Analytical Chemistry

a measurement science
consist of a set of powerful ideas and methods
The Nature of Analytical useful in all fields of science and medicine
Chemistry On July 4, 1997, the Mars Pathfinder
spacecraft delivered the Sojourner rover
by on Ares Vallis, Mars in its quest for
information on the nature of the red planet.
Rey Y. Capangpangan, Ph.D.
Chem 20 : Analytical Chemistry (Lec
(Lec))
Chemistry Division
College of Arts and Sciences
Caraga State University

Mars: Through the eyes of the Sojourner rover 2

APXS Instrumentation aboard the Sojourner Chemical Analysis

Key experiment aboard the

Sojourner used the Alpha
Qualitative analysis
proton X-ray spectrometer reveals the chemical identity of the species
APXS combines 3 advanced (elements and compounds) in a sample
instrumental techniques:
Rutherford backscattering
spectroscopy
Proton emission
spectroscopy Quantitative analysis
X-ray fluorescence determines the relative amounts
of each substance in a sample

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1
 Analyte
the components of a sample
to be determined
(or the constituent of interest)
5 6
Establishing Economic Value
Determining the amount of:
silver in coin
oil in shale
protein in animal feedstock
7 8
Role of Analytical Chemistry
Establishing economic value
Determining health hazards
Diagnosing diseases
Controlling quality
Complying to environmental standards
Relating properties to composition or structure
Conducting research
Determining Health Hazard
Establishing the concentration
(amount) of:
sulfur dioxide in emissions from a
coal-burning power plant
pesticide residue on fresh fruit and
vegetables
2
 Disease Diagnosis Quality Control

In drug and processed-food

Clinical determinations of:
manufacturing*, analysis of:
Glucose
raw materials
Urea
additives
Concentrations of O2 and CO2
finished products
ionized calcium in blood
serum (parathyroid disease)
*to achieve a constant, pre-determined quality of product,
and to ensure product meets acceptable standards

9 10

Quality Control Quality Control

Monitoring of household gas supply for:

Analysis of steel* during its mercaptan content
production to adjust concentrations *to ensure that the gas has a sufficiently noxious odor to warn of
of: dangerous leaks

carbon Farmers tailor fertilization and irrigation

nickel schedules* from quantitative analysis
chromium of:
*to achieve desired strength, hardness, corrosion resistance, and ductility plants *to meet changing plant needs during the growing
season
soil
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3
 Complying to Environmental Standards Relating properties to composition or structure

physical and chemical properties of

Measuring automobile exhaust gases*:
alloys depend on their chemical
hydrocarbons
composition
nitrogen oxides
activity or effectiveness of
carbon monoxide
*to assess effectiveness of smog control devices
pharmaceuticals, pesticides, and
herbicides depend largely on their
chemical structure

13 14

Conducting Research Conducting Research

Physiologists study the role of K, Ca, and Na ions in the

following the accumulation of pesticides in the food
chain
tracking the metabolic fate of drugs in the human body
determining the nature of catalytic surfaces to coal to
natural gas
looking for new ways to separate, identify, and
determine the concentration of enzymes and proteins
developing new and better instruments for quantitative
measurements
body fluids of animals in nerve signal conduction,
muscle contraction, and relaxation
Chemists unravel the mechanisms of chemical reactions
by studying the rate of consumption of reactants or
formation products in a chemical reaction
Materials scientists study the impurities in crystalline
germanium and silicon in semiconductor devices
Archaeologists identify the source of volcanic glasses
(obsidian) to trace prehistoric trade routes for tools and
weapons fashioned from obsidian

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4
The Central Role of Analytical Chemistry Quantitative Analytical Methods

Chemical analysis is a
vital tool in medical,
Complete (or Ultimate) analysis
industrial, the amounts of all of the constituents
government, and in the sample are determined
academic
laboratories
Partial analysis
the amounts of only some of the constituents
in the sample are determined

Elemental and molecular analysis

the determination of the amounts of
elements and molecules
17 18

Quantitative Analytical Methods Quantitative Analytical Methods

Classification based on type/nature of the final

Major-constituent (macro) analysis measurement
the constituent determined is present 1. Measurement of mass or volume of sample being analyzed
in high concentration
2. Measurement of some quantity that is proportional to the
amount of analyte in the sample
mass
volume
Trace analysis intensity of light
the constituent determined is present
electrical charge
in low concentration

