ISO 23191:2020
(Main)Traditional Chinese medicine — Determination of selected Aconitum alkaloids by high-performance liquid chromatography (HPLC)
Traditional Chinese medicine — Determination of selected Aconitum alkaloids by high-performance liquid chromatography (HPLC)
This document specifies methods for the determination of the selected Aconitum alkaloids, including aconitine, mesaconitine, hypaconitine, benzoylaconine, benzoylmesaconine, benzoylhypaconine, yunaconitine, deacetyl-yunaconitine and crassicauline A.
Médecine traditionnelle chinoise — Dosage d'alcaloïdes d'aconit (Aconitum) sélectionnés par chromatographie liquide à haute performance (CLHP)
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INTERNATIONAL ISO
STANDARD 23191
First edition
2020-07
Traditional Chinese medicine —
Determination of selected Aconitum
alkaloids by high-performance liquid
chromatography (HPLC)
Médecine traditionnelle chinoise — Dosage d'alcaloïdes d'aconit
(Aconitum) sélectionnés par chromatographie liquide à haute
performance (CLHP)
Reference number
©
ISO 2020
© ISO 2020
All rights reserved. Unless otherwise specified, or required in the context of its implementation, no part of this publication may
be reproduced or utilized otherwise in any form or by any means, electronic or mechanical, including photocopying, or posting
on the internet or an intranet, without prior written permission. Permission can be requested from either ISO at the address
below or ISO’s member body in the country of the requester.
ISO copyright office
CP 401 • Ch. de Blandonnet 8
CH-1214 Vernier, Geneva
Phone: +41 22 749 01 11
Email: copyright@iso.org
Website: www.iso.org
Published in Switzerland
ii © ISO 2020 – All rights reserved
Contents Page
Foreword .iv
Introduction .v
1 Scope . 1
2 Normative references . 1
3 Terms and definitions . 1
4 Abbreviated terms . 1
5 Principle . 2
6 Reagents . 3
7 Apparatus . 3
8 Sampling . 4
9 Test procedures . 4
9.1 General . 4
9.2 HPLC-MS procedure . 5
9.2.1 Reference solution . 5
9.2.2 Test solution. 5
9.2.3 Chromatographic conditions . 5
9.2.4 HPLC-MS conditions . 5
9.2.5 Determination . 5
9.3 HPLC-DAD procedure . 5
9.3.1 Reference solution . 5
9.3.2 Test solution. 6
9.3.3 Chromatographic conditions . 6
9.3.4 Determination . 6
10 Test report . 6
Annex A (informative) Typical HPLC conditions . 7
Annex B (informative) Typical chromatogram of the selected Aconitum alkaloids .9
Annex C (informative) Typical Aconitum herb used in traditional Chinese medicine .12
Annex D (informative) Reference values of national and regional limits of Aconitum
alkaloids in Aconitum carmichaelii lateral root .13
Bibliography .14
Foreword
ISO (the International Organization for Standardization) is a worldwide federation of national standards
bodies (ISO member bodies). The work of preparing International Standards is normally carried out
through ISO technical committees. Each member body interested in a subject for which a technical
committee has been established has the right to be represented on that committee. International
organizations, governmental and non-governmental, in liaison with ISO, also take part in the work.
ISO collaborates closely with the International Electrotechnical Commission (IEC) on all matters of
electrotechnical standardization.
The procedures used to develop this document and those intended for its further maintenance are
described in the ISO/IEC Directives, Part 1. In particular, the different approval criteria needed for the
different types of ISO documents should be noted. This document was drafted in accordance with the
editorial rules of the ISO/IEC Directives, Part 2 (see www .iso .org/ directives).
Attention is drawn to the possibility that some of the elements of this document may be the subject of
patent rights. ISO shall not be held responsible for identifying any or all such patent rights. Details of
any patent rights identified during the development of the document will be in the Introduction and/or
on the ISO list of patent declarations received (see www .iso .org/ patents).
Any trade name used in this document is information given for the convenience of users and does not
constitute an endorsement.
For an explanation of the voluntary nature of standards, the meaning of ISO specific terms and
expressions related to conformity assessment, as well as information about ISO's adherence to the
World Trade Organization (WTO) principles in the Technical Barriers to Trade (TBT) see www .iso .org/
iso/ foreword .html.
This document was prepared by ISO/TC 249, Traditional Chinese medicine.
Any feedback or questions on this document should be directed to the user’s national standards body. A
complete listing of these bodies can be found at www .iso .org/ members .html.
iv © ISO 2020 – All rights reserved
Introduction
Aconitum is a genus of about 350 species of flowering plants belonging to the family of Ranunculaceae.
