Sludge, treated biowaste and soil - Determination of linear alkylbenzene sulfonates (LAS) by high-performance liquid chromatography (HPLC) with fluorescence detection (FLD) or mass selective detection (MS)

This Technical Specification specifies a method for the determination of linear alkylbenzene sulfonate (LAS) in sludge, treated biowaste and soil using high-performance liquid chromatography (HPLC) with a fluorescence detector (FLD) or a mass selective detector (MS).
This Technical Specification specifies the determination of the sum of LAS. Under the conditions specified in this Technical Specification, typically a limit of detection of 20 mg/kg (expressed as dry matter) for sludge and of 0,2 mg/kg to 0,5 mg/kg for soil and treated biowaste may be achieved.
Lower limits of detection may be achieved by concentrating the extract by solvent evaporation.
NOTE   The single LAS homologues C10 to C14 can be determined by this Technical Specification.

Schlamm, behandelter Bioabfall und Boden - Bestimmung von linearen Alkylbenzolsulfonaten (LAS) mittels Hochleistungs-Flüssigkeitschromatographie (HPLC) mit Fluoreszenzdetektion (FLD) oder massenselektiver Detektion (MS)

Diese Technische Spezifikation legt ein Verfahren zur Bestimmung der linearen Alkylbenzolsulfonate (LAS) in Schlamm, behandeltem Bioabfall und Boden mittels Hochleistungs Flüssigchromatographie (HPLC) und Detektion durch Fluoreszenz (FLD) oder Massenspektrometrie (MS) fest.
Diese Technische Spezifikation beschreibt die Bestimmung der Summe aller LAS. Unter den in dieser Technischen Spezifikation festgelegten Bedingungen kann in der Regel eine Nachweisgrenze von 20 mg/kg (angegeben als Trockenmasse) für Schlamm und eine Nachweisgrenze von 0,2 mg/kg bis 0,5 mg/kg für Boden und behandelten Bioabfall erreicht werden.
Niedrigere Nachweisgrenzen können erreicht werden, wenn der Extrakt durch Verdampfen des Lösemittels konzentriert wird.
ANMERKUNG   Nach dieser Technischen Spezifikation können die LAS Homologen C10 bis C14 einzeln bestimmt werden.

Boues, biodéchets traités et sols - Détermination des alkylbenzènesulfonates linéaires (LAS) par chromatographie liquide à haute performance (CLHP) avec détection par fluorescence (FLD) ou détection sélective de masse (SM)

La présente Spécification technique spécifie une méthode pour la détermination des alkylbenzènesulfonates linéaires (LAS) dans les boues, les biodéchets traités et les sols par chromatographie liquide à haute performance (CLHP) avec détection par fluorescence (FLD) ou détection par spectrométrie de masse (SM).
La présente Spécification technique spécifie la détermination de la somme des alkylbenzènesulfonates linéaires (LAS). Dans les conditions définies dans la présente Spécification technique, on peut généralement atteindre une limite de détection de 20 mg/kg (exprimée par rapport à la matière sèche) pour les boues et de 0,2 mg/kg à 0,5 mg/kg pour les sols et les biodéchets traités.
Il est possible d'atteindre des limites de détection inférieures en augmentant la concentration de l'extrait par évaporation de solvant.
NOTE   La présente Spécification technique permet de déterminer les homologues individuels C10 à C14 des alkylbenzènesulfonates linéaires (LAS).

Blato, obdelani biološki odpadki in tla - Določevanje linearnih alkilbenzen sulfonatov (LAS) z uporabo tekočinske kromatografije visoke ločljivosti (HPLC) s fluorescenčno detekcijo (FLD) ali masno selektivno detekcijo (MS)

Ta tehnična specifikacija opredeljuje metodo za določevanje linearnih alkilbenzen sulfonatov (LAS) v blatu, obdelanih bioloških odpadkih in tleh z uporabo tekočinske kromatografije visoke ločljivosti (HPLC) s fluorescenčno detekcijo (FLD) ali masno selektivno detekcijo (MS). Ta tehnična specifikacija opredeljuje določevanje vsote linearnih alkilbenzen sulfonatov. Pod pogoji iz te tehnične specifikacije se običajno lahko dosežejo meje detekcije 20 mg/kg (izraženo kot suha snov) za blato in od 0,2 mg/kg do 0,5 mg/kg za tla in obdelane biološke odpadke. Nižje meje detekcije se lahko dosežejo s koncentracijo izvlečka z izhlapevanjem raztopine.

