Spring 2024 CHEM 120-Spectrophotometry
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Spring 2024 CHEM 120
Spectrophotometry: Pigments, light, and concentration Discussion Questions
Make sure that your responses to these discussion questions is legible, if your responses are illegible they will
not receive credit. We suggest that you type your responses. Using the data that you collected and wrote in your
lab notebook during lab, organize your data into the required tables or graphs. Make sure that you format the
table properly, i.e. include a title and labels for your columns, etc. Also make sure your plots are properly
formatted and clearly labeled.
1.
In a single table, tabulate your calibration curve data points. Make sure to include all proper labels and units for your solutions, concentrations, and Absorbance values. Also include your λ
max
value (2 points).
Solution
Concentration (M)
Absorbance at λ
max 668.5 nm
Stock
0.0003000
0.337
50%
0.0001500
0.183
25%
0.0000750
0.095
12.5%
0.0000375
0.042
Table 1 Chlorophyll solutions with corresponding concentrations and measured absorbance at 668.5 nm
2.
Attach plot for your calibration curve. Please format your graph including a title, axis-labels and units. Include your best fit line displaying the appropriate slope, y-intercept and R
2
value. (2 points).
Figure 1 Graph of concentrations of chlorophyll solutions and measured absorbances at 668.5 nm
3.
While doing the calibration curve plot for Spectrophotometry experiment, Zack and Mario had a conceptual discussion about how they should plot their data. Zach thinks that they should force the trendline for the absorbance vs concentration through the origin. On the other hand, Mario thinks (0,0) should be added. Is Zack or Mario correct? Using your data, explain why. Is adding (0,0) equivalent to forcing the trendline through the origine? Please provide your answer in paragraph form (6 points)
Zach is incorrect. Arbitrarily forcing the trendline through the origin as Zach suggests completely
changes the slope and the y-intercept of the trendline. This equation is therefore inaccurate and
gives an inaccurate value for absorbance. In contrast, Mario is correct. Adding the point (0,0) is valid when the y-intercept could feasibly be this point. In this case, we measured the absorbance of a blank cuvette, which has a concentration and absorbance of 0. Since the data point exists, we could feasibly add this point without qualitatively affecting the data.
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Related Questions
The spectroscopic data in the table is generated with five solutions of known concentration.
Concentration (M)
0.0133
m=
0.0266
0.0532
0.106
0.213
Absorbance
0.1271
What is the intercept of the linear regression line?
0.08531
0.5388
1.069
Use a spreadsheet program, such as Microsoft Excel, to graph the data points and determine the equation of the best-fit line.
1.954
What is the slope of the linear regression line formed by these points?
M-1
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Pre-Lab Questions- To be completed BEFORE lab as an entry slip. These questions are based on
content presented in the lab procedure and in lecture.
1. Consider the electromagnetic spectrum. (Reference Site)
-- Increasing Frequency (v)
10² 10%
10
10² 10%
Trays
1044
10
10-
10
UV
30*
10
IR
Visible spectrum
10²
10*
30
Microwave FM AM
Radio waves
10° 10²
10 H 10²
Increasing Wavelength (2)→
10
Increasing Wavelength in m
a. What type of radiation has the longest wavelength?
10 10² 10 voto
b. What type of radiation has the highest energy?
Long radio waves
c. Looking at the emission line spectrum for hydrogen in Figure 1. Which colored line
corresponds to the emission of the highest-energy photon?
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We are stuck on 9. We get how Q is 16 and rxn goes left.
at eq
2+ x. 2 + x. -2 x
We get x= 1. Therefore
k = 4. Please help us.
Questin 9. See conditions from 8
This is not a graded question as it is a practice question . I am 60 years old and helping my son prepare for the AP exam in a few months. We do questions at the back of the textbook by Zumdahl and Zumdahl
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The students conducted the assay for LDH activity using the two serum samples. Each cuvette (path length 1 cm) contained 3ml of a suitable assay buffer (including pyruvate as substrate). 20 microlitres of either serum was added to the cuvette and the absorbance values immediately recorded at the optimum wavelength for a period of 5 minutes (absorbance readings taken every 30 seconds).