The therapeutic use of as many as 76 species of Aconitum herbs in traditional Chinese medicine have
been documented for a very long time. Among the Aconitum herbs, the most well known are processed
Aconitum carmichaelii lateral root (附子), Aconitum carmichaelii root (川乌) and Aconitum kusnezoffii
root (草乌). Of approximately 80 000 traditional Chinese medicine formulae, around 10,2 % contain
Aconitum herbs. It is one of the most frequently used groups of herbal medicines in traditional Chinese
medicine. Typical Aconitum herbs used in traditional Chinese medicine are shown in Annex C, Table C.1.
Aconitum herbs contain Aconitum alkaloids which have anti-inflammatory, analgesic and cardiotonic
activities. The Aconitum alkaloids are a double-edged sword, however. At present, international trade
in Aconitum products is restricted to a few nations due to the high natural toxicity of crude Aconitum
products. Unprocessed Aconitum is highly toxic. Expert processing is required to reduce (but not
eliminate) toxicity, and where individual national regulatory schemes do not ban the herb, it is generally
restricted to a high-risk category, such as Schedule 2 in the Hong Kong Chinese Medicine regulations.
Also, there are sporadic cases of Aconitum alkaloid poisoning due to misuse reported all over the world.
Nonetheless, the toxicity of Aconitum herbs can be reduced dramatically with proper processing (such
as repeated boiling or steaming), prolonged decoction and dose control. However, testing standards for
Aconitum alkaloids have not yet been harmonized on an international level, and regulatory authorities
in many nations do not adequately differentiate highly toxic forms from less-toxic forms (or even non-
toxic forms) of Aconitum herbs.
Six kinds of Aconitum alkaloids [aconitine (AC), mesaconitine (MA), hypaconitine (HA), benzoylaconine
(BAC), benzoylmesaconine (BMA) and benzoylhypaconine (BHA)] are commonly used as chemical
markers for quality control of aconite, determined by the high-performance liquid chromatography
[1]
(HPLC) method . The AOAC Official Method 2008.11 also requires the determination of three Aconitum
alkaloids, AC, MA and HA, in dietary supplements and raw botanical materials by LC/UV detection
[2]
with confirmation by LC/MS/MS . Nevertheless, poisoning cases are still occasionally reported. From
[3]
1989 to 2010, 140 cases of Aconitum poisoning, including one fatal case, were reported in Hong Kong .
Additionally, 17 cases were reported in Taiwan from 1990 to 1999, 2017 cases in China from 1989 to
[4]
2008 and 121 cases in Korea from 1995 to 2007 . Multiple reasons for Aconitum poisoning exist and
include overdoses, inadequate processing, Aconitum contamination in other herbs, dispensing and
management errors and hidden risk factors. In the 17 cases reported in Hong Kong, yunaconitine (YAC),
crassicauline A (CCA) and 8-deacetyl-yunaconitine (DYA) were detected instead of AC, MA and HA in the
[3,4]
urine samples of the Aconitum poisoning patients . As a result, these alkaloids are considered to be
[5]
hidden risk factors and should be covered in laboratory screenings for toxic compounds . Therefore,
a method to simultaneously determine the levels of these nine alkaloids is needed for quality control of
[6]
the herb and its products in order to ensure the safe use of these medicinal materials .
This document aims to build a systematic and practical international standard for the determination of
Aconitum alkaloids with the goal of standardizing the global market, to ensure safe and effective use in
clinics and to reduce cases of Aconitum alkaloid poisoning.
As national implementation may differ, national standards bodies are invited to modify the limit values
of selected Aconitum alkaloids in their national standards. Examples of national and regional values are
given in Annex D.
INTERNATIONAL STANDARD ISO 23191:2020(E)
Traditional Chinese medicine — Determination of
selected Aconitum alkaloids by high-performance liquid
chromatography (HPLC)
1 Scope
This document specifies methods for the determination of the selected Aconitum alkaloids, including
aconitine, mesaconitine, hypaconitine, benzoylaconine, benzoylmesaconine, benzoylhypaconine,
yunaconitine, deacetyl-yunaconitine and crassicauline A.
2 Normative references
The following documents are referred to in the text in such a way that some or all of their content
constitutes requirements of this document. For dated references, only the edition cited applies. For
undated references, the latest edition of the referenced document (including any amendments) applies.
ISO 3696, Water for analytical laboratory use — Specification and test methods
World Health Organization, Quality control methods for herbal materials, 2011
3 Terms and definitions
No terms and definitions are listed in this document.
ISO and IEC maintain terminological databases for use in standardization at the following addresses:
— ISO Online browsing platform: available at https:// www .iso .org/ obp
— IEC Electropedia: available at http:// www .electropedia .org/
4 Abbreviated terms
For the purposes of this document, the following abbreviated terms apply.
AC aconitine
BAC benzoylaconine
BHA benzoylhypaconine
BMA benzoylmesaconine
CAS chemical abstracts service
CCA cra
...
