils of the solvent (methylene chloride)

for the 160

for the 120 we want 10 plus 15 ....for the 80 we want 15 plus 10

for the 50 6.5+18.75 mils

for the 20 we want... this is pretty diluted now

2.5 mils this and 22.5 mils of the solvent

and this will give us our standard

so ..now to run the initial calibration

we simply run an injection of this , an injection of this , an

injection of this and a injection of this

I know when I....I wasn't a very smart kid and I would be confused by this big jar these little jars

I would go

"but wait a minute.. it this is a big jar and this is a small jar""

it's one microliter of each no matter what you do............ so the size of the container doesn't matter

what you do

Like I say, I wasn't a very smart kid

this is an approach which will work.

I see no scientific, particular reason why this approach wouldn't work

there are problems with it. It is never actually done this way.

the problems are related to the chemicals themselves

the 8270 is a list of hazardous waste.

The compounds on the 8270 list are carcinogenic and toxic

these are like bad guys

you don't go out and buy a container of 2-methylnaphthalene.

you don't go buying benezidine

let's buy some 2-nitrophenol

no.... you don't buy this stuff

anything that comes in to the lab is eventually

going to be toxic waste

any... any hazardous compound that's produced

ends up as hazardous waste

so you don't want this stuff hanging around your lab

and then another factor is that a container of this at 200ppb

would last a laboratory about 100 years

you use a microliter at a time

and another point, and this is on a technical , scientific side

is that the standards are unstable when they're mixed.

most of the 8270 compounds are very stable chemically but some of them are not

and when they're mixed together

some of the compounds will combine with other compounds

and the result of this is that the standards

are not stable when they are mixed

you mix them and they're good

the next day they're good, the third day ...they're probably still good

but a standard that is a this week old ....they start to break down and you really can't use them when they get old

you have to be constantly making them up fresh

the way this works in a real laboratory is the standards are purchased at 2000 ppm

they come in the mix so that they come purchased and diluted 2000 ppm

but there separated such that they will be stable

in the mix

they are not stable when they're mixed together but they're separated and put in groups that will be stable together like for example the

benezidine is separated all by itself

the phenols will all be together

the PAHs will all be together

the mix consists of five very small break off vials. The five vials together form the 8270 calibration curve

the way this starts out is that the analyst gets the five break off vials and carefully labels five autosampler vials

the autosampler vials ....the most important label

is the 'primary control number' to go into the log for the EPA ..

this EPA mandated and everything has to be documented

So the first thing you do is document the standards of the EPA

they are given an ascension number on the jar and

then an ascension number in a book

and so it would have ascension number and then maybe a label for what it is or not

and in the description goes into the book

so the vials are broken off, transferred into the appropriate vial.... the first one's broken

off transferred

the second one is broken off.... transferred

third one's broken off.... transferred

fourth one's broken off..... transferred

fifth one's broken offf... transferred

it has to be done such that they don't get contaminated

we have these five standards

we want to dilute them down to make a calibration curve

and we want them to have half a milliliter total

remember we picked the 25 mils

we said "well let's make 25 mils"

this is like.... has to be a half a ml.

so we want to have a half a ml of these guys mixed together for 20

we want to have half a ml of these guys mixed together for a 50

a half a ml of these guys mixed together for an 80

a half a ml these guys mixed together for 120

and you have to have half a ml mixed together for 160

so how are we going to do this?

let's start with our 20

we want to make a 20 standard and

we want a half a ml total and want to put it in one of these

okay we want 20 standard

we want 20 standard and we want it made.....

these guys diluted all appropriately such that it comes out as 20 part per million these are at 2000 ppm

we want to dilute these five guys down equally

so that when we are done we will have 20 ppm and a half a mill total

okay this is how works

concentration times volume equals concentration times volume

we want 0.5 mls as the final volume

we want 20 ppm as the final concentration

we are starting out with 2000 ppm and the volume is our variable

so we do the calculations and we find the volume to be .005 milliliters.

"0.005 milliliters!"