General Information

Status
Published
Publication Date
21-Feb-2012
Current Stage
9060 - Closure of 2 Year Review Enquiry - Review Enquiry
Due Date
02-Dec-2020
Completion Date
02-Dec-2020

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SLOVENSKI STANDARD
SIST-TS CEN/TS 16189:2012
01-junij-2012
%ODWRREGHODQLELRORãNLRGSDGNLLQWOD'RORþHYDQMHOLQHDUQLKDONLOEHQ]HQ
VXOIRQDWRY /$6 ]XSRUDERWHNRþLQVNHNURPDWRJUDILMHYLVRNHORþOMLYRVWL +3/& V
IOXRUHVFHQþQRGHWHNFLMR )/' DOLPDVQRVHOHNWLYQRGHWHNFLMR 06

Sludge, treated biowaste and soil - Determination of linear alkylbenzene sulfonates

(LAS) by high-performance liquid chromatography (HPLC) with fluorescence detection

(FLD) or mass selective detection (MS)
Schlamm, behandelter Bioabfall und Boden - Bestimmung von Linearen

Alkylbenzolsulfonaten (LAS) mittels Hochleistungs-Flüssigkeitschromatographie (HPLC)

mit Fluoroszenzdetektion (FLD) oder massenselektiver Detektion (MS)

Boues, bio-déchets traités et sols - Détermination des alkylbenzènesulfonates linéaires

(LAS) par chromatographie liquide à haute performance (CLHP) avec détection par
fluorescence (FLD) ou détection sélective de masse (SM)
Ta slovenski standard je istoveten z: CEN/TS 16189:2012
ICS:
13.030.20 7HNRþLRGSDGNL%ODWR Liquid wastes. Sludge
13.080.10 .HPLMVNH]QDþLOQRVWLWDO Chemical characteristics of
soils
SIST-TS CEN/TS 16189:2012 en,fr,de

2003-01.Slovenski inštitut za standardizacijo. Razmnoževanje celote ali delov tega standarda ni dovoljeno.

---------------------- Page: 1 ----------------------
SIST-TS CEN/TS 16189:2012
---------------------- Page: 2 ----------------------
SIST-TS CEN/TS 16189:2012
TECHNICAL SPECIFICATION
CEN/TS 16189
SPÉCIFICATION TECHNIQUE
TECHNISCHE SPEZIFIKATION
February 2012
ICS 13.030.01
English Version
Sludge, treated biowaste and soil - Determination of linear
alkylbenzene sulfonates (LAS) by high-performance liquid
chromatography (HPLC) with fluorescence detection (FLD) or
mass selective detection (MS)
Boues, biodéchets traités et sols - Détermination des
Schlamm, behandelter Bioabfall und Boden - Bestimmung

alkylbenzènesulfonates linéaires (LAS) par von linearen Alkylbenzolsulfonaten (LAS) mittels

chromatographie liquide à haute performance (CLHP) avec Hochleistungs-Flüssigkeitschromatographie (HPLC) mit

détection par fluorescence (FLD) ou détection sélective de Fluoreszenzdetektion (FLD) oder massenselektiver

masse (SM) Detektion (MS)

This Technical Specification (CEN/TS) was approved by CEN on 24 April 2011 for provisional application.

The period of validity of this CEN/TS is limited initially to three years. After two years the members of CEN will be requested to submit their

comments, particularly on the question whether the CEN/TS can be converted into a European Standard.

CEN members are required to announce the existence of this CEN/TS in the same way as for an EN and to make the CEN/TS available

promptly at national level in an appropriate form. It is permissible to keep conflicting national standards in force (in parallel to the CEN/TS)

until the final decision about the possible conversion of the CEN/TS into an EN is reached.

