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In the Classroom

Using Candy Samples To Learn about Sampling Techniques and Statistical Data Evaluation

Larissa S. Canaes, Marcel L. Brancalion, Adriana V. Rossi, and Susanne Rath* Department of Analytical Chemistry, Institute of Chemistry, State University of Campinas,13084-971 Campinas, SP, Brasil; *raths@iqm.unicamp.br

The first step in any chemical analysis is to obtain an ana- lytical sample of the bulk material. In effect, the reliability of an analytical result is often conditioned by the quality of the original sample. The sample must have the same chemical and physical properties of the raw material, so that it represents well the material that will be analyzed. In these cases, sampling is

directly responsible for the accuracy of the analytical results. The best way to sample would be to obtain large samples at random from the total population, based on the idea that as the sample size approaches the population size the errors decrease to zero (1). In practice, some factors—such as measurement costs and facilities for manipulating huge amounts of the bulk material— make it impractical to select large, essentially unlimited samples.

A typical random sample is usually far smaller than desired, rais-

ing concerns about how accurately the sample really represents the bulk material. This doubt can be answered by statistical analysis of the data (2). These facts justify the important need for students to un- derstand all the challenges involved in sampling techniques, the first step of any chemical analysis. In spite of that, students in classroom experiments are usually presented with homogeneous samples, so they tend to believe that sampling and statistical analyses are not problems that they have to deal with. Some references do describe ways to present to students how random sampling works and how it could be representative (1, 3), while

others propose different exercises emphasizing statistical analysis

of data (4–6). However, these exercises usually require extended

laboratory periods or are very theoretical. In 2000, Ross (7) proposed a simple, fast, and didactic classroom exercise using colored candies, which could easily demonstrate the effect of sample and particle size in sampling. However, this paper does not address statistics; this is more fully explored by Vitha and Carr (2). Inspired by these papers (2, 7), we developed and imple- mented a more complete classroom exercise for undergraduate and beginning graduate students to explore both sampling and statistics. It is an easy, interesting exercise that takes ~1.5 hours to demonstrate the effects involved in sampling techniques (sample amount and particle size and the representativeness of the sample in relation to the bulk material). This exercise also includes a simple statistical approach to commonly used parameters (mean, median, standard deviation, errors, quartiles, and confidence limits), presentation of results, graphs (histogram, box-plot, and whisker plot) and related tests (normality, outliers, significance) using parametric and non-parametric statistical methods.

Procedure

Materials For the sampling exercise, we used sugar-coated, round

chocolate candies available in several colors, sizes, and types. All

of the candies were purchased at a local market; the quantities

given should be adequate for undertaking this activity in a class of 10–35 students. The variety of sizes, shapes, and colors is important for representing heterogeneity in data.

• 10 packages (104 g) of M&M 1 candies

• 1 package (98 g) of M&M candies with peanuts

• 2 packages (35.2 g) of M&M Minis 2

• 1 package (80 g) of Confeti 3 chocolate candies

• Disposable gloves

• Plastic tray or paper plates

• Paper cups (50 and 200 mL capacities)

The gloves, plastic tray or paper plates, and paper cups were used in all the sample manipulation with hygiene in mind, so that the samples could be eaten at the end of the experiment. This exercise has been developed and used with students in both an undergraduate classical analytical chemistry course and a graduate course, Statistics in Analytical Chemistry, from 2000 to present.

Data Acquisition

In the first step, students were divided into ten groups and each group was responsible for data acquisition from one bag of the regular M&M candies. Each group counted the candies, separating and reporting them by color. The students were asked to compile the data and to start the statistical evaluation.