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5
 Quantitative Analytical Methods Miscellaneous Analytical Methods
1. Gravimetric methods: determine the mass of the Measurement of such quantities as:
analyte or some compound chemically related to it
1. mass-to-charge of molecules (mass spectrometry)
2. Volumetric methods: the volume of solution
containing sufficient reagent to react completely 2. rate of radioactive decay
with the analyte is measured
3. heat of reaction
3. Electroanalytical methods: involve the measurement
of electrical properties, such as potential, current, 4. rate of reaction
resistance, and quantity of electrical charge
5. thermal conductivity
4. Spectroscopic methods: measurement of the
interaction between electromagnetic radiation and 6. optical activity
analyte atoms or molecules or on the production of
such radiation by analytes 7. refractive index
21 22

Types of Samples and Methods Classification of Analysis by Sample Size

Sample size Constituent types

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6
Classification of Analysis by Constituent Type Steps in Quantitative Analysis

What questions are asked to determine the

information needed to answer questions?

What part of that information can be

supplied by analysis?

What methods should be used for the

analyses?

Overall Plan or Goal

25 26

Steps in Quantitative Analysis Steps in Quantitative Analysis

What is the nature of the sample?

How many components are there in the Laboratory sample must
sample? accurately represent the
What is the expected concentration composition of the bulk sample
level of the analyte? Laboratory sample weighs <1 g
What is the level of accuracy required?
Analysis results of sample are used to
How samples should be analyzed? characterize the entire material from
What is the cost and time required? which the sample was taken

What equipment and technical

expertise are available?

1. Choosing a Method 2. Collecting the sample

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7
 Sampling Sampling homogeneous vs heterogeneous materials

the process of collecting a small mass Homogeneous material Heterogeneous material

of a material whose composition easy more difficult
accurately represents the bulk composition is uniform requires carefully
of the material being sampled prescribed procedures
select only the desired that are designed to
size of the sample for the ensure proper
Reliability of final results laboratory representation in the
reliability of the sampling step final sample

29 30

Acceptable Methods for Sampling Bulk Materials Sampling Bulk Materials

1. ASTM: The American Society for Testing and 1. Obtaining a gross sample
Materials
2. Reducing the gross sample to a
2. NBS: The National Bureau of Standards
transportable size
3. AOAC: The Association of Official Analytical Chemists
3. Obtaining a laboratory sample
4. AOCS: The American Oil Chemists Society

31 32

8
 The gross sample is obtained by random selection Cone and Quarter Method

1. Obtaining a gross sample

A chute may be designed that siphons off a certain

fraction of the material (~1 ton) which , taken as a
whole must be representative of the entire load of ore 1. The gross sample is piled into a cone which is
flattened and quartered
2. Reduce gross sample to a size suitable for transportation

Crushing sample to get uniform size followed by 2. Opposite quarters are retained, mixed, and the
sampling procedure that removes a fixed portion of procedure repeated until an appropriate sample is
the gross sample obtained (5-25 lbs)
Cone and quarter method

Example: Unloading ore from a ship

33 34

Obtaining a Laboratory Sample Steps in Quantitative Analysis

In the laboratory a similar procedure is used to further reduce

the sample size that can be analyzed
1. 3 or more samples are prepared and analyzed simultaneously
more confidence can be placed in a number representing the Preparing a laboratory sample
results of 3 measurements than in one representing 1
measurement Defining replicate samples
in case a sample is ruined during the analysis, the analysis can be
continued with the remaining 2 samples Preparing solutions
2. Prevent contamination during the sample collection process
(especially when the constituents to be determined are present in very
low concentration trace analysis)
All sample-handling equipment must be cleaned before use
Special care is required in handling corrosive substances (they
often react with their containers and the equipment used to handle
them) 3. Processing the sample
35 36

