MS

Chapter 3.1

A Comparative Study on the Influences of Tea Hydration for the Method Efficiency of Pesticide Multiresidues Using Three Sample Preparation Methods/GC–MS/MS Chapter Outline 3.1.1 Introduction 3.1.2 Experimental Method 3.1.2.1 Three Different Sample Preparation Methods 3.1.2.2 Aged Sample Preparation 3.1.2.3 Incurred Sample Preparation 3.1.3 Experimental Results and Discussion 3.1.3.1 Comparison of Accuracy and Precision for Fortification Recovery Experiments by the Three Methods

497 499 499 499 499 500

3.1.3.2 Correlation Comparison of the Three Methods’ Extraction Efficiency With Pesticide log Kow Values 3.1.3.3 General Analysis of Method Applicability 3.1.3.4 Comparison of the Cleanup Efficiency for the Three Methods 3.1.4 Conclusions References

501 504 508 508 512

500

3.1.1 INTRODUCTION Tea is rich in physiologically active compounds, such as polyphenols, alkaloids, tea pigments, aromatic substances, amino acids, and vitamins, and it possesses such functions as reducing blood sugar, blood pressure, antithrombosis and antiatherosclerosis, bacteriostat, improving immunity, and is antitumor, in other words, a naturally healthy drink. More than 2 billion people from 150-plus countries and regions drink tea, making tea one of the three most popular drinks in the world. Tea is mostly planted in warm temperate zones and subtropical areas which are susceptible to threats from plant diseases and insect pests year-round. For the purpose of plant disease and insect pest prevention, chemical pesticides are widely used, leading to contamination with pesticide residues. To hold this in check, countries worldwide have prescribed pesticide maximum residue limits in tea (MRL). For instance, the EU has stipulated 453 varieties, Japan 268, and Germany 530. Hence, the acceleration of the development of techniques for pesticide multiresidues in tea from GC [1] to GC–MS [2], GC–MS/MS [3], LC–MS/MS [4],UPLC–MS/MS [5], and GC×GC-ToF [6], and so on. Tea has a complex matrix with pigments, polyphenols, alkaloids, and some lipids that act as interferences for determination of pesticide residues in tea. The question of what pretreatment techniques should be adopted to sufficiently extract pesticide multiresidues in tea matrices, lowering interference from foreign matter to the greatest extent, is one of the most difficult projects in the tea residue analytical field. For pesticide residues in tea, two modes of extraction are usually adopted, one of which is using common organic solvents or their composite mixing solvents, such as acetonitrile, hexane, cyclohexane, ethyl acetate, acetone, dichloromethane, or methanol. Alternatively, making an extraction in a mode such as homogeneous [7], oscillating [8], ultrasonic [4], vortex [9], accelerated solvent extraction [10], matrix solid-phase dispersion [11], head space solid-phase microextractions Analysis of Pesticide in Tea. http://dx.doi.org/10.1016/B978-0-12-812727-8.00008-0 Copyright © 2018 Chemistry Industry Press. All rights reserved. Published by Elsevier.

497

498 3  Study on the Influences of Tea Hydration

[6], pressurized solvent extraction, or dispersive liquid–liquid microextraction [12]. Acetonitrile extraction, among those listed earlier, possesses a wide range of pesticide polarities and less interference from coextracting matters, making it the first choice of solvents for pesticide multiresidue extraction in years past. The most widely used cleanup techniques for multiresidues in tea is solid phase extraction (SPE), which features lesser loss of pesticide residue and good cleanup results. The cleanup fillers mainly used include Carb-NH2 [13], Carb-PSA, and Florisil [1]. Pang et al. [14,15] employed acetonitrile homogeneous extraction and SPE cleanup and established a simultaneous determination method for 653 pesticides in tea, with an appraisal study of different kinds of analytical conditions, among which the limit of detection (LOD) by GC–MS for 490 pesticides was 1.0–500 µg/kg, while the LOD by LC–MS/ MS for 448 pesticides was 0.03–4820 µg/kg. For GC–MS determination of 490 pesticides at the low fortification level of 0.01 –100 µg/kg, the average recoveries for 94% of pesticides fall within 60%–120%, with RSD for 77% pesticides below 20%; for LC–MS/MS determination of 448 pesticides, the average recoveries for 91% of pesticides fall within 60%–120%, with RSD for 76% pesticides below 20%. The new-type SPE cartridge Cleanert TPT, composed of three ingredients: graphitized carbon black (PestiCarb), polyamine silica, and amide polystyrene, is high in cleanup efficiency and is of good reproducibility and repeatability in analytical results. Other common cleanup techniques include liquid–liquid extraction [8], DSPE [3], GPC [10], DLLME [12]. Schurek et al. [6] adopted head space solid phase microextraction in conjunction with the GC×GC-TOF approach for determination of 36 pesticide residues in tea. Ana Lozanoa et al. [3] adopted GC–MS/MS and LC–MS/MS for respective determination of 86 pesticide residues in green tea, black tea, dark tea, and jasmine flower tea, with a much lower limit of detection with LC–MS/MS than GC–MS/MS. Another way of extraction is first hydrating samples before adopting organic solvents for extraction. For hydration of tea leaves, adding water quantities varies from 1 mL/g to 10 mL/g as different hydration methods are used. The varieties of pesticides determined by these analytical methods vary from more than a dozen to more than 100. For example, Hong-Ping Li et al. [8] used acetone to extract the hydrated tea leaves, employed 5% NaCl water solutions (1:1:5, v/v) liquid–liquid partitioning for cleanup, and made a GC determination of 84 pesticide residues, with recoveries 65%–120% and RSD 0.34%–16% at the fortification concentration of 0.02–3.0 mg/kg. Zhiqiang Huang et al. [2] used acetone, ethyl acetate, and hexane mixing solutions for extraction and GPC in conjunction with SPEC for cleanup, making a GC–MS determination of 102 pesticide residues in tea, with recoveries 59.7%–120.9% and RSD 3.0%–20.8% at the fortification concentrations of 0.01–2.5 µg/m. Ana Lozano et al. [3] used acetone oscillation for extraction and PSA and DSPE for cleanup after salting out, making a GC–MS/MS and LC–MS/MS determination of 86 pesticide residues in black tea, green tea, dark tea, and jasmine flower tea, with recoveries 70%–120% and RSD less than 20% at the fortification concentration of 10–100 µg/kg. LOD is 0.1–210 µg/kg. For comparison of the superiority of these two modes of extraction, there are only two papers that have been found for appraisal study, one of which is Pang et al. [16], who use hydration (4 mL/g) homogeneous extraction and SPE cleanup for determination of 201 pesticide residues in tea. The comparison of the hydration method with the other two has found that both accuracy and precision with the hydration method is poor. Tomas Cajka et al. [17] proposed that tea samples be added with water and made to sit still for 30 min, which effectively helps improve the extraction efficiency of pesticide residues in tea. For a further probe into the influences of hydration on the method efficacy of tea multiresidues, we chose M1 to be equivalent to Cajka’s method [17] and used acetonitrile for extraction after tea hydration, then used part of the extraction solution and hexane liquid–liquid partitioning for cleanup. The extraction of M2 is identical to that of M1, but it takes all the extraction solution and cleans up with SPE, with cleanup procedures the same as those of M3. M3 is equivalent to Pang’s method [14]: sample extraction through pure acetonitrile and SPE for cleanup. There only exist cleanup differences with M1 and M2, while extraction differences only exist with M2 and M3. Through recovery experiments at three fortification levels, aged sample determination experiments, and field incurred sample determination, aspects such as extraction efficiencies, cleanup results, and the applicability of the process to different methods have been studied and compared. These results reflect the concrete influences of hydration and nonhydration on the method efficiency. Recoveries from 456 pesticides at three fortification levels and the log Kow function correlation diagram of them with pesticide polarities have both been proven. First, tea leaf hydration is indeed capable of increasing extraction efficiency of certain polarity pesticides—for instance, 24 pesticides of strong polarities, with recoveries of M2 higher than those of M3. Hydration also lowers the extraction efficiency of some nonpolar pesticides, however. Taking the 28 nonpolar pesticides for an example, recoveries with M3 are higher than M2. The appraisal of overall extraction efficiencies of 456 pesticides has found that the hydration method does more harm than good. In addition, interference from coextraction after tea hydration increases greatly, leading to a decrease of method efficiency. Fortification experiments of uniform limit 0.010 mg/kg confirm that for the percentage of pesticides that comply with EU SANCO/12495/2011 technical requirements of 70%–120%

