Determination of norethynodrel in sea water

Determination of norethynodrel in sea water

Topic: Determination of norethynodrel in sea water

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Dear writer,

This assignment is in two parts. First part is a statistical part which I have done it, second part is as below: (also two examples will be provided "from lecture and from friend")(pictures and graphs also will be provided))

You must write a report concerning your data. The report will have an introduction, procedure, results, discussion and conclusion. The discussion must end in a conclusion regarding the comparitive values of hormone concentration at the different sites. It should also include a justification for the order of concentrations.

Determination of norethynodrel in sea water
Aim: To determine the concentration of norethynodrel in sea water by HPLC-MS.

Introduction
Norethynodrel is a progestin or synthetic progestagen which is used in oral contraceptive. There is significant concern that this and other hormonal based medications are effecting the breeding cycles of animals, in particular in the oceans. These chemicals are found in out domestic waste and pass through waste water treatment facilities (WWTF) into the water ways. This experiment determined the norethynodrel concentration in the sea water at three locations in major city (See map below, red dots indicate locations, red arrow indicates flow of water out of the bay and into the sea). Three replicate samples were taken from each of these locations, mouth of the river which passes through the city; near the outfall of the city’s WWTF and in a cove in the national park on the opposite side of the bay to the city.

Instructions
Click your results icon below to see the correct answers to the statistical analysis of your data. Also included are graphs of hormone and d-hormone calibration curve and the residuals for the regression analysis and map of bay, all downloadable.
You must write a report concerning your data. The report will have an introduction, procedure, results, discussion and conclusion. The discussion must end in a conclusion regarding the comparitive values of hormone concentration at the different sites. It should also include a justification for the order of concentrations.

Determination of norethynodrel in sea water
Aim: To determine the concentration of norethynodrel in sea water by HPLC-MS.
Results
Click here to download a file which contains the correct calculations for Part 1.
Click here to download a picture of the calibration curve for the norethynodrel (right click on picture in new window to save)

Click here to download a a picture of the residual plot for the norethynodrel (right click on picture in new window to save)

Click here to download a picture of the calibration curve for the d-norethynodrel (right click on picture in new window to save)

Click here to download a picture of the residual plot for the d-norethynodrel (right click on picture in new window to save)

Click here to download the map in the introduction (right click on picture in new window to save)

Introduction
Norethynodrel is a progestin or synthetic progestagen which is used in oral contraceptive. There is significant concern that this and other hormonal based medications are effecting the breeding cycles of animals, in particular in the oceans. These chemicals are found in out domestic waste and pass through waste water treatment facilities (WWTF) into the water ways. This experiment determined the norethynodrel concentration in the sea water at three locations in major city (See map below, red dots indicate locations, red arrow indicates flow of water out of the bay and into the sea). Three replicate samples were taken from each of these locations, mouth of the river which passes through the city; near the outfall of the city’s WWTF and in a cove in the national park on the opposite side of the bay to the city.

In this experiment, 100 litres of sea water was collected from each site. Three (3) 20 litre aliquots were taken from each of these 100 litre samples. Each aliquot was spiked with one (1) litre of 2.1014 ppm d-norethyndrel solution. d-norethyndrel is the deuterated analog of norethyndrel and is used in this experiment to determine the efficiency of the ethyl acetate extraction. The sub-sample was then passed through an extraction filter tube which collected all the organic material. This concentrates the organic material into the smaller organic extract. The organic compounds were then extracted from the filter tube with ethyl acetate. A new filter tube was used for each sub-sample. The ethyl acetate extract was collected in a 5-ml volumetric flask and made to volume with ethyl acetate. This extract was analysed by HPLC-MS. The individual organic compounds in the ethyl acetate extract were separated by HPLC and each separated species and its concentration analysed by MS. The peaks due to norethyndrel and d-norethyndrel were identified and measured. Each extraction was analysed 4 times by HPLC-MS.
17b-Methyltestosterone was used as an internal standard of the HPLC-MS. The results presented below are the normalised area of norethyndrel and d-norethyndrel, adjusted for this internal standard.
Calibration solutions
Ten solutions of known concentration of norethyndrel in ethyl acetate were prepared by dilution of a standard solution. Nine solutions of known concentraton of d-norethyndrel in ethyl acetate were prepared by dilution of a standard solution. A blank solution was also prepared. SIMS (Selective Ion Mass Spectroscopy) was used to distinguish the peak areas of the norethyndrel and d-norethyndrel in the chromatographs.
Each calibration solutions of norethyndrel and d-norethyndrel were recorded 4 times.
There are NO graphic outliers in the calibration curves of norethyndrel or d-norethyndrel
Results
Calibration curve – norethyndrel
Solution (ppm)    Normalised peak area (SIMS)    Standard Deviation
Blank (0 ppm)    0    40
40    15968    52
120    37082    106
240    83925    241
360    115351    345
480    163732    508
640    213886    678
800    262113    714
1000    336636    1059
1200    396558    1076
Calibration curve – d-norethyndrel
Solution (ppm)    Normalised peak area (SIMS)    Standard Deviation
Blank (0 ppm)    1563    39
250    49976    149
300    59040    178
325    64416    177
350    69352    196
375    73739    203
400    78296    229
450    88209    244
800    155508    434
Samples
Sample    Normalised peak area – norethyndrel (SIMS)    Normalised peak area – d-norethyndrel (SIMS)
WWTF – No. 1    328063,332917,328534,330187    79911,76924,77003,77172
WWTF – No. 2    314796,316131,330628,315374    76528,77313,76036,76882
WWTF – No. 3    317200,317906,316699,316532    77548,77948,76697,77559
National Park – No. 1    34810,35358,34896,35500    78031,76844,76926,77356
National Park- No. 2    34214,34276,34524,34789    77764,76981,76966,76659
National Park- No. 3    41183,41877,41420,41806    77558,78181,78053,76709
River – No. 1    79879,81218,80864,79737    76492,77006,76598,77124
River – No. 2    82788,83002,82224,83674    77730,77344,78150,77654
River – No. 3    83556,84677,84852,84411    78260,79122,78780,80035
(SIMS = Selective Ion Mass Sepectroscopy)
Calculate the concentration of norethyndrel in each sea water sample in ug/litre and hence the average concentration in these areas. In your discussion, comment on how the results were obtained (statistical analysis). Discuss the different levels found at the sites in the bay and suggest an explanation for these differences.
Click here to download a file which contains all these values

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