CEN members are the national standards bodies of Austria, Belgium, Bulgaria, Croatia, Cyprus, Czech Republic, Denmark, Estonia,

Finland, France, Germany, Greece, Hungary, Iceland, Ireland, Italy, Latvia, Lithuania, Luxembourg, Malta, Netherlands, Norway, Poland,

Portugal, Romania, Slovakia, Slovenia, Spain, Sweden, Switzerland, Turkey and United Kingdom.

EUROPEAN COMMITTEE FOR STANDARDIZATION
COMITÉ EUROPÉEN DE NORMALISATION
EUROPÄISCHES KOMITEE FÜR NORMUNG
Management Centre: Avenue Marnix 17, B-1000 Brussels

© 2012 CEN All rights of exploitation in any form and by any means reserved Ref. No. CEN/TS 16189:2012: E

worldwide for CEN national Members.
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CEN/TS 16189:2012 (E)
Contents Page

Foreword ..............................................................................................................................................................3

Introduction .........................................................................................................................................................4

1 Scope ......................................................................................................................................................5

2 Normative references ............................................................................................................................5

3 Terms and definitions ...........................................................................................................................5

4 Principle ..................................................................................................................................................5

5 Interferences ..........................................................................................................................................6

5.1 Interferences from sampling ................................................................................................................6

5.2 Interferences by HPLC-FLD and HPLC-MS .........................................................................................6

6 Reagents .................................................................................................................................................6

7 Apparatus ...............................................................................................................................................9

8 Sample storage and sample pretreatment ....................................................................................... 10

8.1 Sample storage ................................................................................................................................... 10

8.2 Sample pretreatment .......................................................................................................................... 10

9 Procedure ............................................................................................................................................ 10

9.1 Extraction ............................................................................................................................................ 10

9.1.1 Extraction of dried sludge ................................................................................................................. 10

9.1.2 Extraction of dried soil, sediment and treated biowaste ................................................................ 10

9.2 Concentration (optional) .................................................................................................................... 11

9.3 Clean-up (optional) ............................................................................................................................. 11

9.4 Blank test ............................................................................................................................................. 11

9.5 HPLC analysis ..................................................................................................................................... 11

9.5.1 General ................................................................................................................................................. 11

9.5.2 Fluorescence detection ...................................................................................................................... 12

9.5.3 Mass selective detection .................................................................................................................... 12

9.6 Calibration ........................................................................................................................................... 12

9.6.1 General ................................................................................................................................................. 12

9.6.2 Initial calibration ................................................................................................................................. 12

9.6.3 Verification of calibration ................................................................................................................... 13

9.7 Analysis of samples and identification ............................................................................................ 13

10 Calculation and expression of results .............................................................................................. 13

10.1 General ................................................................................................................................................. 13

10.2 Calibration ........................................................................................................................................... 14

10.3 Calculation ........................................................................................................................................... 14

11 Precision .............................................................................................................................................. 15

12 Test report ........................................................................................................................................... 15

Annex A (informative) Repeatability and reproducibility data ..................................................................... 16

A.1 Materials used in the interlaboratory comparison study ............................................................... 16

A.2 Interlaboratory comparison results .................................................................................................. 17

Annex B (informative) Examples of chromatographic conditions and chromatograms .......................... 18

B.1 Isomeric separation of LAS ............................................................................................................... 18

B.2 Homologue separation of LAS .......................................................................................................... 19

Annex C (informative) Examples of clean-up procedures ........................................................................... 20

C.1 Clean-up procedure based on graphitised carbon black (GCB) columns .................................... 20

C.2 Clean-up procedure based on strong anion exchange (SAX) columns ........................................ 20

Bibliography ..................................................................................................................................................... 21

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Foreword

This document (CEN/TS 16189:2012) has been prepared by Technical Committee CEN/TC 400 “Project

Committee - Horizontal standards in the fields of sludge, biowaste and soil”, the secretariat of which is held by

DIN.

Attention is drawn to the possibility that some of the elements of this document may be the subject of patent

rights. CEN [and/or CENELEC] shall not be held responsible for identifying any or all such patent rights.

The preparation of this document by CEN is based on a mandate by the European Commission

(Mandate M/330), which assigned the development of standards on sampling and analytical methods for

hygienic and biological parameters as well as inorganic and organic determinants, aiming to make these

standards applicable to sludge, treated biowaste and soil as far as this is technically feasible.