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499

and RSD <20%, only 5.0% are accounted for with M1 hydration method, but 50% with M3 nonhydration method. (M1 fails to meet the requirement.) Although the influence is not obvious for determined values of high-concentration fortification, it has a relatively big influence on RSD of the method. For instance, with 456 pesticide aged samples, there are 158 pesticides with their added RSD <10% for M1, accounting for 35%; there are 381 for M3, making up 84%, and M3 is 2.4 times superior to M1. Additionally, tea leaf hydration greatly lowers the sensitivity of the method. In a study of 456 pesticide aged tea samples, the signal/noise ratio of each pesticide was statistically calculated and the adding of signal/noise ratios of 456 pesticides was averaged, with M1 being 940 and M3 6781, showing that hydration lowered the sensitivity 7 times. This is also the main cause of failure of detection at 0.010 mg/kg to meet EU SANCO/12495/2011 technical requirements. The log Kow values of 329 pesticides have been found in this study of 456 pesticides, among which there are only 39 polarity pesticides and strong polarity pesticides with log Kow values <2.0, accounting for 12%. There are 290 other pesticides of medium polarity, weak polarity, and nonpolarity, accounting for 88%. Therefore, hydration methods only increased the extraction efficiency of certain polarity pesticides while losing extraction efficiency for the majority of pesticides of medium polarity and nonpolarity, which is not worthwhile.

3.1.2  EXPERIMENTAL METHOD 3.1.2.1  Three Different Sample Preparation Methods Method 1(M1): hydration + oscillating extraction + hexane liquid/liquid partitioning cleanup for part of extraction solution. It is equivalent to Cajka’s method [17]; method 2 (M2): hydration + oscillating extraction + overall extraction solution SPE cleanup, that is, the extraction is identical with M1, and cleanup is identical with M3; method 3 (M3): pure acetonitrile homogeneous extraction + overall extraction solution SPE cleanup. It is equivalent to Pang’s method [14]. The differences in the three methods are demonstrated in Fig. 3.1.1. Fig. 3.1.1 shows that M1 and M3 are two totally different methods, while M2’s extraction is identical with M1 and its cleanup is identical with M3, so the test results of M2 can testify to the advantages and disadvantages of M1 and M3.

3.1.2.2  Aged Sample Preparation Four hundred and fifty-six pesticide mixing solutions are uniformly sprayed onto blank oolong tea powders, which are stored in the dark and aged 30 days. Quarter sampling is adopted, and a parallel determination of 456 pesticide concentrations is made for the sample. When RSD <4% (n = 10) is determined for each pesticide, it is considered to be a uniformly prepared aged tea samples.

3.1.2.3  Incurred Sample Preparation Market-available 18 pesticides were sprayed in accordance with spraying procedures onto tea trees grown in the field. A parallel spray was conducted on two experimental fields. The first picking began 24 h after spraying, with picking

FIGURE 3.1.1  Sketch of the differences of M1, M2 and M3.

500 3  Study on the Influences of Tea Hydration

continuing once every day, lasting 1 month. On the 10th day after spraying the tea is picked to be an incurred sample and is prepared into uniform (RSD < 4%, n = 10) incurred samples.

3.1.3  EXPERIMENTAL RESULTS AND DISCUSSION This experiment used three sample preparation methods, M1, M2, and M3. (1) For 456 pesticides in oolong tea, a fortification experiment was conducted at three levels of high, medium, and low residues, with recoveries and RSD data tabulated in Supplemental Tables 1–3; (2) for 456 pesticides, S/N of three fortification levels for oolong tea matrix standards were tested, with S/N raw data tabulated in Supplemental Tables 4–6; (3) for 456 pesticides in oolong tea aged samples, the concentration values and S/N were tested, with finding values, RSD (n = 5), and S/N data tabulated in Supplemental Table 7; (4) for 160 pesticides determined by Cajka’s method [17], using two methods for sample preparation, recoveries determined at three fortification levels and RSD data were tabulated in Supplemental Table 8; (5) for 18 pesticide green tea incurred samples, three methods were used for sample preparation and their true content was tested and tabulated in Supplemental Table 9. Due to the huge quantity of data, supplementary materials have been provided for the reader to reference or trace the source when necessary.