9
 Preparing Solid Sample Preparing Solid Sample

1. Grinding to decrease particle size It is a good idea to remove occasional water

2. Mixing to ensure homogeneity before weighing samples for analysis:

3. Storing for various lengths of time before analysis* hydrated

4. Drying samples before starting an analysis adsorbed on the surface
(alternatively, the moisture content of the sample can be
determined at the time of the analysis in a separate *occasional water is water that can be removed from the sample without
analytical procedure) causing structural changes

Occasional water in solids

*any loss or gain of water changes the chemical composition of the solid

37 38

Water content of solid samples Drying of Samples before Weighing

Water content of most samples is not constant, 1. Heat* the sample in an oven to volatilize the
varying with changes in: water
atmospheric humidity 2. After a sufficient time (~1 to 2 hours), the
sample is removed and allowed to cool in a dry
temperature atmosphere
particle size
*the best temperature for volatizing the water depends on how firmly the
Sample-handling procedures water is attached to the water

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10

Analysis on a wet basis
When samples cannot be dried because they
decompose at the temperatures necessary to
drive off the water:
analyze the sample on a wet or as received gas), the sample container must be kept in a second
basis
the water content can be determined by
analyzing another portion of the same sample
41
Defining Replicate Sample
Replicate samples, or replicates
portions of a material of approximately the same size
that are carried through an analytical procedure at the
same time and in the same way
43
Preparing Liquid Sample
Liquid samples present a related set of problems during
the preparation step:
if such samples are allowed to stand in open
containers, the solvent may evaporate and change
the concentration of the analyte
if the sample is a gas dissolved in a liquid (e.g. blood
sealed container to prevent contamination by
atmospheric gases
Extraordinary measures, including sample manipulation
and measurement in an inert atmosphere may be required
to preserve the integrity of the sample
42
Defining Replicate Sample
Most chemical analyses are performed on
replicate samples whose masses or volume
have been determined by careful
measurements with an analytical balance or a
precise volumetric device
Replication improves the quality of the results
and provides a measure of reliability
Quantitative measurements on replicates are
averaged, and various statistical tests are
performed on the results to establish reliability
44
11
 Preparing Solution Examples of Insoluble Materials

Most analyses are performed on solutions of

the sample made with a suitable solvent.
Silicate minerals
Ideally, the solvent should dissolve the
entire sample, including the analyte, rapidly High-molecular-weight polymers
and completely
Specimens of animal tissue
The conditions of dissolution should be
sufficiently mild to prevent loss of analyte

45 46

Conversion of the analyte into soluble form Dissolving Sample in Water

Heating with aqueous solutions of strong

acids, strong bases, oxidizing agents, reducing Samples composed entirely of soluble salts
agents or combinations of such reagents may be dissolved directly in water

Igniting of sample in air or oxygen Some salts, although soluble, dissolve slowly
at room temperature and mild heating may be
Performing a high-temperature fusion of the necessary
sample in the presence of fluxes

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12
 Dissolving Sample in Acids Dissolving Sample in Organic Solvents

In many instances, at least a portion of the sample

will not dissolve in water.
Strong acids will usually render the sample soluble:
Hydrochloric acid, HCl Samples containing organic compounds
dilute sulfuric acid, H2SO4
generally do not dissolve in water.
dilute perchloric acid, HClO4 Choice of organic solvents depend on their
compatibility with sample treatment and
hot concentrated sulfuric acid, H2SO4
measurement
Nitric acid, HNO3
Aqua regia (a mixture of HCl and HNO3)
Hydrofluoric acid, HF suited for dissolving silicate ores

49 50

Fusion Fusion

1. Mixing a finely ground sample with solid flux

Samples that fail to dissolve in aqueous or in an inert metal crucible
organic solvents must be fused dissolved in a
molten solvent (called a flux) that has acidic, 2. Heating until the flux melts, at which time it
basic, or oxidizing properties becomes very reactive and dissolves the
Potassium pyrosulfate, K2S2O7 sample
Sodium carbonate
3. Cooling the melt and dissolving in water or
Sodium peroxide dilute aqueous acid

51 52

13
 Why do molten solvents exhibit greater dissolving power
than aqueous solvents?
1. The temperature of a molten flux is several hundred
degrees higher than the temperature of a boiling
aqueous solution
Does the soluble form of the sample have a property that is
proportional to the analyte concentration that is
measureable?
Example: Determination of Mn in steel