According to the CEN/CENELEC Internal Regulations, the national standards organizations of the following

countries are bound to announce this Technical Specification: Austria, Belgium, Bulgaria, Croatia, Cyprus,

Czech Republic, Denmark, Estonia, Finland, France, Germany, Greece, Hungary, Iceland, Ireland, Italy,

Latvia, Lithuania, Luxembourg, Malta, Netherlands, Norway, Poland, Portugal, Romania, Slovakia, Slovenia,

Spain, Sweden, Switzerland, Turkey and the United Kingdom.
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CEN/TS 16189:2012 (E)
Introduction

The anionic surfactant LAS (Linear Alkylbenzene Sulfonate) is found in the environment due to the use of LAS

in detergents. For more than 30 years LAS has been the largest single surfactant used in detergents, and the

use continues on a high level.

Although LAS is readily biodegradable during wastewater treatment, considerable amounts may still be found

in sludge of municipal origin. By the use of sludge for soil improvement LAS may end up in the agricultural

soil, where a rapid biodegradation takes place.

The method describes the determination of LAS in sludge, soil, treated biowaste and neighbouring fields. LAS

is the sodium salt of alkylbenzene sulfonic acids, and it consists of a mixture of the homologues C -LAS,

C -LAS, C -LAS, C -LAS and C -LAS. LAS is determined as the sum of the homologues.

11 12 13 14

This Technical Specification is applicable and validated for several types of matrices as indicated in Table 1

(see also Annex A for the results of the validation).

Table 1 — Matrices for which this Technical Specification is applicable and validated

Matrix Materials used for validation
Sludge Municipal sewage sludge
Biowaste Fresh compost
Soil Sludge amended soil

WARNING — Persons using this Technical Specification should be familiar with usual laboratory

practice. This Technical Specification does not purport to address all of the safety problems, if any,

associated with its use. It is the responsibility of the user to establish appropriate safety and health

practices and to ensure compliance with any national regulatory conditions.

IMPORTANT — It is absolutely essential that tests conducted according to this Technical

Specification be carried out by suitably trained staff.
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CEN/TS 16189:2012 (E)
1 Scope

This Technical Specification specifies a method for the determination of linear alkylbenzene sulfonate (LAS) in

sludge, treated biowaste and soil using high-performance liquid chromatography (HPLC) with a fluorescence

detector (FLD) or a mass selective detector (MS).

This Technical Specification specifies the determination of the sum of LAS. Under the conditions specified in

this Technical Specification, typically a limit of detection of 20 mg/kg (expressed as dry matter) for sludge and

of 0,2 mg/kg to 0,5 mg/kg for soil and treated biowaste may be achieved.

Lower limits of detection may be achieved by concentrating the extract by solvent evaporation.

NOTE The single LAS homologues C to C can be determined by this Technical Specification.

10 14
2 Normative references

The following documents, in whole or in part, are normatively referenced in this document and are

indispensable for its application. For dated references, only the edition cited applies. For undated references,

the latest edition of the referenced document (including any amendments) applies.

EN 15934, Sludge, treated biowaste, soil and waste — Calculation of dry matter fraction after determination of

dry residue or water content
EN 16179, Sludge, treated biowaste and soil — Guidance for sample pretreatment

EN ISO 22892, Soil quality — Guidelines for the identification of target compounds by gas chromatography

and mass spectrometry (ISO 22892)

ISO 8466-1, Water quality — Calibration and evaluation of analytical methods and estimation of performance

characteristics — Part 1: Statistical evaluation of the linear calibration function

3 Terms and definitions
For the purposes of this document, the following terms and definitions apply.
3.1
analyte

mixture of homologues (i. e. C -LAS, C -LAS, C -LAS, C -LAS and C -LAS) where each homologue

10 11 12 13 14

consists of a mixture of four to six isomers depending on the length of the alkyl group

Note 1 to entry: The dominant homologues in detergents and environmental samples are C -LAS and C -LAS. C to

11 12 10
C refers to the chain length of the linear alkyl group.
4 Principle

After pretreatment, the test sample is extracted by shaking with methanol. If necessary, interfering compounds

are removed from the extract by a clean-up on a suitable column.