Parametric and Nonparametric Approaches

In order to show the students the theoretical and practical differences between parametric and non-parametric approaches, the raw data were statistically evaluated by comparison of param- eters, representations, and by the application of tests from both statistical methods. The results were also statistically compared to an average composition of the bags, provided at the manu- facturer’s Web site. 1

Sample Amount Effect—Part I

Next, all the M&M candies from the 10 bags that had been sorted by color were put together in a large plastic tray to simu- late the population of a “bulk material”. The students obtained this composition by aggregating the data of all 10 bags. Then, the groups collected “samples” of the bulk material, using two different sizes of cups (50 and 200 mL). Each group sampled five times. In the next step, a reducing procedure by quartering was used to enable the students to evaluate how representative each kind of sampling is. For the quartering procedure, the candies were uniformly spread inside a circular tray and then divided radially into quarters. The opposite quarters were combined. This process was repeated two more times. The statistical treat- ment of data was the same as used before.

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In the Classroom

Sample Amount Effect—Part II

All the candies were returned to the tray and some different candies (purple in color; 0.99% of the total in the bag) from another manufacturer were added to the mix. In our study, Con- feti 3 candies were used because they are very similar to M&Ms in terms of size. The two sampling procedures used in Part I and described above were also used in Part II, although just the purple candies were counted and analyzed statistically, because this condition simulates an analyte present in low concentra- tion as well as mimicking the effect of using different sampling methods.

Particle Size Effect The last observations explored the influence of different particle sizes on a sampling method, using a simple visual exer- cise. Students combined candies of different sizes (two bags of M&M peanut candies, one bag of M&M Minis, and some of the regular M&Ms used before) in a large glass jar, and mixed them very well to qualitatively observe the size distribution after different mixing procedures.

Data Acquisition

After counting and compiling the raw data, the students were asked to organize these data by percentage of candies of each color. We instruct students to be aware of significant figures and observe that the smallest unit possible is one candy. As the num- ber of candies per bag is about one hundred, the number of can- dies of each color has two significant figures. Thus, the percentage of candies must be represented by a maximum of two significant digits, without decimals. Table 1 reports the results from a typical classroom exercise used for the statistical treatments. In this exercise, each bag was considered as a sample origi- nating from a total population (in the manufacturer’s produc- tion line), the colors were the property being measured and the counting results were the measurements in a total of ten replicates represented by the number of bags used.

The Parametric Approach

In order to introduce the concepts important to a paramet- ric approach, the different ideas of random (or indeterminate), systematic (or determinate), and gross errors were presented. The instructor discussed how different types of error affect the final results. In the present experiment, a gross error is exemplified by accidentally dropping some or all of the candies (loss of sample):

as a consequence the experiment must be restarted with a new bag. In some instances one set of measurements apparently lies an abnormal distance from other values in a random sample from a population. Such measurements, called outliers, may be related to human errors and must be removed or corrected because they interfere with the precision and accuracy of the results. In a sense, this definition leaves it up to the analyst to de- cide what will be considered abnormal. The students were asked about possible outliers. It was explained that before abnormal observations can be singled out, it is necessary to characterize normal observations; a statistical test that can identify outliers should be used. Nevertheless, the students pointed out some possible outliers from Table 1; after they evaluated the data us- ing Dixon’s Q-test (8) at a 95% significance level (P = 0.05), no values were rejected. After that, the students were asked to represent the results in a simple way, retaining the sampling information—that is, the average value for the frequency of each color and the distribution about the mean. For this, they used the arithmetic means (x ) and the confidence interval of the error distribution for two different confidence intervals (95 and 99%), presenting these statistical concepts (8). The calculated parameters—means, standard de- viation (s), and relative standard deviation (RSD) or coefficient of variation (CV), as well as Student’s t values—are shown in Table 2, using the data from Table 1. Note that the mean, s, and RSD are presented with three significant figures—the total number of candies was higher than 1000 (four significant figures) and the number of candies of each color was given with three significant figures. For the confidence interval the results were rounded.