2. Since the solvent itself is the acid, base, or oxidizing Mn is oxidized to MnO4 before the absorbance of
agent in a fusion, its concentration is 2 to 10 times the colored solution is measured
larger than it would be in an aqueous solution

3. The absence of water can greatly increase the acid or

base strength of a substance

53 54

Steps in Quantitative Analysis

Selective versus Specific Techniques

Interference (or interferents) are

species other than the analyte
that affect the final measurement
species that cause errors in analysis
by enhancing or attenuating
(making small) the quantity being
measured
Selective method: reactions that apply for a
few analytes
Specific method: reactions that work only for
one analyte

4. Eliminating Interferences
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14
 Eliminating Interferences
1. Selecting the method
2. Masking
3. Separation
57
Standard or Accepted Methods of Analysis
Methods that have been carefully tested and
evaluated under controlled conditions
59
Selecting the Method
Know the general composition of the sample
to be analyzed
Know which methods possibly can be used
Refer to standard or accepted methods of
analysis
Refer to review articles that evaluate and
summarize new methods and applications
58
Masking
The elimination of an interference from a
substance by converting it to a non-interfering
form
Example: a method for determining copper based on
measuring the amount of iodine produced when Cu2+
reacts with iodide ion, Fe3+ interferes by also
oxidizing iodide to iodine
2Fe3+ + 2I 2Fe2+ + I2
60
15
 Masking Separation

Precipitation

Electrodeposition
Ferric ion can be masked by adding fluoride
ion, which forms a stable complex ion with
Extraction
iron (III), FeF63 and prevent its reduction.
Ion exchange
FeF63 + I no reaction
Volatilization

Chromatography
61 62

Steps in Quantitative Analysis Examples

Most methods require the analyte

to be in a particular form that is
suitable for measurement
The exact property or substance
measured depends on the type of
method selected
Redox titration: the analyte must be in a
particular oxidation state before the titration
begins
Colorimetric method: a colorless analyte must
be converted to a colored product for the
measurement

5. Treating the sample and

measuring the analyte
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16
 Steps in Quantitative Analysis

The procedure used to calculate the

amount or concentration of an analyte
depends on the analysis method Significant Figures
absolute measurements where the (Review)
amount or concentration of analyte is
determined directly from the measured
value by

relative measurements where the Rey Y. Capangpangan, Ph.D.

amount or concentration is determined Chem 20 : Analytical Chemistry (Lec
(Lec))
by comparison of the measured value Chemistry Division
for the unknown with similar values College of Arts and Sciences
obtained for known amounts of analyte Caraga State University

6. Calculating the amount

65

What is significant figures? What is significant figures?

There are 2 kinds of Approximate: weight,

numbers: heightanything
Exact: the amount of MEASURED. No
money in your account. measurement is perfect.
Known with certainty. Uncertainty

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 When to use significant figures? How do I know how many significant figures?

When a measurement is recorded only
those digits that are dependable are
written down.
If you measured the width of a paper
with your ruler you might record 21.7cm.
To a mathematician 21.70, or 21.700 is the
same.
69
Rule: All digits are significant starting
with the first non-zero digit on the
left.
Exception to rule: In whole numbers
that end in zero, the zeros at the end
are not significant (unless they follow a
decimal point).
70

How do I know how many significant figures? How do I know how many significant figures?

Exercise
7 1 2nd Exception to rule: If
40 1 zeros are sandwiched
0.5 1
between non-zero digits,
0.00003 1
7 x 105 1 the zeros become
7,000,000 1 significant.

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18
 How do I know how many significant figures? How do I know how many significant figures?

Exercise
3rd Exception to rule: If 1.2 2 3401 4
2100 2 2100 2
zeros are at the end of a

56.76
4.00


4
3


2100.0
5.00


5
3

number that has a

0.0792
7,083,000,000


3
4


0.00412
8,000,050,000


3
6

decimal, the zeros are

significant.

73 74

What about calculation with significant figures? What about calculation with significant figures?