The extract is analysed by high performance liquid chromatography (HPLC) on a C - or C -column and

8 18
detection by fluorescence (FLD) or mass spectrometry (MS).

The identification is based on the retention times of the homologues and of the isomers of each homologue.

Another identification point is the pattern/fingerprint of the homologues, and the isomer fingerprint of each

homologue if a C -column is used for HPLC. By use of MS detection the relative intensities of two diagnostic

ions may also be used for the identification (optional).
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The quantification is based on an internal standard procedure. The internal standard (C -LAS) is taken

through the whole analytical procedure.

Depending on the type of matrices from which LAS is extracted, different analytical pathways can be applied.

An overview of the analytical procedure for the matrix of interest is shown in Table 2.

5 Interferences
5.1 Interferences from sampling

Use sampling containers of materials (preferably glass or steel) that do not significantly affect the sample

during the contact through sampling and storage. Plastic containers may be used if it has been proven that

they do not significantly affect the sample.
5.2 Interferences by HPLC-FLD and HPLC-MS

The chromatographic analysis can be done on a C - or a C -reverse phase column, and the choice of

8 18

column determines the separation obtained. On the C -column (with methanol in mobile phase) the LAS

homologues are separated, however, there is no separation of the isomers. On the C -column (with

acetonitrile in mobile phase) the homologues are separated and there is a partial separation of the isomers of

each homologue. This is illustrated by the chromatograms in Annex B.

The selectivity of the fluorescence as well as the mass selective detector is high; however, interference from

co-eluting substances may occur. It is essential that the interfering peaks are not included in the calculations.

A peak is excluded if the retention time differs from the LAS standard mixture. Interfering peaks can best be

detected when a C -column is used for the LC analysis, due to the partial separation of the isomers. The

C -column is mandatory when fluorescence is used, due to the higher selectivity obtained. The interfering

peaks can usually be detected by comparing the fingerprints of the sample with the fingerprints of the LAS

standard mixture, although the isomer and homologue distribution in the environmental samples may differ

from the distribution in the standard mixture.

The highest selectivity is obtained by the use of a C -column and the MS detector. However, for most

applications the separation on a C -column is sufficient, when MS is used. When all isomers are eluted in one

peak, the integrations are less complicated, resulting in a higher precision and a lower limit of detection.

Table 2 — Choice of analytical procedure
Matrix FLD MS
C-column C-column C-column C -column
8 18 8 18
Sludge No Yes Yes Yes
Soil No (Yes) Yes Yes
Treated biowaste No (Yes) Yes Yes

For FLD the limit of detection will generally be inadequate for this type of matrix.

6 Reagents
6.1 General
Use only reagents of recognized analytical grade, unless otherwise specified.

The purity of the reagents used shall be checked by running a blank determination as described in 9.4.

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SIST-TS CEN/TS 16189:2012
CEN/TS 16189:2012 (E)
6.2 Methanol, CH OH; HPLC-grade.
6.3 Acetonitrile, C H N; HPLC-grade.
2 3
- +
6.4 Ammonium acetate, c(CH COO NH ) = 0,01 mol/l.
3 4
6.5 Mobile phases for HPLC
6.5.1 For isomeric separation on C -column
 Mobile phase A: 0,01 mol/l ammonium acetate (6.4);
 Mobile phase B: Acetonitrile (6.3).
6.5.2 For homologue separation on C -column
 Mobile phase A: 0,01 mol/l ammonium acetate (6.4);
 Mobile phase B: Methanol (6.2).
6.6 Reagents for clean-up procedures
6.6.1 Clean-up procedure based on strong anion exchange (SAX)
6.6.1.1 SAX column
6.6.1.2 Acetic acid, CH COOH
6.6.1.3 Hydrochloric acid, HCl
6.6.1.4 Methanol, CH OH
6.6.2 Clean-up procedure based on graphitised carbon black (GCB)
6.6.2.1 GCB column
6.6.2.2 Hydrochloric acid, HCl