.005 milliliters is 5 microns... that's not a problem

we have this syringe that will do 5 µL

so we put 5 µL of this one in it

5 µL of this one in it

5 µL of this one in it

5 µL of this one in it and we're done with that one.

set that one aside

okay we need one that's 50 ppm

so we want a vial that's 50 ppm of each of these 50 ppm and

we want .5 mL when we are done

so let's take a look at this

concentration times volume equals concentration times volume our final volume is .5 mils

our final concentration is 50 ppm

our initial concentration is 2000 ppm

how much do we need to add

well... we end up with 0.0125 mL that's 12.5 µL ....not a problem

we take this guy,,, put 12.5 µL from this one ,

12.5 µL from this one

12.5 µL of this

12.5 µL from this one in here and we're done

moving forward... we want to make an 80

take a vial,...label it..... this also needs to go in your EPA standards control log

incidentally this needs to be logged in also

label it as an 80 opportunity how much do we need...well same thing

concentration times volume equals concentration times volume

80 x .5 ml concentration times volume 2000 ppm what's our volume going to be? Our volume is going to be 0.020 which is 20ul

so we put in 20 µL of this

20 µL of this

20 µL of this

20 µL of this

20 µL of this and we're done .....so go on to the next one

120 which is 30 µL so 30 ul of each one

and our 160 is 40 µL each one

okay labeled and everything

and we end up with their five points

we've added each of the five to each of our five standards so we're done with these ...cap these off real good

put them away somewhere where they're safe... they're expensive

and we have our 20, our 50, our 80, our 120, and our 160

these are all nicely labeled

they go in the our control log for the EPA

but we are not quite done with them are we.

we need to have a half a ml for each one so

so 160 ....we have added 200 we need to add 300 to dilute it up to 500 ul

in theory you could bring it up to volume at a half a ml

but in actual fact you really can't do that

it you need to calculate and add it

so we add 300ml to the "160"

so we add 350 mils to the "120"

we add 400 mils to the "80"

we add 437 mils to the "50" and we add 475 mls to the "20"

and then we are done

let me give you a quick walk-through on how to actually do this

The way this is actually done is..... I think this is kind of important

it's a trick but it's not anything that's completely insurmountable

so.... starting from scratch let me give it to you

as a lab tech... as a veteran lab tech

let me give you the technique you use

you get a solid displacement pipette

start with the ... 20...

solid displacement pipette

turn it so that it will give you 5 µL (microliters)

we take the first one

draw 5 µL ....put it in the first vial, labeled of course,

go through a series of rinses

the rinses are dirty, not so dirty clean clean super clean

so you go through it (noise, noise etc) and you're done with the first one

take out the second one...take out 5 µL ...add it ...

go through the series of rinses..... take the next one

add it...go to the series of rinses

take the next one

add it ...go through the series of rinses

take the last one .....add it ....go through the series of rinses

take the last one, add it

go through the series of rinses

okay so this one's done

take your 50

you want to dial the 50 to 12.5

dial your solid displacement pipette to 12.5

go 50 ( noise) go through a series of rinses

go 50 (noise) go to your series of rinses

50 go through your series of rinses

50 go through your series of rinses

50 go through your series of rinses

I like solid displacement pipettes because they're easy to clean

you can just reuse the same sleeve

just clean it which you really can't do very well

the alternative would be to use a disposable tip

And then the way to add the methylene chloride

to bring it up to volume at .5ml

is to get a pipette,, automated,, adjustable, pipette with 1 mL,

dial in the first volume you want

air out the pipette

the methylene chloride is very volatile

and it will tend to want a blow out

of the pipette and mess up your volumes

so let it come to equilibrium

the whole.... let the pipette come to equilibrium

and then do the first one

punch in a new number....do the second one

punch in a new number....do the third one

punch in a new number....do the fourth one

punch in a new number....do the last one

and throw the pipette tip away

it's all methylene chloride

so you don't need to use bunch of different tips

so we have our five points

we have our 20 50 80 120 160

we get some samples in to run.

so we line them up

we have a prep blank it has to have half of a ml

we have three samples.... they each have a half a mill in them

and we have a couple of spiked samples.... a couple of quality control samples

they each have a half a ml in them

in the half a milliliter later is extremely important

the half a milliliter is not an approximate half a milliliter

It is exactly at half a milliliter

and you should be able to look across them and see that they all have the exact same volume

exact.... exact same volume

it's not any great trick

but it's important to understand

that it is very important that the volumes be right

now the last step is.... we take a ...an internal standard

the internal standard comes at 4000 ppm

and we put 5 µL in

each one so we put 5 µL of internal standard

in each one of them

remember that the internal standard quantitation

is based upon the ratio of the internal standard to the total volume

so the volume has to be exactly .5

and the internal standard has to be exactly 5 µL

so be careful..... this is a good time to be as careful as you can

And now ... like next week.... you need make a continuing calibration standard at 50

just follow the same directions

take a vial, put in 12.5 uL of each one

add 437.5 µL to bring it up to 500

add 5 µL of internal standard ...exactly of the same internal standard

and you can hit the iCal every time

if the other variables are correct

if the GC is working properly you can hit the iCal every time

it's.....its fairly ....it's actually pretty satisfying

the initial calibration is complicated

but when it comes out... which it will... if you're careful

it's cool

it's cool