Table 1. Comparison of the Dispersion of Colors in the Candy Samples

 

Candies Classified by Color, %

 

Groups (Bags)

Candies per Bag

Blue

Brown

Green

Orange

Red

Yellow

1

127

29

13

13

10

18

17

2

127

23

11

12

16

8

30

3

128

27

14

13

20

8

18

4

119

15

9

15

26

16

19

5

114

11

19

11

23

12

24

6

118

10

15

11

24

20

20

7

115

15

14

10

28

11

22

8

119

10

13

12

30

17

18

9

115

18

11

14

24

17

16

10

114

12

10

12

27

13

26

Total

1196

In the next exercise the students had to verify whether a sig- nificant difference existed between the means reported for each color (n = 10 bags) and the values supplied by the manufacturer (used as the actual values, μ), in order to evaluate whether the data obtained from the sampling experiment accurately represent

the bulk material or bulk sample or, in this case, a large popula- tion, denoted by the provided values. Significance testing was introduced and a null hypothesis (H 0 ) was formulated: the two population means are equal. It is important to emphasize that to accept a hypothesis does not mean that it is true, only that we do not have evidence to believe otherwise. Thus hypothesis tests are usually stated in terms of both a condition that is doubted (null hypothesis) and a condition that is believed (alternative hypothesis). In our study the alternative hypothesis would be that the two population means are not equal. The students are asked to test the hypothesis using a t-test (8) for each color and

a significance level of 0.05. The significance level, P, defines the

sensitivity of the test. A value of P = 0.05 means that we inadver-

tently reject the null hypothesis 5% of the time when it is in fact true. The choice of P is somewhat arbitrary, although in practice

a value of 0.05 is commonly used in analytical chemistry.

In the Classroom

The comparisons were made by using the critical values of t for 9 degrees of freedom 2.26 and 3.25, respectively, for significance levels of P = 0.05 and P = 0.01 (8). Significant differences were observed for both significance levels between the samples evaluated and the population (supplied by the manufacturer) just for the green amount of candies, the color that also presented the lowest standard deviation and, as a con- sequence, the lowest confidence interval (see Table 2). For the green candies, the null hypothesis is rejected as it is statistically understood, since a data distribution with a smaller dispersion means greater precision near the arithmetic mean. In this way, any value that is not very close will not be contained inside the confidence interval provided by the Gaussian distribution curve. The opposite effect could be observed for bigger dispersions of data (see the data for the blue and orange candies). New information was introduced to the students. Bags 1–3 and bags 4–10 came from sample batches A and B, respec- tively. Now it was asked whether a significant difference existed between the two sample batches of the candies in relation to each color (see Table 3). In this case, we compared two sample

means x A and x B , which correspond to sample batches A and B,

Table 2. Parametric Approach to Analyzing the Data

 

Candies Classified by Color, %

 
 

Data Parameters a

Blue

Brown

Green

Orange

Red

Yellow

Values supplied by the manufacturer b

14.3

14.3

14.3

21.4

14.3

21.4

Mean x (n =10)

 

17.0

12.9

12.3

22.8

14.0

21.0

Standard deviation (s )

007.06

02.88

01.49

06.03

04.22

04.47

RSD c

41.5

22.3

12.1

26.4

30.1

21.3

Confidence interval: d P = 0.05

17±5

13±2

12±1

23±4

14±3

21±3

Confidence interval: d P = 0.01

17±7

13±3

12±2

23±6

14±4

21±5

Student’s t = | x – μ|n ½ /s (t statistic values)

01.2

01.4

04.9

0.84

0.23

0.28

Is there a significant difference? e

No

No

Yes

No

No

No

a Based on the data reported in Table 1.

b These mean values are reported at the manufacturer’s Web site: http://global.mms.com/br/about/products/milkchocolate.jsp (accessed Jun 2008).

c RSD is the relative standard deviation, and is given by 100(s/ x ).

d The confidence interval is determined by ( x t ) s/n ½ < μ < ( x + t ) s/n ½ , where t is the critical value of Student’s t-test. Confidence levels of 95% and

99% are represented by probability values in which P = 0.05 or 0.01, respectively. e Comparison of the mean value supplied by the manufacturer and the mean value of the ten bags evaluated in this experiment.