6.6.2.3 Tetramethylammonium hydroxide, C H NO (CAS-RN 10424-65-4 ); pentahydrate.

4 13
6.6.2.4 Formic acid, HCOOH
6.6.2.5 Dichloromethane, CH Cl
2 2
6.6.2.6 Methanol, CH OH
6.7 Nitrogen, N , for solvent evaporation of sufficient purity.
6.8 Standards for calibration
6.8.1 General
The standards shall be stored in a freezer at a temperature of (–18 ± 3) °C.
CAS-RN Chemical Abstracts Service Registry Number.
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6.8.2 C -LAS, sodium linear undecylbenzene sulfonate, C H SO Na; 99 %.
11 17 27 3

6.8.3 C -LAS, sodium linear dodecylbenzene sulfonate, C H SO Na; 99 % (CAS-RN 2211-98-5).

12 18 29 3
6.8.4 C -LAS, sodium linear tridecylbenzene sulfonate, C H SO Na; 99 %.
13 19 31 3

6.8.5 C -C -LAS mixture of homologues and isomers, highest possible purity (CAS-RN 69669-44-9,

10 14
CAS-RN 25155-30-0).
6.9 Internal standard, C -LAS
Octylbenzene sulfonic acid, sodium salt C H SO Na (CAS-RN 6149-03-7).
14 21 3

The internal standard shall be stored in the freezer at a temperature of (–18 ± 3) °C.

6.10 Internal standard solution

Prepare the internal standard solution of the internal standard (6.9) by dilution to about 1 000 mg/l in methanol

(6.2).

It is essential that the same internal standard solution is used for calibration standard solutions and for

samples, blank tests and internal quality control samples.

Store the internal standard solution in a dark place at a temperature of (4 ± 3) °C. The solution is stable for at

least two years.
6.11 Stock solutions

Prepare individual stock solutions of 1 000 mg/l to 5 000 mg/l in methanol (6.2), either from solid standard

substances or from solutions with a certified concentration. Prepare stock solutions of C -LAS (6.8.2),

C -LAS (6.8.3) and C -LAS (6.8.4).
12 13

Prepare a calibration mixture by mixing stock solutions of C -LAS, C -LAS and C -LAS containing equal

11 12 13
concentrations of each homologue.

Prepare a stock solution of C to C -LAS mixture (6.8.5) of 1 000 mg/l to 5 000 mg/l in methanol (6.2). This

10 14
solution is only for identification.

Store the stock solutions and the calibration mixture in a dark place at a temperature of (4 ± 3) °C. The

solutions are stable for at least two years.
6.12 Calibration standard solutions
6.12.1 General

Calibration standard solutions are prepared from the calibration mixture (6.11) by diluting with a 1:1 mixture of

methanol (6.2) and ammonium acetate (6.4).

The calibration range is different for sludge (see 6.12.2) and for soil or treated biowaste (see 6.12.3).

Store the calibration standard solutions in a dark place at a temperature of (4 ± 3) °C.

NOTE A diluted C - to C -mixture is prepared for the identification of the C and C homologues, which are not

10 14 10 14
present in the calibration mixture.
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6.12.2 Sludge samples

For sludge samples the calibration standards are prepared for concentrations from 5 mg/l to 500 mg/l. The

internal standard solution (6.10) is added to a concentration of 10 mg/l.
6.12.3 Soil and treated biowaste samples

For samples of soil or treated biowaste the calibration standards are prepared for concentrations from

0,05 mg/l to 5 mg/l. The internal standard solution (6.10) is added to a concentration of 1 mg/l.

7 Apparatus
7.1 General

All equipment that comes into contact with the sample or extract shall be free from LAS. Glassware shall be

cleaned by heating, at least for 2 h at 450 °C.
7.2 Usual laboratory glassware

7.2.1 Screw cap glass flask with polytetrafluoroethylene (PTFE) seal; volume 20 ml and 100 ml.

7.2.2 Round-bottom flasks, volume 100 ml and 250 ml.
7.2.3 Test tubes and vials
7.3 Shaking device

Reciprocating shaker with horizontal movement (suitable for (250 ± 20) strokes per minute).

7.4 Evaporator
Rotary evaporator, turbo evaporator or Kuderna Danish .
7.5 Freeze drying apparatus

7.6 High-performance liquid chromatograph with fluorescence or mass selective detector

The HPLC system is equipped with a C - or C -reverse phase chromatographic column. The dimensions

8 18
should be sufficient to separate the LAS as described below.
NOTE Two examples for HPLC-columns are given in Annex B.

