Table 1. Wavelengths of lightFigure 2. Density of water vs. temperature.Laboratory Measurements and ProceduresIntroductionTheoryExperimental ObjectivesExperimental MethodsMass MeasurementsTemperature MeasurementPipette TechniqueMeasure DensityPrepare methylene blue standards of several concentrationsPrepare a standard curve and measure an unknownPrelab QuestionsQuestionsData SheetBalance CalibrationTemperature MeasurementPipette Technique (use DI‑100 balance)Measure Density (use DI‑800 balance)Prepare methylene blue standards of several concentrationsMeasure absorbance at 660 nm using a spectrophotometer.Lab Prep NotesTable 2. Reagent list.Table 3. Equipment listTable 4. Methylene Blue Stock SolutionSetupLaboratory Measurements and ProceduresIntroductionMeasurements of masses, volumes, and preparation of chemical solutions of knowncomposition are essential laboratory skills. The goal of this exercise is to gainfamiliarity with these laboratory procedures. You will use these skills repeatedlythroughout the semester.TheoryMany laboratory procedures require preparation of chemical solutions. Mostchemical solutions are prepared on the basis of mass of solute per volume of solution(grams per liter or Moles per liter). Preparation of these chemical solutions requiresthe ability to accurately measure both mass and volume. Preparation of dilutions is also frequently required. Many analytical techniquesrequire the preparation of known standards. Standards are generally prepared withconcentrations similar to that of the samples being analyzed. In environmental workmany of the analyses are for hazardous substances at very low concentrations (mg/Lor µg/L levels). It is difficult to weigh accurately a few milligrams of a chemical withan analytical balance. Often dry chemicals are in crystalline or granular form witheach crystal weighing several milligrams making it difficult to get close to the desiredweight. Thus it is often easier to prepare a low concentration standard by diluting ahigher concentration stock solution. For example, 100 mL of a 10 mg/L solution ofNaCl could be obtained by first preparing a 1 g/L NaCl solution (100 mg in 100 mL).One mL of the 1 g/L stock solution would then be diluted to 100 mL to obtain a 10mg/L solution.Absorption spectroscopy is one analytical technique that can be used to measurethe concentration of a compound. Solutions that are colored absorb light in the visiblerange. The resulting color of the solution is from the light that is transmitted.According to Beer's law the attenuation of light in a chemical solution is related to theconcentration and the length of the path that the light passes through.logoPbcPe� �=� �� �where c is the concentration of the chemical species, b is the distance the light travelsthrough the solution,  is a constant Po is the intensity of the incident light, and P isthe intensity of the transmitted light. Absorption, A, is defined as:logoPAP� �=� �� �In practice Po is the intensity of light through a reference sample (such as deionizedwater) and thus accounts for any losses in the walls of the sample chamber. Fromequation and it may be seen that absorption is directly proportional to theconcentration of the chemical species.A bce=The instrument you will use to measure absorbance is a Hewlett Packard (HP)model 8452A diode array spectrophotometer. The diode array spectrophotometer usesa broad-spectrum source of incident light from a deuterium lamp. The light passesthrough the sample and is split by a grating into a spectrum of light that is measuredby an array of diodes. Each diode measures a bandwidth of 2 nm with 316 diodescovering the range from 190 nm to 820 nm. The wavelengths of light and their colorsare given in Table 1. The light path for the diode array spectrophotometer is shown inFigure 1.Figure 1. Diagram of light path in diode array spectrophotometer.The HP 8452Aspectrophotometer has a photometric range of 0.002 - 3.3 absorbance units. Inpractice absorbance measurements greater than 2.5 are not very meaningful as theyindicate that 99.7% of the incident light at that wavelength was absorbed. Conversely,an absorbance of 0.002 means that 0.5% of the incident light at that wavelength wasabsorbed.When measuring samples of known concentration the Spectrophotometer software(page Error: Reference source not found) calculates the relationship betweenabsorbance and concentration at a selected wavelength. The slope (m), intercept (b)and correlation coefficient (r) are calculated using equation through .The slope of the best fit line is( )22x ynxymxxn� �-=-���The intercept of the line isb my x= -The correlation coefficient is defined asTable 1. Wavelengths of lightcolor wavelength (nm)ultra violet 190-380violet 380-450blue 450-490green 490-560yellow 560-590orange 590-630red 630-760Diode ArraySpectrograph LensShutterGratingSlitSample CellSource LensDeuterium Lamp( ) ( )2 22 2x ynxyrx yx yn n� �-=� �� �� �� �- -� �� �� �� ��� �� �where x is the concentration of the solute (methylene blue in this exercise), y is theabsorbance, and n is the number of samples. Experimental ObjectivesTo gain proficiency in:1) Calibrating and using electronic balances2) Digital pipetting3) Preparing a solution of known concentration4) Preparing dilutions5) Measuring concentrations using a UV-Vis spectrophotometerExperimental MethodsMass MeasurementsMass can be accurately measured with an electronic analytical balance. Perhapsbecause balances are so easy to use it is easy to forget that they should be calibratedon a regular basis. It is recommended that balances be calibrated once a week, afterthe balance has been moved, or if excessive temperature variations have occurred. Inorder for balances to operate correctly they also need to be level. Most balances comewith a bubble level and adjustable feet. Before calibrating a balance verify that thebalance is level. The environmental laboratory is equipped with balances manufactured by DenverInstruments. To calibrate the Denver Instrument balances:1) Zero the balance by pressing the tare button.2) Press the MENU key until "MENU #1" is displayed.3) Press the 1 key to select Calibrate.4) Note the preset calibration masses that can be used for calibration on the bottomof