Table 3. Parametric Comparison of the Two Random Samples of Candies

 

Candies Classified by Color, %

 

Parameters

Blue

Brown

Green

Orange

Red

Yellow

Sample batch A Mean, x A (n = 3)

26

13

13

15

11

22

Sample batch B Mean, x B (n = 7)

13

13

12

26

15

21

Sample batch A standard deviation, s A

3.0

1.5

0.58

5.0

5.8

7.2

Sample batch B standard deviation, s B

3.0

3.4

1.8

2.5

3.2

3.5

Is there a significant difference?

Yes

No

No

Yes

No

No

Note: Significance of the comparison of the mean values between the two sample batches ( x A x B ) evaluated at P = 0.5.

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In the Classroom

respectively. Taking the null hypothesis that the two means are

equal, we need to test whether (x A x B ) differs significantly from zero. First the F-test was applied for the comparison of standard deviations (8). Both samples had standard deviations that did not differ significantly, which allows calculation of a pooled estimate of standard deviation from the two individual standard deviations, s A and s B . In turn, the value of t was obtained and compared with the critical value of t, using 8 degrees of freedom [(n A + n B ) − 2]. Typical statistical tests incorporate assumptions about the underlying normal (Gaussian) distribution of data, and hence rely on distribution parameters. Statistical values such as means, standard deviations, and confidence limits are, strictly speaking, for a large population size. In analytical chemistry,

35 30 25 20 15 10 5 0 blue brown green orange red yellow Fraction
35
30
25
20
15
10
5
0
blue
brown
green
orange
red
yellow
Fraction of Candy (%)

Color of Candy

Figure 1. Box-and-whisker plots for fractions (%) of each color of candies. (The black squares inside the boxes represent the means).

Table 4. Comparison of the Five-Number Summary for Each Color

Sample

Minimum,

Lower

Median,

Upper

Maximum,

Color

%

Quartile, %

%

Quartile, %

%

Blue

10

11

15

23

29

Brown

09

11

13

14

19

Green

10

11

12

13

15

Orange

10

20

24

27

30

Red

08

11

13

17

20

Yellow

16

18

19

24

30

Table 5. Comparison of Results Obtained by Parametric and Nonparametric Approaches

Sample

Parametric

Nonparametric

Color

(Mean), %

(Median), %

Blue

17.0

15

Brown

12.9

13

Green

12.3

12

Orange

22.8

24

Red

14.0

13

Yellow

21.0

19

we generally deal with small sets of data, sometimes fewer than five results, and in some instances we are interested in methods that do not require the assumption of normally distributed data. Methods that make no assumptions about the shape of a data set’s distribution are called nonparametric or distribution- free methods.

A Nonparametric Approach

A nonparametric approach to data analysis uses the same

data (Table 1); however, instead of the mean, the students are now asked to calculate the median for each color from the 10 bags. In addition, the lower and upper quartiles should be calcu- lated, as well as the smallest (minimum) and the greatest (maxi- mum) values in the distribution. Values categorized into these five rankings are then represented in a simple visual way by a box- and-whisker plot (8) (Figure 1), where the immediate visuals are the center, the spread, and the overall range of distribution. A box-and-whisker plot consists of a rectangle (the box) with two lines (the whiskers) extending from opposite edges of the box, and a further line in the box, crossing it parallel to the edges. The ends of the whiskers indicate the range of the data, the edges of the box from which the whiskers protrude represent the upper and lower quartiles, and the line crossing the box represents the median of the data. A box-and-whiskers plot, accompanied by a numerical scale, is a graphical representation of the five-number summary, thus, the data set is described by its extremes, its lower and upper quartiles, and its median (see Table 4). The plot shows at a glance the spread and the symmetry of the data (8). After considering these results, no values were rejected. The comparison of results obtained by parametric and non- parametric approach is shown in Table 5. The differences between the mean and the median are not significant, indicating that the data can be drawn from a normal distribution, which makes sense for the sampling exercise used. One method of testing this hypothesis is by using a χ 2 test. This method, unfortunately, is only reliable in cases with at least 50 data points.

