Fundamental Techniques and Measurements Mass Measurements Volume Measurements

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Fundamental Techniques and Measurements Ø Mass Measurements Ø Volume Measurements Ø Preparation of a

Fundamental Techniques and Measurements Ø Mass Measurements Ø Volume Measurements Ø Preparation of a solution of known concentration Ø UV-Visible Spectrophotometer

Electronic Balance Ø What does an electronic balance measure? _____ force Ø If you

Electronic Balance Ø What does an electronic balance measure? _____ force Ø If you took an electronic balance with a capacity of 100 g to the moon what would its range be? _____ 600 g

Mass: Electronic Balance Ø Accuracy Ø 4 to 6 significant digits ä Sources of

Mass: Electronic Balance Ø Accuracy Ø 4 to 6 significant digits ä Sources of error ä Ø Calibration Ø Use known mass Ø Check weekly or when balance is moved ä ä ä Balance must be calibrated and maintained in same orientation in _____ field gravity hydroscopic chemicals: dry to constant mass first (will increase in mass rapidly as they reabsorb water on the balance!) When preparing a solution of a given concentration it may be difficult to get report actual! the exact mass desired_____ evaporation of wet samples

Electronic Balance M o d el D I-100 D I-800 D I-5000 Cap acity

Electronic Balance M o d el D I-100 D I-800 D I-5000 Cap acity Reso lutio n 100 g 0. 0001 g 800 g 0. 01 g 5000 g 0. 1 g Ø For maximum accuracy use balance with lowest _______ capacity possible! Ø Don’t forget to clean the balance if you spill any chemicals!!!!!!

Volume Ø Volumetric flask 0. 16 m. L Øaccuracy of ______/100 m. L Ø

Volume Ø Volumetric flask 0. 16 m. L Øaccuracy of ______/100 m. L Ø Graduated cylinder 1 m. L Øaccuracy of ______/100 m. L Ø Beaker 5 m. L Øaccuracy of _____/100 m. L Ø Pipette What will accuracy of solution be if you use pipette, volumetric flask, and electronic 1% balance? ____ What controls the Pipette accuracy? _______ 0. 6% for 100 -1000 µL Øaccuracy of ± _____ 0. 8% for 10 -100 µl Øaccuracy of ± _____

Digital Pipettes Ø Air displacement Ø Do not directly contact fluid volume Øavoids contamination

Digital Pipettes Ø Air displacement Ø Do not directly contact fluid volume Øavoids contamination of pipette Øavoids sample carryover Ø Require air tight connection between tip and body

Pipette Workings piston cylinder Pipette tip

Pipette Workings piston cylinder Pipette tip

Pipettes: Sources of Error Ø Jetting Ø Incorrect transfer technique (getting too much sample)

Pipettes: Sources of Error Ø Jetting Ø Incorrect transfer technique (getting too much sample) Wipe tip on container to remove droplets Ø Contamination from previous samples Ø Viscous fluids Ø Hot or cold fluids Ø Fluids with high vapor pressure Keep Pipette vertical!

Preparation of Solutions Ø Example: Prepare 100 m. L of a 30 m. M

Preparation of Solutions Ø Example: Prepare 100 m. L of a 30 m. M solution of methylene blue. Ø The molecular weight of methylene blue (C 16 H 18 N 3 SCl) is 319. 87 g. = concentration conversion volume mass

Preparation of Dilutions Ø Prepare 100 m. L of a 300 µM solution from

Preparation of Dilutions Ø Prepare 100 m. L of a 300 µM solution from the 30 m. M solution mass Ø Conservation of _____ = 1 m. L

Preparation of Solutions Ø Fill volumetric flask half way with distilled water Ø Add

Preparation of Solutions Ø Fill volumetric flask half way with distilled water Ø Add reagent (could be solid or liquid) Ø Mix Ø Fill volumetric flask to the line Ø Mix Ø Verify that volume didn’t change (if necessary refill to line)

UV-Visible Spectrophotometer Ø Theory Ø Instrument Ø Sample requirements Ø Software

UV-Visible Spectrophotometer Ø Theory Ø Instrument Ø Sample requirements Ø Software

Light Attenuation by an Aqueous Solution P 0 P P is light intensity (photons/s)

Light Attenuation by an Aqueous Solution P 0 P P is light intensity (photons/s)

Theory: Light Attenuation = f(? ) Ø For a given excitation process, a molecule

Theory: Light Attenuation = f(? ) Ø For a given excitation process, a molecule absorbs only one discrete amount of energy: expect very narrow absorption lines. Ø Different vibrational and rotational states yield _______ broad absorption lines. Ø Exponential decay with distance

A= bc Ø Po - _____ incident light intensity Ø P light intensity after

A= bc Ø Po - _____ incident light intensity Ø P light intensity after passing through sample path length (1 cm) Ø b - _______ Ø c - _______ concentration Ø - ______ coefficient (function of extinction wavelength and molecule)

Absorption Spectra Ø Absorption Spectra for Methylene Blue Ø Broad peaks blue Ø Absorbs

Absorption Spectra Ø Absorption Spectra for Methylene Blue Ø Broad peaks blue Ø Absorbs _____, red looks ______

Instrument Light Path

Instrument Light Path

Absorbance Measurement Limitations lamp Ø Po is a function of the _____. Ø If

Absorbance Measurement Limitations lamp Ø Po is a function of the _____. Ø If absorbance is high what is P? ______ small Ø Suppose A = 3, what is Po/P? _____ 1000 Ø Suppose I create samples of higher and higher concentration. What will happen to the absorbance measurements? minimum (non zero) P that There is a _____ can be measured by an instrument. A _______ doesn’t keep increasing! Amax 3

Sample Requirements Ø Sipper cell Øperistaltic pump draws sample into sipper cell Ørequires a

Sample Requirements Ø Sipper cell Øperistaltic pump draws sample into sipper cell Ørequires a few m. L to displace previous cell contents sample Light source pump detector

Software Ø Reference (single sample) Ø subtracts absorbance of sample cell and reference solution

Software Ø Reference (single sample) Ø subtracts absorbance of sample cell and reference solution Ø usually distilled water or reagent blank Ø Standards (multiple samples) calibration Ø used to create a _____ curve Ø Samples (multiple samples) Ø after sampling, standards can be used to estimate the concentration of samples

Maximum Absorbance: P 0 is measured as reference! Ø Max absorbance f() lamp intensity

Maximum Absorbance: P 0 is measured as reference! Ø Max absorbance f() lamp intensity Ø ________ detector sensitivity Ø ________ cell absorbance Ø ________ reference absorbance Ø ________ acceptable error Ø ________ Ø absorbance readings that exceed this value will not be used in analysis

Standards your name general description rinse time sample concentrations select number of samples by

Standards your name general description rinse time sample concentrations select number of samples by moving this control

Samples enter sample descriptions here select number of samples by moving this control

Samples enter sample descriptions here select number of samples by moving this control