Welcome back to the Human Anatomy and Physiology lab! This video describes the protocol that we carry in this lab at USF to investigate the digestion of fat by pancreatin and to test for the presence of any signs of digestion under different experimental parameters, including the addition of bile salts.
What’s up everyone it’s nails again and in this video i will show you the protocol that i will be using to investigate the digestion of fat by lipase this video is part of a three part series on the digestion of food stuff through enzymatic actions so make sure to watch my other videos to learn more about search and protein digestion there is quite a bit of chemistry
Involved with this protocol and as you will see there’s a number of steps that you will have to pay attention to to ensure the success of this experiment alright enough talking let’s get started first this is what you will need to perform this experiment a set of seven disposable culture tubes a sharpie to write the id of each tube di water a solution of 1% then
Creating some litmus creme which is basically a whipping cream to witch litmus powder has been added to turn the cream blue and some bile salts in addition to this you will need some disposable pipettes a clamp to handle the tubes when using the hot plate a microspatula and some ice here i’m following the protocol described in the human emp lab manual by mariam
And smith as usual you will also need a lab coat some gloves and a pair of glasses before doing anything i would highly recommend turning on your water bath this thing can take a little while to get up to temperature and turning it on right now will save you some time here we want to set it to 37 degrees celsius also you may want to turn on your hot plate since we
Will be using it shortly now we can label all the tubes so we can identify them when we prepare the different solutions according to the protocol described in the manual we want to label the first five tubes 1 l 2 l and up to 5 l the last two tubes should be labeled for b and 5 beats each tube represents a specific treatment including 2 controls and 5 tests now
That everything is labeled let’s proceed to the experiment in tube 1l you want to add about five drops of di water and five drops of been created and just for good measure don’t forget to cap your vial to prevent accidental contamination moving on to tube to l the second control add five drops of di water and five drops of flip new screen in both tubes 5l and 5b
Add five drops of paint created because we will incubate these tubes in ice we won the paint creating to be as cold as possible before adding litmus cream and vitals that’s why we partially prepare them in advance before we get back to them once all the other tubes are ready now we can prepare to three l’s add five drops of paint create into it and then if water
Is boiling on your hot plate grab the tube with the clamp and drop it in the boiling water let it sit for about five minutes meanwhile we can prepare to 4l add 5 drops of litmus cream and 5 drops up and created and as usual don’t forget to cap the tube we can also prepare tube for b by also adding 5 drops of pay plus cream and 5 drops of paint creating the only
Difference with tube 4l is that we will also add a pinch of bile salt to it once added make sure to stir the solution a bit to mix all the chemicals it’s been five minutes now so we can turn the hot plate off and we can take to brielle out of the boiling water to put it back in the holder tube 3 l is almost complete add five drops of whitmer screen to it and cap
It remember tubes 5 l and 5 b now that they are nice and cold we can take care of them into 5 l add three drops of liquid screams and like the other tubes don’t forget to cap it into 5 b also add 5 drops of litmus creme but unlike tube 5 l we will also add a pinch of bile salt to it once added make sure to stir the solution a bit to mix all the chemicals and
Put the tube back into the ice we can finally proceed to leaving all the tubes except you 5a and 5b in the water bath for about one hour all right it’s been about an hour now so we can take all of our tubes out of the water bath to look at the results oh and don’t forget to bring the tube sitting in the ice along wow those results look excellent check this out we
Can clearly see that some of the solutions turned pink or red oil others remain blue also in the tube that turned to a vivid red we can see that some sort of a foam formed at the surface let’s quickly discuss those results tube when l was a control containing paint creating only so it is totally normal that it shows no color at all since the color would come from
The litmus creep tube 2 l which was also a control only contain litmus cream so in the absence of an enzyme to digest the fat containing the cream the ph was not expected to change and as a result the solution remained blue in tube 3 l we initially boiled the pancreatic this in nature the enzyme which was therefore not capable of digesting the fast present in the
Cream that we eventually added later and as a consequence the solution remained blue thus indicating no change in the ph of the solutions and no digestion moving on the tube for l we see a moderate positive test for the digestion of the cream by pancreatic this makes perfect sense since we incubated this tube at 30 degrees celsius which turns out to be around the
Optimal temperature for pancreatin to act upon fat however the lack of an emulsifier made the digestion of the fat containing the cream difficult for the enzyme thus resulting in an incomplete digestion within the incubation period of one hour that we used then into 5 l we see that the solution remained mostly blue with a very faint hint of think this is indicative
Of an almost entirely incomplete digestion of the fat present in the cream that resulted in a very mild change in the ph of the solution this can be explained by the fact that at 0 degree celsius pancreatin is not as effective at digesting fat as it is at 37 degrees moving on to tube for b where all the action happened the solution turned to a very vivid red thus
Indicating a strong decrease in its ph that resulted from the complete digestion of the fat containing the cream also i like to pour l we see that the addition of bile salts allowed for the emulsification of fat sauce facilitating digestion last but not least until 5 li we see that the solution took a moderately pink tint thus showing a mildly positive test for
The digestion of fat these can be explained by the fact that even when bile salts were present as zero degrees celsius pink region is not as effective at digesting fat as it is at 37 degrees that’s pretty much it for this video hopefully you managed to follow along and to obtain the same great results by the way don’t forget to check out my other videos if you want
To learn about starch and protein digestion see you next time
Transcribed from video
Fat digestion by pancreatin and bile By Nils at USF