Sample Amount Effect

At this point the students are told of the relationships of the operations involved in sampling and analysis. The concepts of primary sample (bulk sample), reduced sample, subsample, laboratory sample, and test sample are discussed. It is pointed out that the term “sample” implies the existence of sampling error, which arises from a lack of homogeneity in the popula- tion. Since sampling error is always associated with analytical

error, it must be isolated by the statistical procedure of analysis of variance (9). It will be assumed in our experiment that all the candies from the 10 bags represent the bulk sample and the task now is to obtain the laboratory sample from the bulk sample of the material.

In this part of the exercise the students discussed and simu-

lated different conditions of sampling (using different containers and reducing by quartering) from a bulk sample with known composition, which was obtained by mixing all the candies in a large container. The data collected and the basic statisti- cal parameters calculated are presented in Table 6, in terms of percentages. The values of the relative errors for each sampling procedure are shown in parentheses. The same statistical treat- ments applied before were also used in this case.

In the Classroom

Table 6. Dispersion of Candies Reported by Color for Each Sample

 

Candies Classified by Color, % (Relative Error)

 

Sampling Methods

Total Candies

Blue

Brown

Green

Orange

Red

Yellow

Observed means of the population (µ)

 
 

1196

17.0

12.9

12.3

22.8

14.0

21.0

Sampling with a small container (50 mL cup)

 

1 0040

15 (−12)

08 (−38)

15 (−22)

10 (−56)

20 (−43)

32 (−52)

2 0049

16 (−5.9)

16 (24)

04 (−67)

33 (45)

06 (−133)

25 (19)

3 0043

18 (5.9)

12 (−7.0)

19 (54)

19 (−17)

21 (50)

11 (−47)

4 0042

17 (0)

12 (−7.0)

17 (38)

21 (−7.9)

09 (−35)

24 (14)

5 0035

14 (−18)

06 (−53)

20 (63)

06 (−74)

23 (64)

31 (48)

Sampling with a large container (200 mL cup)

 

1 0177

15 (−12)

11 (−15)

16 (30)

22 (−3.5)

11 (−21)

25 (19)

2 0153

18 (5.9)

12 (−7.0)

11 (−11)

20 (−12)

14 (0)

25 (19)

3 0185

24 (41)

15 (16)

10 (−19)

19 (−17)

13 (−7.1)

19 (−9.5)

4 0191

15 (−12)

17 (32)

12 (−2.4)

20 (−12)

17 (21)

19 (−9.5)

5 0192

19 (12)

09 (−30)

15 (22)

24 (5.3)

12 (−14)

21 (0)

Reducing by quartering

1

0265

16 (5.9)

14 (8.5)

12 (2.4)

23 (0.0088)

13 (7.1)

22 (4.7)

Note: Values in parenthesis are the relative errors, calculated by 100| x – μ|/μ, where µ is the known value of the bulk sample.

Table 7. Comparison of the Percentage of Purple Candies in Each Sample

 

Purple Candies or “Analyte”, % (Relative Error)

 

Samples

Actual Values (µ)

Sampled with a 50 mL Cup

Sampled with a 200 mL Cup

Reduced by Quartering

1 0.99

2.5 (152)

1.0 (1.0)

0.98 (–1.0)

2 0.99

0.0 (–100)

1.9 (92)

3 0.99

1.9 (92)

0.49 (–50)

Note: Values in parenthesis are the relative errors, calculated by 100| x – μ|/μ, where µ is the known value of the bulk sample with purple candy added.

It is possible to observe in these experiments that the number of candies sampled influences the values of the relative errors. Whereas with the small sampling cup the relative errors varied from 133 to 64%, with the larger cup the values range between 30 to 41%. Thus, the larger container resulted in a rela- tive error about three times smaller than provided by the smaller one. These numbers elucidate to the students the improvement in sampling caused by increasing the sample amount from ap- proximately 42 to 180 candies per collection. For reducing by quartering—even though the procedure was only made once, in contrast to the five replicates for the other samplings—the relative error observed was 0.0088 to 8.5%, still smaller than the values observed with the other sampling procedures. This is because reducing by quartering results in a larger sample (265 candies in this case) and because this method was developed to optimize a sampling condition, resulting in smaller errors (8). Another discussion topic concerns dispersion of data points resulting when the same sample procedure is used. The values were graphically presented (Figure 2), so students can observe that a greater dispersion usually occurs with a smaller sampling con- tainer than with a larger one (blue and brown were exceptions).