Addition/Subtraction
Rule: When adding or
2.45cm + 1.2cm = 3.65cm,
subtracting measured
Round off to = 3.7cm
numbers, the answer can have
no more places after the
7.432cm + 2cm = 9.432 round to
decimal than the LEAST of the
9cm
measured numbers.

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 What about calculation with significant figures? What about calculation with significant figures?

Multiplication/Division
Rule: When multiplying
or dividing, the result can 56.78 cm x 2.45cm = 139.111 cm2
have no more significant Round to 139cm2
figures than the least
reliable measurement. 75.8cm x 9.6cm = ?

77 78

Rounding off data

In rounding a number ending in 5, always

round it off so that the result ends with an Errors and
even number. Treatment/Evaluation of
Example: 0.635 rounds to 0.64
0.625 rounds to 0.62
Analytical Data
by

Rey Y. Capangpangan, Ph.D.

It is strongly recommended not to round Chem 20 : Analytical Chemistry (Lec
(Lec))

your data in the middle of calculation. You Chemistry Division

College of Arts and Sciences
can round it off in your final answer (after Caraga State University

the calculation is complete).

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 Types of Error Systematic (determinate) Errors

No analysis is free of error or uncertainty 1. Instrument errors

Systematic Error (determinate error)
The error is reproducible and can be discovered and corrected
Random Error (indeterminate error)
Caused by uncontrolled variables, which cannot be
defined/eliminated
Failure to calibrate, degradation of parts in the instrument,
power fluctuation, variation in temperature, etc.
Can be corrected by calibration or proper instrumentation maintenance
2. Method errors
- Errors due to no ideal physical or chemical behavior
-Completeness and speed of reaction, interfering side
reactions, sampling problems
Can be corrected with proper method development
3. Personal errors
Occur where measurements require judgement, result from
prejudice, color acuity problems
Can be minimized or eliminated with proper training and experience

81 82

Detection of Systematic Errors Random (indeterminate) Errors

No identifiable cause: Always present, cannot be

1. Analysis of standard samples eliminated; the ultimate limitation on the
determination of a quantity (sources of
2. Independent analysis : Analysis uncertainty).
using a reference method or
reference laboratory Example: reading a scale on an instrument
caused by the finite thickness of the lines on the
3. Blank determination scale; electrical noise

4. Variation in sample size : detects The accumulated effect causes replicate

constant error only measurements to fluctuate randomly around the
mean; Give rise to a normal or Gaussian curve;
Can be evaluated using statistics

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 Sources of Random (indeterminate) Errors Possible Combinations of Four Equal-Sized Uncertainties

Small undetectable uncertainties produce a

detectable random error
Example
Four small random errors combine to give an
overall error
Each error has an equal probability of occurring
and that each can cause the final result to be high
or low by a fixed amount U

85 86

Statistical treatment of Random Errors Random Errors follow a Gaussian or normal distribution

We are 95% certain that the true value falls within 2 (infinite
Statistical analysis is based on the assumption that population) if there is no systematic error.
random errors in chemical analysis follow Gaussian, or
normal, distribution
Mean,
Analytical data can follow distributions other than the
Gaussian distribution
Experiments where there is either a successful outcome or failure
produce data that follow the binomial distribution
Radioactive or photon-counting experiments produce results that
follow the Poisson distribution
Gaussian distribution is used to approximate these
distributions (the approximation becomes better in the limit
of a large number of experiments)

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 Properties of the Gaussian Curve
Population mean,
Population standard deviation,
Parameters refers to quantities
such as and that define a
population or distribution
Statistic refers to an estimate of a
parameter that is made from a
sample of data
89
How to determine Error?
Mean average or arithmetic mean.
Median arrange results in increasing or
decreasing order.
Precision S, RSD, CV
Accuracy Error, relative error
91
How to determine Error?
Accuracy closeness of measurement to its
true or accepted value.
Systematic or determinate errors affect
accuracy
Precision agreement between 2 or more
measurements of the sample made in
exactly the same way
Random or indeterminate errors affect
precision.
90
Accuracy
Absolute error (E) diff. between true and measured value.
E = xi - xt
Where xi = experimental value and xt = true value
Example:
xi = 19.78 ppm Fe and xt = 20.00 ppm Fe
E = 19.78 20.00 ppm = -0.22 ppm Fe
(-) value too low; (+) value too high
Relative error (Er) expressed as % or in ppt.
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23
 Analytical Procedures Precision