Protein concentration of serum sample
(mg/ml)
Change in absorbance at optimum wavelength per minute
Control serum (C)
8
-0.04
Diseased serum (D)
7.8
-0.6
1c. Using the molar absorption coefficient of NADH as 6220 M-1 cm-1, and by application of the Beer-Lambert law, estimate the enzyme activity in the two samples (C and D). Express activity as moles per second.
1d. Estimate the specific activity of the two samples (moles per second per microgram).
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To determine the amount of copper and the identity of the copper compound in the unknown sample, a 100-mL solution containing 120.8 mg of the sample was initially prepared. Then, 5.00 mL of this solution was pipetted and diluted with deionized water in another 100-mL volumetric flask. The absorbance of this diluted solution was determined three times at the analytical wavelength. The data is in the table below.
Unknown sample
Trial 1
Trial 2
Trial 3
Absorbance (AU)
0.378
0.379
0.377
What is the corrected absorbance?
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10. The concentration of haemoglobin in a blood sample was determined by
spectrophotometry. A standard curve of the absorbance at 412 nm of several solutions
of known haemoglobin concentrations was created. The data for the standard curve
is shown below.
a. Calculate the linearity coefficient (r), y-intercept and slope.
b. What is the concentration (in ug/mL) of haemoglobin in your sample if the
absorbance obtained at 412 nm was 0.303?
Absorbance Concentration of standard solution (pg/ml)
(412nm)
0.069
1
0.113
2
0.201
4
0.377
8.
0.730
16
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To determine the amount of copper and the identity of the copper compound in the unknown sample, a 100-mL solution containing 120.8 mg of the sample was initially prepared. Then, 5.00 mL of this solution was pipetted and diluted with deionized water in another 100-mL volumetric flask. The absorbance of this diluted solution was determined three times at the analytical wavelength. The data is in the table below.
Unknown sample
Trial 1
Trial 2
Trial 3
Absorbance (AU)
0.388
0.389
0.388
What is the Corrected Absorbance of each trial? How do you get the corrected absorbance?
arrow_forward
To determine the amount of copper and the identity of the copper compound in the unknown sample, a 100-mL solution containing 120.8 mg of the sample was initially prepared. Then, 5.00 mL of this solution was pipetted and diluted with deionized water in another 100-mL volumetric flask. The absorbance of this diluted solution was determined three times at the analytical wavelength. The data is in the table below.
Unknown sample
Trial 1
Trial 2
Trial 3
Absorbance (AU)
0.375
0.374
0.376
What is the concentrartion of Cu in the analyzed solution?
arrow_forward
To determine the amount of copper and the identity of the copper compound in the unknown sample, a 100-mL solution containing 120.8 mg of the sample was initially prepared. Then, 5.00 mL of this solution was pipetted and diluted with deionized water in another 100-mL volumetric flask. The absorbance of this diluted solution was determined three times at the analytical wavelength. The data is in the table below.
Unknown sample
Trial 1
Trial 2
Trial 3
Absorbance (AU)
0.388
0.389
0.388
What is the concentration of Cu in analyzed solution (mg/L) for each trial?
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A student weighed out 0.150 g of protein powder and dissolved it in 100 mL of water (Solution 1). The student then diluted this solution by transferring 1 mL into a 25 mL flask and diluting with water (Solution 2). Finally, 1 mL of that solution was transferred to a test tube and combined with 4 mL Bradford reagent. The absorbance of the solution in the test tube was 0.187.
Assuming that the best fit linear line of the standard curve was y = 0.04144 x + 0.01521 (μ g mL), calculate the percent protein by mass in the original protein powder.
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O Homework in Chem101 - due Su X
My Questions | bartleby
( Periodic Table – Royal Society of
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A learn.maricopa.edu/courses/1132256/modules/items/19018052
CHM151 17003 > Modules > Weeks 8 & 9 - Chapter 7 > Homework in Chem101 - due Sunday night
CG 2020 FALL CRED
Question 37 of 4
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Complete the balanced neutralization equation for the reaction below:
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Here is the protocol for a UV-Vis spectrophotometer to detect water and chlorine-carbon.