3.1.3.1  Comparison of Accuracy and Precision for Fortification Recovery Experiments by the Three Methods The three methods of M1, M2, and M3 are used to conduct a comparative experiment on the fortification recoveries at three levels (0.01, 0.1 and 1.0 mg/kg) for 456 pesticides determined in Pang’s method [14], with the test raw data listed in Supplemental Tables 1–3. The three methods of M1, M2, and M3 are used to conduct a comparative experiment on fortification recoveries at three levels for 160 pesticides determined by Cajka’s method [17], with data listed in Supplemental Table 8. Pesticide quantities and ratios with their fortified recoveries 70%–120% and RSD ≤ 20% in conformance with EU SANCO/12495/2011 technical requirements tabulated in Supplemental Tables 1–3 and Supplemental Table 8 are listed in Table 3.1.1. Table 3.1.1 shows that in terms of EU SANCO/12495/2011 technical requirements with recoveries (REC) 70%–120% and RSD ≤20%, two items of REC and RSD at three fortification levels with M3 meet the technical standards of EU SANCO/12495/2011, with pesticide quantities determined in excess of those with M2 and M1. In a practical experiment, one notable difference is also discovered. For M1 and M2, take 2 g oolong tea samples, with supernatant about 8–8.5 mL obtained through final centrifuging. For M1, take 1 mL acetonitrile supernatant (equivalent to 0.2 g tea) for cleanup; for M2, take overall acetonitrile supernatant (equivalent to 2 g tea) for cleanup. In fact, for 2 g samples, 8–8.5 mL supernatants are actually obtained, and based on this calculation, 1 mL acetonitrile is equivalent to 0.235–0.250 g tea; on the contrary, if 1 mL acetonitrile supernatant is equivalent to 0.2 g tea, 8–8.5 mL supernatant is equivalent to 1.6–1.7 g tea. Based on this calculation, due in part to extraction solutions adopted with M1, the analytical results from the pesticides are falsely higher than those of M2 at 17.5%–25.0%, with an average of 21.25%.

TABLE 3.1.1 Recoveries, RSD and Ratios (%) That Meet EU SANCO/12495/2011 Technical Requirements 1.00 mg/kg

0.10 mg/kg

0.01 mg/kg

AVE REC 70%–120%

RSD ≤ 20%

AVE REC 70%–120%

RSD ≤ 20%

AVE REC 70%–120%

RSD ≤ 20%

M1

121(75.5)

159(99.4)

109(67.9)

144(89.9)

23(14.5)

24(15.1)

M2

134(84.3)

155(97.5)

99(62.3)

128(80.5)

18(11.3)

52(32.7)

M3

145(90.6)

160(100)

121(75.5)

156(97.5)

80(49.7)

112(69.8)

M1

335(73.5)

387 (84.9)

313 (68.6)

287 (62.9)

121 (26.5)

156(34.2)

M2

353 (77.4)

313 (68.6)

358 (78.5)

345 (75.7)

350 (76.8)

287 (62.9)

M3

404 (88.6)

399 (87.5)

417(91.4)

413 (90.6)

325 (71.3)

303 (66.4)

Fortified concentrations For 160 target pesticides

For 456 target pesticides

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TABLE 3.1.2 Total Average Recoveries for Pesticides at Three Fortification Levels by Three Methods Fortification levels, mg/kg

M1a

M2

M3

1.0

61.8%

85.3%

88.0%

0.1

69.2%

84.9%

89.3%

0.01

52.7%

86.3%

83.4%

Average

61.2%

85.5%

86.8%

a

Deducting falsely high number 20.0%.

The same thing also happened in comparison with M1 and M3, where the acetonitrile extraction solutions added with M3 are 30 mL and the supernatants obtained through centrifuging are 25 mL. Theoretically, when M1 and M3 are compared, the analytical results with M1 should be falsely higher by 20.0% than those with M3. See Table 3.1.2. If, in deference to the deduction about the falsely high number 20.0%, it is deducted from the analytical results brought about by the procedure differences of M1, the average extraction efficiency ranking for the three methods is M3 > M2 > M1.

3.1.3.2  Correlation Comparison of the Three Methods’ Extraction Efficiency With Pesticide log Kow Values 3.1.3.2.1  Correlation Comparison of 456 Pesticide Fortified Recoveries With Pesticide log Kow Values For the purpose of evaluating the correlations of the recoveries and pesticide polarities of the three methods, log Kow values have been found for 329 pesticides out of 456 pesticides, with their range: −0.77 to +8.20. Take the log Kow values as horizontal ordinate and recoveries as vertical ordinate to draw the scatter diagram (eliminate recoveries being zero and the outliers exceeding 150%), and the correlation diagram of the recoveries of the three methods at three fortification levels and pesticide polarities is determined (see Fig. 3.1.2). To further prove that hydration influences the extraction efficiency

FIGURE 3.1.2  Correlation distribution for pesticide recoveries of M1, M2 and M3 at three fortification levels with log Kow in the range of −0.77–8.20.

502 3  Study on the Influences of Tea Hydration

TABLE 3.1.3 Distribution of Recoveries for 329 Pesticides by log Kow Sector log Kow range

Pesticide polarities and number (Total:329)