1. 2-3 replicates are performed and carried out through Standard Deviation (s) for small data set:
the entire experiment. Results may vary and must
calculate the central or best value for data set.
2. Mean arithmetic mean or average
3. Median - arrange results in increasing or decreasing Standard error of
order, the middle value of replicate data the mean
4. Rules:
For odd # values, median is the middle value Standard Deviation () of population: for finite/large set of data
For even # values, median is the average of the two
middle values

Where is the mean or average of the population

93 94

Precision Example

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24
 Example How to improve data precision

Averaging results
Improvement gained by averaging is limited
To increase precision by a factor of 10 times
requires 100 times as many measurements
Decrease s, since sm is directly proportional to
s
More precision in individual operations
Change procedure
Use more precise measurement tools

97 98

Pooling data to improve the reliability of s as a measure of Pooling data to improve the reliability of s as a measure of
precision precision

Several subsets of data can be pooled (combined) to get

a better estimate of the population standard deviation
than using only one data set
Assumption: same sources of random error in all
measurements
Samples have similar compositions
Samples have been analyzed in exactly the same way
Samples are randomly drawn from the same population and
thus have a common value of

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 To Calculate Spooled Example

spooled is the weighted average Deviations from the mean for

of the individual estimates each subset are squared
The squares of the deviations
of all subsets are then
summed and divided by the
appropriate number of
degrees of freedom
The pooled s is obtained by
taking the square root of the
resulting number
1 degree of freedom is lost
for each subset
The number of degrees of
freedom for the pooled s is
equal to the total number
of measurements minus
the number of subsets

101 102

Example

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26
 How sure are we that the true value is close to our z Values
measurement average?

The Confidence interval (CI) is an expression stating that the

true mean, , is likely to lie within a certain distance from
the measured mean.
We find a general expression for the confidence interval of the
true mean based on measuring a single value x. Thus,

Rarely do we estimate the true mean from a single

measurement, however. Instead, we use the experimental
mean x of N measurements as a better estimate of .

105 106

Example Solution

Determine the 80% and 95% confidence intervals for the mean
value (1100.3 mg/L) for month 1 in the example. Assume that in
each part, s = 19 is a good estimate of .

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27
 Finding the CI when is unknown Finding the CI when is unknown

To account for the variability of s, we use the important

statistical parameter t, which is defined in exactly the same
The confidence interval for the mean, of N
way as z, except that s is substituted for . For a single replicate measurements can be calculated from t
measurement with result x, we can define t as as shown:

For the mean of N measurements

where t is students t.
(tabulated)
109 110

Values of Student t Exercise

111 112

28
 Exercise Exercise

113 114

Paired t Test for Comparing Individual Differences Comparison of Two Standard Deviations (precision) using F-
test

Comparing two methods: Do the two

methods give the same answer within
experimental error?

Rules:
Always put the larger standard deviation in the
numerator, thus Fcalculated always 1
If Fcalculated > Ftable (95%), the difference is significant.

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29
 Example Critical values of F

117 118

Q-Test: Statistical tool for rejection of outliers Q-Test: Statistical tool for rejection of outliers

xq = questionable result Example

xn = nearest neighbour
w = the spread (largest value smallest
value)
Note: Always arrange the data according to increasing order

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 Recommendations for Treating Outliers Recommendations for Treating Outliers

1. Re-examine carefully all data relating to the outlying result to see 4. If more data cannot be secured, apply the Q test to the existing
if a gross error could have affected its value. This set to see if the doubtful result should be retained or rejected
recommendation demands a properly kept laboratory notebook on statistical grounds.
containing careful notations of all observations
2. If possible, estimate the precision that can be reasonably 5. If the Q test indicates retention. consider reporting the median
expected from the procedure to be sure that the outlying result of the set rather than the mean. The median has the great
actually is questionable. virtue of allowing inclusion of all data in a set without undue
influence from an outlying value. In addition, the median of a
3. Repeat the analysis if sufficient sample and time are available.
normally distributed set containing three measurements
Agreement between the newly acquired data and those of the
provides a better estimate of the correct value than the mean of
original set that appear to be valid will lend weight to the notion
the set after the outlying value has been discarded.
that the outlying result should be rejected. Furthermore, if
retention is still indicated, the questionable result will have a
small effect on the mean of the larger set of data.