1.Dissolve the water and chlorine-carbon compounds in a solvent, such as water.
2.Prepare a standard solution of known concentration that is similar to the sample being measured.
3.Calibrate the spectrophotometer using the standard solution.
4.Measure the absorbance of the sample using the spectrophotometer.
5.Calculate the concentration of the compounds in the sample using the calibration curve obtained from the standard solution.
How is the spectrophotometer calibrated with standard solutions? When is the blank solution placed in the spectrophotmeter?
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The data in the table are used to create a calibration curve for the determination of RNA from
its absorbance at 260 nm.
Use a spreadsheet and the least-squares method to determine the slope and y-intercept of the
best straight line fit to the calibration curve. Do not subtract the blank reading when creating
the calibration curve.
m=
RNA
(µg)
0.00
10.29
20.58
30.87
41.16
mass=
Absorbance at 260
nm
0.227
0.452
0.786
1.078
1.296
Hg-1
Using the slope and intercept of the calibration curve, determine the quantity of RNA in a sample that gives an absorbance of
0.523.
µg
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QUESTION 4
4. The main difference between "in-time" and "in-space" spectrophotometer design is that the former uses timed
mirrors to alternate measurements between reference and sample using the same detector, while in-space uses a
beamsplitter to send light through a different path to the same detector which is able to retrieve the signal using lock-
in amplification.
A. False
B. True
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The spectrophotometer is set at 600 nm. The absorbance of each cuvette is read. The data is
shown below: Cuvette 1 is the clank
Cuvette #
ml albumin
Absorbance
0.
0.2
.063
3
0.4
.163
4.
0.6
.187
0.8
.253
1.0
.289
Using this data, plot an Excel graph showing absorbance (y-axis) vs ml albumin (x-axis). Find the
correlation coefficient (r). R2 > 0.99 = excellent data; r2 0.98-0.95 good data r2 0.94 -0.90 =
fair data. How would you describe this data?-
To find the relative sensitivity, for each of tubes 2-6 (neglect blank), perform the following
operation absorbance (Lowry)/Absorbance (Biuret x 0.5 gram albumin (Biuret)/0.05 gram
(Lowry) x 200 ml (Lowry)/100 mL (Biuret). This implifies to Relative sensitivity = 20 dilution
factor x absorbance (Lowry)/Absorbance (Biuret). Report your relative sensitivity as an average
of all 5 values.
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Enter this data set in Excel, construct an appropriate plot, develop a trendline (not forcing the trendline through 0,0), and determine a linear equation. If an unknown sample exhibits an absorbance of 0.517, determine
the concentration of the compound in this sample.
mM
A
0
0
0.48 0.21076
0.96 0.4004
1.44 0.6072
1.92 0.7546
2.4 0.9086
O 1.00 mM
O 1.10 mM
O 1.20 mM
O 1.40 mM
O 1.30 mM
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Plot a professional quality standard curve (absorbance vs. protein amount in µg) with an appropriate title and caption underneath the plot from the data give above. The caption should include the best-fit data.
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Table 1. pH of water samples
Sample No.
Water Samples
pH
1
Tap water from household
6.59
2
Water from creek
7.35
3
Water from river
7.35
4
Unknown sample
7.00
Table 2. Absorbance of Samples
Table 3. Calibration Curve Data*
Sample No.
Absorbance
Absorbance
3-
Concentration of PO4
Ions, ppm
(1 pt each)
1
1.33
1.0
0.050
2
3.26
2.0
0.093
0.89
3.0
0.137
4
0.099
4.0
0.185
5.0
0.230
*Provided by the instructor
II. Graph
Determination of Phosphate Concentration
3-
1. Using Excel, plot the concentration of PO4* (x-axis) vs. absorbance (refer to Table 3) The olot
must show the equation of the best-fit (or trend) line of the graph. Attach the graph below
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Using the data below, please create a calibration graph and determine the concentration of
the unknown sample.
Standard #1
Standard #2
Standard #3
Unknown sample
Concentration
1.5 x 10 M
2.0 x 103 M
2.5 x 10 M
?