1.0 mg/kg

0.1 mg/kg

0.01 mg/kg

M1

M2

M3

M1

M2

M3

M1

M2

M3

<1

Strong polarity

14

29.0

103.3

80.9

42.2

91.1

85.8

37.9

64.0

51.8

1–2

Polarity

25

38.4

101.3

82.0

65.3

87.2

82.7

75.0

87.3

71.1

2–3

Medium polarity

65

68.8

86.6

85.9

78.7

83.3

85.1

60.5

86.9

81.2

3–4

95

82.7

80.7

86.5

82.9

80.7

86.4

55.0

81.8

77.7

4–5

72

83.1

77.6

85.6

84.4

79.3

86.7

57.0

79.9

76.5

5–6

Weak polarity

27

73.5

71.7

84.4

74.9

72.0

81.1

47.3

64.1

70.1

>6

Nonpolar

31

70.2

62.7

87.2

59.4

63.5

88.8

48.8

61.2

64.8

63.7

83.4

84.6

69.7

79.6

85.2

54.5

75.0

70.5

AVE

of pesticides of different polarities, 329 pesticideswere divided into 7 sectors per log Kow values <1, 1–2, 2–3, 3–4, 4–5, 5–6, and >6 to statistically calculate the corresponding pesticide number, average recoveries, RSD, and mean S/N values for each sector (see Table 3.1.3). Recoveries, RSD, and S/N values can be traced to Supplemental Tables 1–6. Fig. 3.1.2 shows that the number and ratios (%) of pesticides with recoveries 70%–120% at three fortification levels are 240 (73%), 232 (71%), and 94 (29%) for M1; 251 (76%), 265 (81%), and 256 (78%) for M2; and 296 (90%), 303 (92%), and 236 (72%) with M3. There are more varieties with M3 than M1 and M2. Moreover, the absolute majority of pesticide recovery values are relatively concentrated, which demonstrates that the M3 method has relatively wide application for a sufficient and balanced pesticide extraction of different polarities. Table 3.1.3 shows: (1) there are 39 pesticides of relatively strong polarity in the range of log Kow <1 and 1–2, and at the three fortification levels. The recoveries for the overall ranking of the three methods is M2 > M3 > M1 because hydration has increased the extraction efficiency of these pesticides, enabling M2 > M3; the hexane liquid–liquid partitioning adopted for M1 has lowered the partitioning ratios of these polarity pesticides, causing recoveries to drop greatly; (2) there are 232 pesticides of medium polarity in the range of log Kow of 2–3, 3–4, and 4–5. At the high and medium fortification levels, the extraction efficiency for pesticides in these three ranges is balanced with M3, and their average recoveries are higher than M1 and M2, while with M2 pesticide recoveries within these three ranges drop regularly because with the drop of pesticide polarity hydration extraction efficiency obviously decreases. (3) There are 58 pesticides of weak polarity and nonpolarity in the range of log Kow 5–6 and >6, and recoveries for these pesticides with M1 and M2 are all obviously lower for than M3. This is because hydration has led to the marked drop of extraction efficiency in pesticides with weak polarity. The aforementioned analysis indicates that the hexane liquid–liquid partitioning adopted for cleanup with M1 sequentially and markedly lowers the pesticide extraction efficiency in the four sectors of 3–4, 2–3, 1–2, and <1.The hydration effect also gradually and sequentially lowers the extraction efficiency of nonpolar pesticides in the sectors of log Kow 5–6 and >6.

3.1.3.2.2  Changing Trend of Pesticide Polarities and Method Recoveries To further prove the correlation of method recoveries and pesticide polarities, as per Table 3.1.3, choose 0.1 mg/kg fortified concentrated samples. Take the average value of recoveries for each sector obtained by the three methods as longitudinal ordinate, and the midvalues of the corresponding log Kow values as horizontal ordinate, to draw a histogram and draw a fortification recovery trend line (Fig. 3.1.3). Next, the three methods of M1, M2, and M3 are independently analyzed. Fig. 3.1.3 shows that for the M1 trend line, the method recoveries increase—before decreasing with the pesticide polarities changing from strong to weak. This is because hydration lowers the extraction efficiency of pesticides with weak polarities, while the adoption of hexane liquid–liquid partitioning for cleanup causes the loss of polar pesticides, thus leading to an arc trend line high in the middle and low at both ends. For M2, hydration procedures are likewise adopted, increasing the extraction efficiency of pesticides with strong polarities and lowering the extraction efficiency of pesticides with weak polarities. Recoveries obviously take a high-tolow trend, demonstrating M2 has a relatively strong selectivity for pesticides of different polarities. For M3, deviations are small for recoveries of pesticides of different polarities without obvious fluctuations, which takes a balanced, stable, and similar-to-straight trend line.

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FIGURE 3.1.3  The recovery change diagram for 7 polarity sectors by the three methods.

We then make another comparative analysis of M1 and M2, M2 and M3, and M1 and M3, respectively. Likewise Fig. 3.1.3 shows that when comparing M1 and M2, the extraction efficiency of polarity pesticides (log Kow < 3.5) presents opposite trends, that is, M1 recoveries increase with the increase of log Kow and M2 recoveries decrease with the increase of log Kow. M1 recoveries begin at pesticides with log Kow > 3.5 and gradually increase until approximating those of M2. Owing to the identical extraction procedures of M1 and M2, such deviations are traced to M1’s adoption of hexane liquid–liquid partitioning for cleanup, which lowers the recoveries of polarity pesticides. Average recoveries for 14 pesticides of strongest polarity were about one-half that of M2. M2’s adoption of the SPE cleanup method, identical to M3, has shown balanced and good recoveries for pesticides of different polarities. Comparing M2 with M3, their trend lines cross at the point of log Kow = 2.6, that is, for 74 polarity pesticides (log Kow < 2.6), M2’s recoveries were obviously higher than M3’s. Beginning at log Kow > 2.6, M2’s recoveries gradually decrease and are lower than M3’s for 255 pesticides of medium and weak polarities. With similar cleanup procedures for M2 and M3, in comparison with M3’s pure acetonitrile extraction, M2’s hydration is capable of increasing the extraction efficiency of compounds of strong polarities while decreasing the extraction efficiency of compounds of weak polarities. Comparing M1 with M3, as well as M1’s trend line overall under the M3 trend line, M1 has lower recoveries than M3 for the absolute majority of pesticides. Therefore, we conclude that hydration increases the extraction efficiency of polarity pesticides but, in the meantime, decreases the extraction efficiency of pesticides of weak polarity and nonpolarity—owing to the hexane partitioning design of the M1 procedures. Moreover, the adoption of hexane liquid–liquid partitioning for cleanup causes lower recovery of polarity pesticides. Therefore, M1’s extraction and cleanup are self-contradictory, which results in an overall extraction efficiency lower than M3’s. The aforementioned analysis shows that M3’s pure acetonitrile extraction achieves relatively good results for pesticides of different polarities, while M1’s hydration increases the extraction efficiency of certain polarity water-soluble pesticides. However, the M1 adoption of hexane liquid–liquid partitioning causes certain loss of extracted polarity pesticides. Overall speaking, M3’s extraction efficiency is superior to M1’s.

3.1.3.2.3  Correlation Comparison of Determination Results of Pesticide Aged Samples With Pesticide log Kow Values The correlation of extraction efficiency of three preparation methods and pesticide polarities regarding aged samples are shown in Table 3.1.4. Table 3.1.4 indicates that there are 23 out of 26 water-soluble pesticides with log Kow values less than 1.8, with the M2 determination concentration higher than M3, accounting for 92.3%; there are 28 fat-soluble pesticides with log Kow values higher than 6.0, each with the M3 determination concentration higher than M2. A typical example for hydration decreasing the extraction efficiency of pesticides of nonpolarity is demonstrated in Table 3.1.5, which compares two kinds of specially composed pesticides DDT and long-lasting organic pollutants (PCB) fortification recoveries and aged samples determination results. Due to the very weak polarity of these two kinds of compounds, the content tested with M2 is far lower than with M3. It has also been found in the experimental process that while carrying out hexane liquid–liquid partitioning with M1, acetonitrile extraction solution, hexane, and 5% NaCl water solutions (1:1:5,v/v) are statically layered after manual oscillation, and there is an emulsion layer between the upper layer of hexane phase and the lower layer of water phase, with their content varying as the determined lots and categories of tea vary. After high-speed centrifuging at 10,000 rpm, this emulsified layer is usually compressed a black oily drop existing between the upper and lower phases. Research demonstrates that when the fat content in the samples exceeds 1% (tea contains 2% phospholipids, glycolipids, sulfatide, triglyceride, etc.), nonpolar pesticides will enter into the third phase, in addition to water and organic phases: an emulsified layer, leading to low recoveries, which is in conformity with the low recoveries

504 3  Study on the Influences of Tea Hydration

TABLE 3.1.4 Extraction Comparison of M2 and M3 for Pesticides of Strong Polarity and Nonpolarity in Aged Samples Aged sample concentrations (µg/kg)

Aged sample concentration (µg/kg)

Log Kow < 1.8

M2

M3

Log Kow

M3/ M2

Log Kow > 6.0

M2

M3

Log Kow

M3/ M2

1.