121 122

Assignment The Least-Squares Method

Regression analysis provides the means

for objectively obtaining such a line and
also for specifying the uncertainties
associated with its subsequent use. We
consider here only the basic method of
least squares for two- dimensional data.

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 The Least-Squares Method Assumptions of the Least-Squares Method

The first is that a linear relationship actually exists

between the measured response y and the standard
Analyte concentration x. The mathematical
relationship describing this assumption is called the
Calibration curve for regression model, which may be represented as
the determination of
isooctane in a
hydrocarbon
mixture. where b is the y intercept (the value
of y when x is zero) and m is the
slope of the line

125 126

Assumptions of the Least-Squares Method Finding the Least-Squares Line

Linear least-squares analysis gives you the
equation for the best straight line among a
set of x,y data points when the x data contain
negligible uncertainty.
As illustrated in Figure 8-9, the vertical
deviation of each point from the
straight line is called a residual.
The least-squares rnethod finds the
sum of the squares of the residuals
SSresid and minimizes these according
to the minimization technique of
calculus. The value of SSresid is found
from

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 Finding the Least-Squares Line Finding the Least-Squares Line

The calculation of slope and intercept is simplified when

three quantities are defined. Sxx, Syy, and Sxy"' as follows:
1. The slope of the line, m:

2. The intercept. b:

Where xi and yi are individual pairs of data for x and y, N is the 3. The standard deviation about regression. sr:
number of pairs. And x and y are the average values for x and
y ; that is,

129 130

Finding the Least-Squares Line Finding the Least-Squares Line

4. The standard deviation of the slope, sm: 6. The standard deviation for results obtained
from the calibration curve, sc:

5. The standard deviation of the intercept. sb:
Equation above gives us a way to calculate the standard
deviation from the mean yc of a set of M replicate analyses of
unknowns when a calibration curve that contains N points is
used; recall that y is the mean value of r for the N calibration
points. This equation is only approximate and assumes that
slope and intercept are independent parameters, which is not
strictly true.

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33
 Finding the Least-Squares Line
The value of sr is related to SSresid by
The standard deviation about regression, also called
the standard error of the estimate or just the standard
error, is a rough measure of the magnitude of a typical
deviation from the regression line.
133
Interpretation of Least-Squares Result
We can also define a total sum of the squares,
SStot as
The coefficient of determination (R2) is a measure of
the fraction of the total variation in y that can be
explained by the linear relationship between x and y.
135
Interpretation of Least-Squares Result
The closer the data points are to the line predicted
by a least-squares analysis, the smaller are the
residuals. The sum of the squares of the residuals,
SSresid, measures the variation in the observed values
of the dependent variable (y values) that are not
explained by the presumed linear relationship
between x and y.
134
Interpretation of Least-Squares Result
The difference between Sstot and SSresid is the
sum of the squares due to regression. SSregr In
contrast to Ssresid, SSregr is a measure of the
explained variation. We can write.
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 Exercise Solution

Solve for a) The slope of the line, b) The intercept b, c) The standard
deviation about regression. Sr, 4) The standard deviation of the slope, sm,
5) . The standard deviation of the intercept. Sb, 6) total sum of the
squares, Sstot, 7) coefficient of determination (R2)

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137

Solution Solution

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 Assignment

Construct a calibration curve using the following

data:
Calculate:
1. Slope
Amount of Corrected
protein Absorbance 2. Y-intercept
0 0 3. The equation of the straight line
5 0.086 4. Linear coefficient (r2)
10 0.183 5. Standard deviation about regression,
14 0.246 Sr.
20 0.326 6. Standard deviation of the intercept, Sb.
25 0.384
7. Total sum of the squares, Stot.
8. Calculate the concentration of the
unknown sample having an absorbance
of 0.201.

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