Absorbance
0.61
0.80
0.98
0.72
a) 1.6 x 10 M
b) 1.8 x 10° M
c) 2.1 x 103M
d) 2.2 x 10 M
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Table 1.0 of data showing calibration standards prepared by dilution and the absorbance
Concentration CuSO4 (M) Volume of 0.5M (ml) Volume of D/Water (ml) Absorbance
0.00
0.10
0.20
0.30
0.40
0.50
II.
0.00
1.00
2.00
3.00
4.00
5.00
5.00
4.00
3.00
2.00
1.00
0.00
1. Construct a graph of concentration against absorbance, clearly show a best fit line.
Find regression of your graph. Show how
0.000
0.406
0.638
0.854
1.202
1.276
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No need to upload any image just give me the answer for the question with proper explanation.
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If Johnny Joestar measured the amount of light absorbed by the analyte in a sample to determine its concentration, what kind of method did he use?
Volumetric method
Gravimetric method
Electroanalytical method
Spectroscopic method
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List three typical examples applications of SFC analysis.
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The concentration of hydronium ions and hydroxide ions can vary widely in a solution, easily from 1 M to 1 x 10-14 M. This variation is too wide to fit on an analog readout. In order to make this into a smaller scale suitable for an analog readout, the log to the base 10 was used so that the read out is focused on powers of 10. Note that the log [1] = 0, and the log of 1 x 10-14 = -14. It was then decided that the numbers should be positive so the sign was changed. The way to measure concentrations of both H+ and OH- was thus defined as -log. This was then referred to as p. Thus, pH = -log[H+], pOH = -log[OH-], and pK = -log[Keq]. You need to figure out how to do this mathematical manipulation on your calculator. The acid-base reaction in pure water is H2O + H2O ⇔ H3O+1 + OH-1 and it was found that in pure water under standard conditions the [H3O+1] = [OH-1] = 1 x 10-7 M Neutral pH is defined as having a [H+] or [H3O+1] of 1 x 10-7 M.
The pH of a neutral solution is thus pH = -log[1 x…
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Do not answer in image format
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Plot (using appropriate scaling, with titles, axis labels, and units where applicable) the
absorbance versus the concentration of each standard on graph paper. The slope of the best-
fit line will be , in A = ,bc (Beer’s Law, where A is absorbance, , the extinction
coefficient, b the path length, and c the concentration of the sample). Produce and submit the standard curve
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■ True or False: in a TOF analyzer, a fragment of low mass reaches the
detector first
▪ In GCMS when we know what analytes we are measuring we use
not know what analytes are in the sample we use
than scan.
▪ A) Scan; SIM
■ B)SIM; scan
• C) SIM; SIM
mode. When we do
mode. SIM allows greater sensitivity
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Table 1. Data on EDTA Standardization
Weight of CaCO3 (g): 0.2003g
Trial
Volume of EDTA (mL)
Molarity of EDTA (M)
1
8.60
2
8.50
8.55
Average Molarity of EDTA (M)
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Plot a calibration graph of absorbance vs concentration of Ca2+ and determine the concentration of Ca2+ in the unknown tap and unknown bottle water samples.
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Wavelength (nm)
a. Which part of the visible light spectrum is most useful (most absorbed) by
chlorophyll b?
b. Which pigment do you think was best at absorbing the red light that was
shown on the leaves when using a red-light filter in the experiment?
Explain.
c. Do you think a greenhouse with green and yellow lights would be very
successful in terms of plant growth? Explain.
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Page of 5 O
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ZOOM
Question 1: Using a well-known protein estimation technique, absorbance reading for a set of
standards were obtained. You were required to plot the data. Which graph would you use? Plot
the data using the graph type.
Protein
Absorbance
concentration
(mg/mL)
(a.u.)
0.01
0.05
0.12
5
10
25
0.25
0.63
1.25
50
Question 2: Enzymes are biological catalysts and perform important biochemical reactions in the
body. A group of scientists discovered a new enzyme – Enzyme X. To understand the working of
this enzyme, they performed biochemical assays to assess the activity of the enzyme. The
results for which are shown in the table below. Based on the information provided in the table,
please state which graph would you use? Plot the data using the graph type.