Sulfallate

153.3

186.3

−0.77

1.2

o,p′-DDE

263.9

329.0

6.47

1.2

2.

Dimethipin

290.2

116.1

−0.17

0.4

cis-Chlordane

186.3

390.4

6.10

2.1

3.

Dicrotophos

224.8

93.5

0.00

0.4

Aldrin

277.5

364.7

6.41

1.3

4.

Oxadixyl

344.2

160.7

0.73

0.5

Bioresmethrin

321.8

423.3

6.14

1.3

5.

2,6-Dichlorodenzamide

941.0

575.7

0.77

0.6

DE-PCB 118

226.5

378.4

6.57

1.7

6.

Phosphamidon-2

290.6

135.4

0.79

0.5

Bromophos-ethyl

286.8

341.7

6.15

1.2

7.

Phosphamidon-1

320.0

162.9

0.79

0.5

Flucythrinate

363.7

494.7

6.20

1.4

8.

Metamitron

225.7

218.5

0.98

1.0

Fenvalerate

483.0

1106.4

6.20

2.3

9.

Demeton-S-methyl

21.1

3.8

1.02

0.2

DE-PCB 101

252.3

355.5

6.16

1.4

10.

Mephosfolan

380.5

253.6

1.04

0.7

alpha-Cypermethrin

842.6

890.9

6.94

1.1

11.

Phthalimide

418.3

157.0

1.15

0.4

Flufenoxuron

242.1

432.8

6.16

1.8

12.

Desisopropyl-Atrazine

183.1

108.5

1.15

0.6

Heptachlor

477.4

735.1

6.66

1.5

13.

Paraoxon-methyl

2417.7

2842.6

1.33

1.2

Cypermethrin

520.1

1215.1

6.00

2.3

14.

Hexazinone

329.5

187.8

1.36

0.6

p,p′-DDD

387.8

683.9

6.02

1.8

15.

Formothion

1079.2

424.6

1.48

0.4

DE-PCB 153

198.5

357.0

6.80

1.8

16.

Atrazine-desethyl

212.4

147.9

1.51

0.7

Lambda-Cyhalothrin

441.3

691.9

7.00

1.6

17.

Propoxur-1

342.6

181.4

1.52

0.5

Bifenthrin

236.7

378.7

6.00

1.6

18.

Propoxur-2

440.4

280.6

1.52

0.6

Esfenvalerate

491.9

930.7

6.22

1.9

19.

Dimethyl Phthalate

387.9

250.1

1.56

0.6

DE-PCB 180

156.6

367.3

6.89

2.3

20.

Thionazin

453.3

306.9

1.58

0.7

o,p′-DDT

572.1

1578.9

6.53

2.8

21.

Metalaxyl

369.4

193.7

1.59

0.5

o,p′-DDD

182.3

389.3

6.42

2.1

22.

Pyroquilon

326.7

166.7

1.60

0.5

Chlorfluazuron

138.0

258.8

6.63

1.9

23.

Metribuzin

552.1

401.5

1.70

0.7

Octachlorostyrene

223.0

321.1

6.29

1.4

24.

Bendiocarb

357.7

280.4

1.70

0.8

Etofenprox

232.6

368.1

7.05

1.6

25.

Methabenzthiazuron

344.1

264.5

1.77

0.8

p,p′-DDE

133.4

346.9

6.96

2.6

26.

Tebuthiuron

376.7

195.7

1.79

0.5

p,p′-DDT

572.1

1578.9

6.36

2.8

27.

Permethrin

237.4

381.9

6.50

1.6

28.

Pyridaben

245.6

396.5

6.37

1.6

Silafluofen

196.3

396.0

8.20

2.0

No.

29. M3/M2 < 1

24

M3/M2 > 1

29

Accounting for

92.3%

Accounting for

100.0%

of these two kinds of specially composed DDT and PCB in the fortified recovery experiment. It illustrates that the watersoluble matrix impurities introduced by hydration interfere withe method efficiency.

3.1.3.3  General Analysis of Method Applicability 3.1.3.3.1  Comparison of the Sensitivity of the Three Methods S/N test results for 456 pesticides in the tea matrices of three fortification levels by three methods are shown in Supplemental Tables 4–6. Average S/N values of these 456 pesticides are listed in Table 3.1.6.

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TABLE 3.1.5 Hydration Decreases the Extraction Efficiency of DDT and PCB Fortified recoveries/0.1 mg/kg

Aged sample determination values (µg/kg)