In the same way, the students also simulated a sample with an analyte present in low concentration, by the addition of 0.99% purple candies to the total material. Once more, the effect of using different sampling procedures was evaluated, as summarized in Table 7.

40 Sample Size: 50 mL 200 mL 30 20 10 0 blue brown green orange
40
Sample Size:
50 mL
200 mL
30
20
10
0
blue
brown
green
orange
red
yellow
Fraction of Candy (%)

Color of Candy

Figure 2. Percentage of candies sampled using the small container (50 mL) and the larger container (200 mL). The solid bar represents the expected value of each color, as provided by the manufacturer. 1

1087

In the Classroom

In the Classroom Figure 3. The size gradient formed by the different-sized candies mixed inside a

Figure 3. The size gradient formed by the different-sized candies mixed inside a glass jar.

The students could note again the reduction in sampling error as the sample amount is increased, denoted by the decrease in relative errors obtained using the small cup, the larger cup, and by quartering. This means that, as the amount sampled becomes larger, it better represents the bulk sample, up to the limit of the entire sample, which represents the actual value of the material.

Particle Size Effect

In this exercise, students easily observe the effect of differ- ent particle sizes during the sampling of solid materials. All the different candies were mixed together inside the flask, resulting in a size distribution in which the smaller candies accumulated at the bottom of the flask, while the bigger candies were more evident at the top of the flask. Figure 3 shows a photograph of this phenomenon. After this visual experiment, the students were questioned about the errors in sampling that might result from this effect, namely, size segregation in real samples. The students were also asked about possible ways of eliminating this type of error. In real applications, for example, the simplest procedure used is a

sequence of three unitary operations: grinding, homogeniza- tion (by mixing), and separation of the samples by different ranges of mesh (defined ranges of particle sizes) before sam- pling occurs.

Conclusions

This classroom experiment with candies has been used for six consecutive semesters in a chemistry course for under- graduates, and in a graduate chemistry course. It successfully introduced the undergraduate students to important concepts of statistics and sampling techniques. This approach is easy to implement and engages students in learning in a stimulating way that is lucid and concrete—as well as tasty—because all the statistical data used was obtained by the students themselves.

Notes

1. M&M candies have chocolate interiors; some also have peanuts

or almonds in the center. For more information, see the manufacturer’s Web page: http://global.mms.com/br/about/products/milkchocolate.jsp (accessed Jun 2008).

2. As the name implies, M&M Minis are smaller-sized than the

conventional version.

3. Confeti candies are manufactured by Kraft Foods Brazil S. A.

Acknowledgments

The authors are grateful to all the students who participated in the exercises, and thank C. H. Collins for language assis- tance.

Literature Cited

1. Cohen, R. D. J. Chem. Educ. 1992, 69, 200–203.

2. Vitha, M. F.; Carr, P. W. J. Chem. Educ. 1997, 74, 998–1000.

3. Cohen, R. D. J. Chem. Educ. 1991, 68, 902–903.

4. Salzsieder, J. C. J. Chem. Educ. 1995, 72, 623–625.

5. Carter, D. W. J. Chem. Educ. 1985, 62, 497–498.

6. Spencer, R. D. J. Chem. Educ. 1984, 61, 555–563.

7. Ross, M. R. J. Chem. Educ. 2000, 77, 1015–1016.

8. Miller, J. C.; Miller, J. N. Statistics for Analytical Chemistry, 3rd ed.; Ellis Horwood PTR Prentice Hall: New York, USA, 1994.

9. Horwitz, W. Pure Appl. Chem. 1990, 62, 1193–1208.

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