Enzyme activity
(%)
35
55
pH
3
4
5
68
6
79
7
98
85
20
8
9
10
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Write the corresponding letters to the following descriptions. Each proposal can be associated with 0 or
1 match and each letter is not necessarily associated with a proposal.
a. [Ar] 4s?
b. 1s 2s2p63s23p3
c. 1s2s2p2
d. [Ne] 3s23ps
e. [Ar] 4s23d10
f. 1s22s!
g. None of those answers
1. I am a halogen
2. I am a transition metal
3. I am alkaline
4. I have exactly 2 single electrons
5. I am the smallest of the atoms presented here
6. I have exactly 3 valence electrons
7.I am the most paramagnetic of the atoms presented here
8. I am the copper
9.I am isoelectronic with Ti2+
IF
cessibility: Good to go
IA
16
SIS
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Related Questions
- The spectroscopic data in the table is generated with five solutions of known concentration. Concentration (M) 0.0133 m= 0.0266 0.0532 0.106 0.213 Absorbance 0.1271 What is the intercept of the linear regression line? 0.08531 0.5388 1.069 Use a spreadsheet program, such as Microsoft Excel, to graph the data points and determine the equation of the best-fit line. 1.954 What is the slope of the linear regression line formed by these points? M-1arrow_forwardPre-Lab Questions- To be completed BEFORE lab as an entry slip. These questions are based on content presented in the lab procedure and in lecture. 1. Consider the electromagnetic spectrum. (Reference Site) -- Increasing Frequency (v) 10² 10% 10 10² 10% Trays 1044 10 10- 10 UV 30* 10 IR Visible spectrum 10² 10* 30 Microwave FM AM Radio waves 10° 10² 10 H 10² Increasing Wavelength (2)→ 10 Increasing Wavelength in m a. What type of radiation has the longest wavelength? 10 10² 10 voto b. What type of radiation has the highest energy? Long radio waves c. Looking at the emission line spectrum for hydrogen in Figure 1. Which colored line corresponds to the emission of the highest-energy photon?arrow_forwardWe are stuck on 9. We get how Q is 16 and rxn goes left. at eq 2+ x. 2 + x. -2 x We get x= 1. Therefore k = 4. Please help us. Questin 9. See conditions from 8 This is not a graded question as it is a practice question . I am 60 years old and helping my son prepare for the AP exam in a few months. We do questions at the back of the textbook by Zumdahl and Zumdahlarrow_forward
- The students conducted the assay for LDH activity using the two serum samples. Each cuvette (path length 1 cm) contained 3ml of a suitable assay buffer (including pyruvate as substrate). 20 microlitres of either serum was added to the cuvette and the absorbance values immediately recorded at the optimum wavelength for a period of 5 minutes (absorbance readings taken every 30 seconds). Protein concentration of serum sample (mg/ml) Change in absorbance at optimum wavelength per minute Control serum (C) 8 -0.04 Diseased serum (D) 7.8 -0.6 1c. Using the molar absorption coefficient of NADH as 6220 M-1 cm-1, and by application of the Beer-Lambert law, estimate the enzyme activity in the two samples (C and D). Express activity as moles per second. 1d. Estimate the specific activity of the two samples (moles per second per microgram).arrow_forwardTo determine the amount of copper and the identity of the copper compound in the unknown sample, a 100-mL solution containing 120.8 mg of the sample was initially prepared. Then, 5.00 mL of this solution was pipetted and diluted with deionized water in another 100-mL volumetric flask. The absorbance of this diluted solution was determined three times at the analytical wavelength. The data is in the table below. Unknown sample Trial 1 Trial 2 Trial 3 Absorbance (AU) 0.378 0.379 0.377 What is the corrected absorbance?arrow_forward10. The concentration of haemoglobin in a blood sample was determined by spectrophotometry. A standard curve of the absorbance at 412 nm of several solutions of known haemoglobin concentrations was created. The data for the standard curve is shown below. a. Calculate the linearity coefficient (r), y-intercept and slope. b. What is the concentration (in ug/mL) of haemoglobin in your sample if the absorbance obtained at 412 nm was 0.303? Absorbance Concentration of standard solution (pg/ml) (412nm) 0.069 1 0.113 2 0.201 4 0.377 8. 0.730 16arrow_forward