Compounds

log Kow

M1

M2

M3

M3/M1

M1

M2

M3

M3/M1

DDT

p,p′-DDD

6.02

76.6

61.5

87.4

1.1

552.0

387.8

683.9

1.2

p,p′-DDT

6.36

70.0

51.7

78.5

1.1

1181.5

572.1

1578.9

1.3

o,p′-DDD

6.42

73.0

50.0

87.0

1.2

351.5

182.3

389.3

1.1

o,p′-DDE

6.47

69.8

64.5

86.8

1.2

320.5

263.9

329.0

1.0

o,p′-DDT

6.53

71.3

56.3

88.7

1.2

1176.3

572.1

1578.9

1.3

p,p′-DDE

6.96

68.3

45.6

87.6

1.3

278.7

133.4

346.9

1.2

DE-PCB 28

5.71

73.9

76.0

94.9

1.3

330.6

316.1

381.6

1.2

DE-PCB 52

5.79

74.1

71.9

94.7

1.3

314.3

311.4

371.7

1.2

DE-PCB 31

5.81

73.9

73.0

94.9

1.3

330.6

316.1

381.6

1.2

DE-PCB 101

6.16

66.4

66.8

94.0

1.4

263.4

252.3

355.5

1.3

DE-PCB 118

6.57

63.1

54.3

94.7

1.5

242.0

226.5

378.4

1.6

DE-PCB 153

6.80

57.9

64.6

92.8

1.6

206.9

198.5

357.0

1.7

DE-PCB180

6.89

53.0

46.3

92.7

1.7

172.1

156.6

367.3

2.1

PCB

Regarding aged samples, there are 379 pesticides that can be determined by all three sample preparation methods. S/N ratios in these 379 pesticides in tea matrices are shown in Supplemental Table 7. The average S/N for these 379 pesticides, taken with M1, M2, and M3, respectively, are sequentially 949, 6096, and 7027. The comprehensive analysis of the aforementioned results shows that S/N ratios obtained from the three sample preparation methods whether for 456 pesticides in the fortification samples or for 379 pesticides in the aged samples are S/N M3 > S/N M2 > S/N M1. Because of transferring of only 1 mL of acetonitrile extraction solution for cleanup with M1, pesticides of low sensitivity cannot be detected or noises incurred by impurities in the matrices covered, causing decreasing method sensitivity. Figs. 3.1.4 and 3.1.5 show extraction ion chromatogram for GC–MS/MS determination of fluoroglycofen-ethyl and oxadixyl, respectively, with adoption of M1 and M3 for sample preparation. In comparison with M3, M1 is low in detection responses, has relatively large deviations for ion abundance ratio, poor peaks, and more interfering peaks close by. In conclusion, from what is described earlier, despite transferring only 1 mL extraction solution for cleanup with M1 and taking the overall quantity (about 8 mL) for cleanup with M3, the concentrations of tea matrix that are finally injected into the instrument with M1 are only one-eighth that of M3, while the average S/N ratios with M1 in aged samples are only one-seventh of M3. This also demonstrates that hydration has increased the interfering matters of coextraction greatly, seriously affecting the method sensitivity.

3.1.3.3.2  Comparison of the Extraction Efficiencies of the Three Methods Aged Tea Samples The single-point quantification method is adopted for determination of the content of 456 pesticides in aged tea samples, with results tabulated in Supplemental Table 7. The number of pesticides detected with the three sample preparation methods respectively are 382 with M1, 404 with M2, and 410 with M3. The content ratio is calculated for the same pesticide

TABLE 3.1.6 Comparison of Average S/N Values From Three Pretreatment Methods and Three Fortification Concentrations Fortification levels/mg/kg

M1

M2

M3

1.00

6234

11127

14224

0.10

432

1656

3048

0.01

73

240

229

506 3  Study on the Influences of Tea Hydration

FIGURE 3.1.4  Fluoroglycofen-ethyl with respective M1 and M3 sample preparation and GC–MS/MS determination.

FIGURE 3.1.5  Oxadixyl with respective M1 and M3 sample preparation and GC–MS/MS determination.

with M3 and M1 to obtain the content ratio of M3/M1, and with M3 and M2 to obtain the content ratio of M3/M2, with the pesticide number statistically computed in the four ratio sectors listed in Table 3.1.7. M3/M1 in Table 3.1.7 shows that there are 201 pesticides with content ratio 0.8–1.2, accounting for 53% in which pesticides can be detected overall by M1 and M3. There are 116 pesticides with ratios greater than 1.2, many more than the 65 pesticides with ratios less than 0.8. It is evident that the extraction efficiency for aged samples with M3 is higher than with M1. M3/M2 shows that there are 176 pesticides with a content ratio 0.8–1.2, accounting for 46% overall in which pesticides can be detected by M2 and M3, and there are 137 pesticides with ratios greater than 1.2, many more than the 87 pesticides with ratios less than 0.8. Thus, it reveals that the extraction efficiency for aged samples with M3 is higher than with M2. In addition, compared with M3, there are 28 pesticides that fail detection with M1; compared with M2, there are 25 pesticides that fail detection with M1. To follow through on what was discussed earlier, nonhydration with M3 is superior

TABLE 3.1.7 Distribution of Pesticide Content Ratios Determined by the Three Sample Preparation Methods for Aged Samples Ratiosector

≤0.6

0.6–0.8

0.8–1.2

≥1.2

The average ratio of overall pesticides that can be determined

M3/M1 pesticide number

13

52

201

116

1.17

M3/M2 pesticide number

36

51

176

137

1.09

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507

TABLE 3.1.8 Distribution of Pesticide Content Ratios Determined by the Three Sample Preparation Methods for Incurred Tea Samples Ratio range

≤0.6

0.6–0.8

0.8–1.2

≥1.2

Average ratios of 18 pesticides

M3/M1 pesticide number

0

4

10

4

1.01

M3/M2 pesticide number

0

5

7

6

1.09

to hydration with M1 and M2, no matter the extraction efficiency of pesticides or for the application ranges of pesticides. That is, M3 is high in extraction efficiency for aged samples and shows a larger range of pesticides that are applicable. Incurred Tea Samples The content of 18 pesticides for determination in incurred tea samples is tabulated in Supplemental Table 9, and the pesticide content ratios determined by the three sample preparation methods are respectively computed, with the pesticide numbers of M3/M1 and M3/M2 in the four ratio sectors listed, in Table 3.1.8. Table 3.1.8 shows that M3/M1 has found 10 out of 18 pesticides with content ratios 0.8–1.2, 4 pesticides with ratios greater than 1.2 and less than 0.8, and no pesticides with ratios less than 0.6. The average ratio of the 18 pesticides is 1.01. Therefore, it can be said that for the majority of pesticides the extraction efficiencies of these two methods are identical, with certain differences among specific pesticides. M3/M2 has found that there are 7 out of 18 pesticides with content ratios 0.8–1.2, and there are 6 and 5 pesticides, respectively, with ratios greater than 1.2 and less than 0.8, and no pesticides with atios less than 0.6. The average ratio of the 18 pesticides is 1.09. There exist certain differences when considering the extraction efficiencies of various pesticides with M3 and M2, but the overall extraction efficiencies are basically identical. Based on what was described earlier, the overall extraction efficiencies are approximately the same although there exist certain differences in extraction efficiencies for different pesticides with incurred samples when comparing the hydration of M1 and M2 with the nonhydration of M3. There are only 18 pesticides for the appraisal of incurred samples. Therefore, it is still beyond our capacity at present to judge which of these three methods is good or bad for the other hundreds of pesticides. This is an issue that needs further study. Comparison of Fortified Recoveries for the Three Sample Preparation Methods Under “Uniform Limit Concentrations” Fortified recovery experimental results from 456 pesticides by the three preparation methods are listed in Supplemental Tables 1–3; the fortified recovery experiment results from testing 160 pesticides by the two methods are tabulated in Supplemental Table 8. Also, pesticide ratios for 10 µg/kg fortification levels and at the same time conforming to recoveries 70%–120% and RSD < 20% are listed in Table 3.1.9. In the fortification experiment of 456 pesticides, pesticides that conform to EU SANCO/12495/2011 technical requirements with M3 account for 50.2%, M2 45.0%, but withM1 only 5%.The method applicability is M3 > M2 > M1: in the fortification experiment of 160 pesticides, M3 is also superior to M1. Comparison of RSD for Pesticides in Aged Tea Samples Determined by the Three Sample Preparation Methods The analytical results from the aged oolong tea samples sprayed with 456 pesticides are shown in Supplemental Table 7, and the RSD data for the pesticides discovered are now divided into four sectors, with distribution shown in Table 3.1.10.