The fluorescence detector shall be suitable to measure at excitation wavelength of 230 nm and emission

wavelength of 310 nm. If a fixed wavelength detector is used, the nearest possible wavelengths shall be used.

The mass selective detector shall be equipped with an atmospheric pressure ionization electro-spray (API-ES)

interface. Use the negative ion mode.

The separation of LAS homologues shall fulfil the following requirements: The five homologues C to C

10 14
shall all be separated to baseline.

Isomeric separation (mandatory for fluorescence detection): C -LAS shall be separated into at least four

chromatographic peaks, although these are not separated to baseline.

Kuderna Danish is an example of a suitable product available commercially. This information is given for the

convenience of users of this Technical Specification and does not constitute an endorsement by CEN of this product.

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CEN/TS 16189:2012 (E)
8 Sample storage and sample pretreatment
8.1 Sample storage

Store the samples in a dark place at a temperature below 10 °C, if possible in a refrigerator.

Determine the content of dry matter in the sample according to EN 15934.

NOTE 1 Freeze-dried samples, if kept sealed, may be stored for a longer period at room temperature (approximately

one month).

NOTE 2 Hygroscopic dried sludge may be preserved by mixing with anhydrous sodium sulfate.

8.2 Sample pretreatment
Pretreat the sample according to EN 16179 if not otherwise specified.

All samples shall be dried, preferably by freeze drying. Conventional drying at 60 °C can be used, but this may

result in crusty hard samples that are difficult to homogenize.
9 Procedure
9.1 Extraction
9.1.1 Extraction of dried sludge

 Weigh accurately 2 g to 3 g (with two decimals) of the test sample and place it in a screw cap flask

(7.2.1).

 Add 100 µl of internal standard solution (6.10) equal to 100 µg of internal standard (6.9).

 Add 10 ml of methanol (6.2), close the screw cap and place the flask on the shaking device (7.3). The

flask shall be placed in horizontal position.
 Shake for at least 30 min with (250 ± 20) strokes per minute.

 Wait for sample to settle, then transfer 500 µl of the extract to a vial (7.2.3) and add 500 µl ammonium

acetate (mobile phase A) (6.5.1).
The extract is now ready for analysis.

In case of high LAS concentrations less of the test sample can be taken for analysis.

The extract can be stored in a refrigerator at (4 ± 3) °C.
9.1.2 Extraction of dried soil, sediment and treated biowaste

 Weigh accurately 10 g to 15 g (with two decimals) of the test sample and place it in a 100 ml screw cap

flask (7.2.1).

 Add 50 µl of internal standard solution (6.10) equal to 50 µg of internal standard (6.9).

 Add 50 ml of methanol (6.2), close the screw cap and place the flask on the shaking device (7.3). The

flask shall be placed in horizontal position.
 Shake for at least 60 min with (250 ± 20) strokes per minute.
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CEN/TS 16189:2012 (E)

 Wait for sample to settle, then transfer 500 µl of the extract to a vial (7.2.3) and add 500 µl ammonium

acetate (mobile phase A) (6.5.1).
The extract is now ready for analysis.
The extract can be stored in a refrigerator at (4 ± 3) °C.

NOTE Other extraction techniques, like ultrasonic extraction, Soxhlet, reflux, microwave or pressurised liquid

extraction may be suitable. However, if using other extraction techniques, the comparability to the method described in this

Technical Specification should be proven.
9.2 Concentration (optional)

In most cases concentration of the extract is not necessary. If lower detection limits are required, this can be

achieved by evaporation of the solvent.

Concentrate the extract on an evaporator (7.4) or by the use of a gentle stream of nitrogen at room

temperature. Since the internal standard (6.9) is used for the calculations, it is not necessary to know the

exact volumes. If necessary, the amount of the internal standard added to the sample can be reduced relative

to the concentration factor to keep the concentration of the internal standard at the same level in the analysis.

9.3 Clean-up (optional)

Clean-up shall be used if compounds are present that can interfere with the analytes or the internal standard

(6.9) in the chromatography, or if those compounds can influence the HPLC-procedure (i. e. contamination of

the detection system). If no or negligible interfering substances are present, clean-up is not necessary.

The selecti
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