- To determine the amount of copper and the identity of the copper compound in the unknown sample, a 100-mL solution containing 120.8 mg of the sample was initially prepared. Then, 5.00 mL of this solution was pipetted and diluted with deionized water in another 100-mL volumetric flask. The absorbance of this diluted solution was determined three times at the analytical wavelength. The data is in the table below. Unknown sample Trial 1 Trial 2 Trial 3 Absorbance (AU) 0.388 0.389 0.388 What is the Corrected Absorbance of each trial? How do you get the corrected absorbance?arrow_forwardTo determine the amount of copper and the identity of the copper compound in the unknown sample, a 100-mL solution containing 120.8 mg of the sample was initially prepared. Then, 5.00 mL of this solution was pipetted and diluted with deionized water in another 100-mL volumetric flask. The absorbance of this diluted solution was determined three times at the analytical wavelength. The data is in the table below. Unknown sample Trial 1 Trial 2 Trial 3 Absorbance (AU) 0.375 0.374 0.376 What is the concentrartion of Cu in the analyzed solution?arrow_forwardTo determine the amount of copper and the identity of the copper compound in the unknown sample, a 100-mL solution containing 120.8 mg of the sample was initially prepared. Then, 5.00 mL of this solution was pipetted and diluted with deionized water in another 100-mL volumetric flask. The absorbance of this diluted solution was determined three times at the analytical wavelength. The data is in the table below. Unknown sample Trial 1 Trial 2 Trial 3 Absorbance (AU) 0.388 0.389 0.388 What is the concentration of Cu in analyzed solution (mg/L) for each trial?arrow_forward
- A student weighed out 0.150 g of protein powder and dissolved it in 100 mL of water (Solution 1). The student then diluted this solution by transferring 1 mL into a 25 mL flask and diluting with water (Solution 2). Finally, 1 mL of that solution was transferred to a test tube and combined with 4 mL Bradford reagent. The absorbance of the solution in the test tube was 0.187. Assuming that the best fit linear line of the standard curve was y = 0.04144 x + 0.01521 (μ g mL), calculate the percent protein by mass in the original protein powder.arrow_forwardO Homework in Chem101 - due Su X My Questions | bartleby ( Periodic Table – Royal Society of how to take a screenshot on an + A learn.maricopa.edu/courses/1132256/modules/items/19018052 CHM151 17003 > Modules > Weeks 8 & 9 - Chapter 7 > Homework in Chem101 - due Sunday night CG 2020 FALL CRED Question 37 of 4 Submit Account Home Complete the balanced neutralization equation for the reaction below: Announcements Dashboa |Modules HCIO-(aq) + CSОН(ag) — rd Concourse Syllabus Courses Grades D2+ N3+ D4+ Cisco Webex Groups 1 2 3 4 7 8 9. Tutoring/Learning Center Calendar Os Do Inbox (s) (1) (g) (aq) History Cs H CI Help • Previous Next » Library 1:03 PM O Type here to search 10/18/2020 +arrow_forwardHere is the protocol for a UV-Vis spectrophotometer to detect water and chlorine-carbon. 1.Dissolve the water and chlorine-carbon compounds in a solvent, such as water. 2.Prepare a standard solution of known concentration that is similar to the sample being measured. 3.Calibrate the spectrophotometer using the standard solution. 4.Measure the absorbance of the sample using the spectrophotometer. 5.Calculate the concentration of the compounds in the sample using the calibration curve obtained from the standard solution. How is the spectrophotometer calibrated with standard solutions? When is the blank solution placed in the spectrophotmeter?arrow_forward
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- Organic Chemistry: A Guided InquiryChemistryISBN:9780618974122Author:Andrei StraumanisPublisher:Cengage Learning
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ISBN:9780618974122
Author:Andrei Straumanis
Publisher:Cengage Learning