TABLE 3.1.9 Pesticide Ratios for 0.010 mg/kg “Uniform Limit Concentrations” and Conforming to Rec70%–120% and RSD < 20% M1

M2

M3

Pesticide varieties

Pesticide number

Ratios/%

Pesticide number

Ratios/%

Pesticide number

Ratios/%

456

23

5.0

205

45.0

229

50.2

160

17

10.6

50

31.2

508 3  Study on the Influences of Tea Hydration

TABLE 3.1.10 Pesticide Number Discovered in Aged Samples and Distributions and Ratios (%) of 4 RSD Sectors RSD%

M1

M2

M3

<10

158(41%)

112(28%)

381(93%)

10–15

136(36%)

209(52%)

14(3%)

15–20

58(15%)

56(14%)

7(2%)

>20

30(8%)

27(7%)

8(2%)

Total

382

404

410

From the aged samples sprayed with 456 pesticides, the pesticide number actually detected is, respectively, 382 with M1, 404 with M2, and 410 with M3. It can seen that M3 > M2 > M1 in terms of sample preparation efficacy. In terms of method ruggedness, there are 381 pesticides with RSD < 10% with M3, accounting for 93%, much more than the ratios with M1 and M2—41% and 28%, respectively. There are 30 and 27 pesticides with pesticide number RSD > 20%, respectively, for M1 and M2—much more than M3 with 8 pesticides. Therefore, when considering precision, M3 is better than M1 and M2 in applicability.

3.1.3.4  Comparison of the Cleanup Efficiency for the Three Methods S/N results obtained from GC–MS/MS determination of 456 pesticide aged samples using the three sample preparation methods are shown in Supplemental Tables 4–6, with their added average S/N values for M1, M2, and M3, respectively, of 942, 5753, and 6781. It is evident that M3 method sensitivity is superior to M1. The color of interfering matters left on the SPE columns can testify this point as proof. See Fig. 3.1.6. In addition, the oolong tea blank matrix extraction solutions prepared by the three methods are fully scanned for their total ion chromatogram (Scan-TIC) (Fig. 3.1.7), among which the contents of M1 (black), M2 (red), and M3(green) are all 0.2 g/mL tea. Fig. 3.1.7 also shows that there are lower baselines and fewer interfering peaks in TIC of M3 than those in TIC of M1 and M2, which also proves that M3 acetonitrile extraction with SPE cleanup sample preparation techniques are superior to hexane liquid–liquid portioning technique with tea hydration extraction. Based on the discussion of the analytical results from the three sample preparation methods, the comprehensive statistical analysis indexes are shown in Tables 3.1.11 and Table 3.1.12.

3.1.4 CONCLUSIONS This experiment has adopted three sample preparation methods: M1, tea hydration + acetonitirle oscillating extraction + hexane liquid/liquid partitioning cleanup for part of extraction solution; M2, tea hydration + acetonitirle oscillating extraction + SPE cleanup; M3, pure acetonitrile homogeneous extraction + SPE cleanup—for a GC–MS/MS determination of 456 pesticide fortified samples, 456 pesticide aged samples, and 18 pesticide field-incurred samples. In this research of 456 pesticides, the log Kow values of 329 pesticides have been determined. Taking the pesticide recoveries as the

FIGURE 3.1.6  Sketch of Cleanert-TPT SPE cleanup effects with samples prepared by the three methods.

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Chapter | 3.1

509

FIGURE 3.1.7  Scan-TIC of blank tea matrix prepared by the three methods.

TABLE 3.1.11 Comparison of the Analytical Results by GC–MS/MS in Four Kinds of Samples Prepared With M1, M2 and M3 Method

Ratio of REC 70%–120%

Ratio of RSD < 20%

Ratio of REC 70%–120% and RSD < 20%

S/N

1.0 mg/ 0.1 mg/ 0.01 mg/ 1.0 mg/ 0.1 mg/ 0.01 mg/ 1.0 mg/ 0.1 mg/ 0.01 mg/ 1.0 mg/ kg kg kg kg kg kg kg kg kg kg

0.1 mg/ 0.01 mg/ kg kg

(1) The fortified recoveries of 456 pesticides M1

73.5

68.6

26.5

84.9

62.9

34.2

63.8

45.8

5.0

6234

432

73

M2

77.4

78.5

76.8

68.6

75.7

62.9

53.8

59.2

45.0

11127

1656

240

M3

88.6

91.4

71.3

87.5

90.6

66.4

80.9

83.8

50.2

14224

3048

229

(2) The fortified recoveries of 159 pesticides M1

75.5

67.9

14.5

99.4

89.9

15.1

75.6

76.3

10.6

1234

145

M3

90.6

75.5

49.7

100.0

97.5

69.8

90.6

80.0

33.1

36410

2459

(4) The analytical results of 456 pesticides in incurred samples (n = 4)

(3) The analytical results of 456 pesticides in aged samples (n = 5)

Method No.s of peaking pesticides

S/N

RSD

the ratios of content

M1

382

942

12.2

≤0.8

M2

404

5753

12.8

0.8–1.2

M3

410

6781

4.1

≥1.2

Pesticide No.s

M3/M1 M3/M2 The ratios of content 65

87

≤0.8

201

176

0.8–1.2

116

137

≥1.2

M3/M1 M3/M2

Pesticide 6 No.s 8

6

4

4

8

510 3  Study on the Influences of Tea Hydration

TABLE 3.1.12 Technical Indexes for M1, M2 and M3 No.

Technical indexes

Sources

M1

M2

M3

Conclusions

1.

Percentage of pesticides with 70%– 120% recoveries

0.01 mg/kg Fortified recovery experiment

26.54%

76.75%

71.27%

M2 > M3 > M1

2.

Percentage of pesticides with RSD < 20%

34.21%

62.94%

66.45%

M3 > M2 > M1

3.

LOD, µg/kg

3.2

1.9

1.5

M3 < M2 < M1

4.

LOQ, µg/kg

10.6

6.2

5.0

M3 < M2 < M1

5.

S/N

Aged sample testing

942

5753

6781

M3 > M2 > M1

6.

Peak height

Aged sample testing

7.6E+03

1.4E+05

1.2E+05

M2 > M3 > M1

7.

Correlation of recoveries VS pesticide polarities

log Kow↑, recovery: firstly ↑finally ↓

log Kow↑, recovery↓

log Kow↑, recovery stale and balanced

(1) Hydration and hexane LLE affect pesticide recoveries; (2) acetonitrile homogeneous and SPE have a sufficient and balanced extraction for pesticides of different polarities

8.

Calibration linear coefficient (R2)

0.9765

0.9995

0.9987

M2 > M3 > M1

9.

Extraction results

Low recoveries, and results tend to be falsely higher for aged sample and incurred sample determinations

Relatively high recoveries, and have good results in extracting watersoluble pesticides of polarities, with extraction matrices of high content of foreign materials

Recoveries fall within the scope of good practices, and have a stale and well-balanced extraction of pesticides

M3 > M2 > M1

S/N = 73

S/N = 7240

S/N = 7229

M3 > M2 > M1

10.

Cleanup results

Incurred sample testing

0.01mg/kg Fortified recovery test Incurred sample testing

Regarding color of the pesticide extraction solution, (1) darkest with M2, M1 next to it, and relatively transparent and clear with M3; (2) with M1 and M2 the extraction solution will have flocculent precipitate after being stored away from exposure to light for a certain amount of time

Oolong tea matrix Scan-TIC

High baselines

Relatively high baselines

Low baselines

Aged samples testing

δRT = 0.009

δRT = 0.004

δRT = 0.004

M3 < M2 < M1

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511

TABLE 3.1.12 Technical Indexes for M1, M2 and M3 (cont.) No.

Technical indexes

Sources

M1

M2

M3

Conclusions

11.

Method fitness (number of peaking pestitcides)

Aged samples testing

382

404

410

M3 > M2 > M1

12.

Method selectivity/ special effect

Aged samples testing

Poor extraction efficiency for pesticides within the range of log Kow < 3.5 and log Kow > 5.4

Poor extraction efficiency for pesticides within the range of log Kow > 3.5

Relatively good extraction efficiency for pesticides within the range of log Kow (−0.77 to 8.20)

Have sufficient extraction for all pesticides with different polarities with M3

longitudinal axis and the log Kow as horizontal axis to establish a mathematical regression equation, these findings are obtained: the extraction efficiency of hydration method M1 presents obvious correlation with pesticide log Kow values and hydration increases extraction efficiencies of certain polarity pesticides, such as the 24 pesticides of strong polarity, but in addition, this has decreased the extraction efficiencies of certain pesticides of weak polarity, such as the 29 pesticides of nonpolarity. The M3 nonhydration method has almost nothing to do with the polarities of pesticides in extraction efficiencies, presenting a straight line nearly parallel to the log Kow horizontal axis, with its extraction efficiencies balanced and sufficient for pesticides of different polarities and with a wider range for application. The statistics of the log Kow values of 329 pesticides also find that there are 14 pesticides with log Kow value <1.0, 25 with log Kow value 1.0–2.0, 65 with log Kow value 2.0–3.0, 95 with log Kow value 3.0–4.0, 72 with log Kow value 4.0–5.0, 27 with log Kow value 5.0–6.0, and 31 with log Kow value >6.0. If pesticides with log Kow values <1.0 are considered to be pesticides of strong polarity and those of 1.0–2.0 polarity pesticides, these two items total 39 pesticides, accounting for 12%. Other medium polarity, weak polarity, and nonpolarity pesticides total 290, accounting for 88%. The ratio of polarity pesticides is far smaller than that of nonpolarity pesticides, proving that hydration does more harm than good, which is the first point. Second, interfering matters from coextraction after tea hydration have increased, leading to an obvious drop in method efficiency. Only part of the extraction solution (1 mL) is transferred for cleanup with M1, which is equivalent to diluting the sample matrix 10 times—while the entire quantity is taken for cleanup with M3, and the target matter concentration is about 9 times larger than that with M1. S/N with M3 is 6781, 940 with M1, and the sensitivity of M3 is 7 times greater than that of M1. The fortification experiment of 0.010 mg/kg (uniform limit) demonstrates that there are 23 pesticides with hydration method M1 that meet the EU SANCO/12495/2011 technical requirements of recoveries 70%–120% and conform to RSD ≤20%, accounting for 5.0%, while there are 229 pesticides with nonhydration M3, accounting for 50%, from which it is evident that M1 has failed to meet the good practice requirement of the EU. For aged samples with residual concentrations above 380 µg/kg, there are 158 pesticides that meet RSD ≤ 10% with M1, accounting for 35%, but 381 with M3, making up 84%, testifying that M3 is 2.4 times superior to M1. In addition, the experiment also found that there are two technical defects with M1’s design: on the one hand, tea hydration increases the extraction efficiencies of polarity pesticides, but on the other hand, hexane liquid–liquid partitioning is adopted for its cleanup, which increases the loss of polarity pesticides extracted in the hexane partitioning. Both M1 and M2 use hydration, but the cleanup adopted by M2 is identical to M3, and M2’s recoveries for polarity pesticides are higher than those with M3, which is a good proof of M1’s defect. Therefore, M1 equipped with hexane liquid–liquid partitioning is self-contradictory; only 1 mL extraction solution transferred with M1 is considered to be equivalent to 0.2 g sample, but the fact is that 10 mL acetonitile extraction solution is finally left to 8.0–8.5 mL, so 1 mL extraction solution taken is approximately equivalent to 2.35 g and computation results with M1 are about 20% falsely higher, which should be deducted from its results. For M1, the extraction solutions left after hydration extraction of different teas also change in volume. If taking part of it for cleanup, the variability is great, but if taking the overall, it is difficult for cleanup, so the variability with M1 is far greater than that with M3. Owing to the reasons stated in the aforementioned three aspects, using tea hydration to increase extraction efficiencies of pesticides has more disadvantages than advantages for the determination of hundreds of multiclasses and multikinds of residue pesticides in tea, which proves it to be unworthy and unadvisable.

512 3  Study on the Influences of Tea Hydration

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