INTERNATIONAL ISO
STANDARD 23191
First edition
2020-07
Traditional Chinese medicine —
Determination of selected Aconitum
alkaloids by high-performance liquid
chromatography (HPLC)
Médecine traditionnelle chinoise — Dosage d'alcaloïdes d'aconit
(Aconitum) sélectionnés par chromatographie liquide à haute
performance (CLHP)
Reference number
©
ISO 2020
© ISO 2020
All rights reserved. Unless otherwise specified, or required in the context of its implementation, no part of this publication may
be reproduced or utilized otherwise in any form or by any means, electronic or mechanical, including photocopying, or posting
on the internet or an intranet, without prior written permission. Permission can be requested from either ISO at the address
below or ISO’s member body in the country of the requester.
ISO copyright office
CP 401 • Ch. de Blandonnet 8
CH-1214 Vernier, Geneva
Phone: +41 22 749 01 11
Email: copyright@iso.org
Website: www.iso.org
Published in Switzerland
ii © ISO 2020 – All rights reserved
Contents Page
Foreword .iv
Introduction .v
1 Scope . 1
2 Normative references . 1
3 Terms and definitions . 1
4 Abbreviated terms . 1
5 Principle . 2
6 Reagents . 3
7 Apparatus . 3
8 Sampling . 4
9 Test procedures . 4
9.1 General . 4
9.2 HPLC-MS procedure . 5
9.2.1 Reference solution . 5
9.2.2 Test solution. 5
9.2.3 Chromatographic conditions . 5
9.2.4 HPLC-MS conditions . 5
9.2.5 Determination . 5
9.3 HPLC-DAD procedure . 5
9.3.1 Reference solution . 5
9.3.2 Test solution. 6
9.3.3 Chromatographic conditions . 6
9.3.4 Determination . 6
10 Test report . 6
Annex A (informative) Typical HPLC conditions . 7
Annex B (informative) Typical chromatogram of the selected Aconitum alkaloids .9
Annex C (informative) Typical Aconitum herb used in traditional Chinese medicine .12
Annex D (informative) Reference values of national and regional limits of Aconitum
alkaloids in Aconitum carmichaelii lateral root .13
Bibliography .14
Foreword
ISO (the International Organization for Standardization) is a worldwide federation of national standards
bodies (ISO member bodies). The work of preparing International Standards is normally carried out
through ISO technical committees. Each member body interested in a subject for which a technical
committee has been established has the right to be represented on that committee. International
organizations, governmental and non-governmental, in liaison with ISO, also take part in the work.
ISO collaborates closely with the International Electrotechnical Commission (IEC) on all matters of
electrotechnical standardization.
The procedures used to develop this document and those intended for its further maintenance are
described in the ISO/IEC Directives, Part 1. In particular, the different approval criteria needed for the
different types of ISO documents should be noted. This document was drafted in accordance with the
editorial rules of the ISO/IEC Directives, Part 2 (see www .iso .org/ directives).
Attention is drawn to the possibility that some of the elements of this document may be the subject of
patent rights. ISO shall not be held responsible for identifying any or all such patent rights. Details of
any patent rights identified during the development of the document will be in the Introduction and/or
on the ISO list of patent declarations received (see www .iso .org/ patents).
Any trade name used in this document is information given for the convenience of users and does not
constitute an endorsement.
For an explanation of the voluntary nature of standards, the meaning of ISO specific terms and
expressions related to conformity assessment, as well as information about ISO's adherence to the
World Trade Organization (WTO) principles in the Technical Barriers to Trade (TBT) see www .iso .org/
iso/ foreword .html.
This document was prepared by ISO/TC 249, Traditional Chinese medicine.
Any feedback or questions on this document should be directed to the user’s national standards body. A
complete listing of these bodies can be found at www .iso .org/ members .html.
iv © ISO 2020 – All rights reserved
Introduction
Aconitum is a genus of about 350 species of flowering plants belonging to the family of Ranunculaceae.
The therapeutic use of as many as 76 species of Aconitum herbs in traditional Chinese medicine have
been documented for a very long time. Among the Aconitum herbs, the most well known are processed
Aconitum carmichaelii lateral root (附子), Aconitum carmichaelii root (川乌) and Aconitum kusnezoffii
root (草乌). Of approximately 80 000 traditional Chinese medicine formulae, around 10,2 % contain
Aconitum herbs. It is one of the most frequently used groups of herbal medicines in traditional Chinese
medicine. Typical Aconitum herbs used in traditional Chinese medicine are shown in Annex C, Table C.1.
Aconitum herbs contain Aconitum alkaloids which have anti-inflammatory, analgesic and cardiotonic
activities. The Aconitum alkaloids are a double-edged sword, however. At present, international trade
in Aconitum products is restricted to a few nations due to the high natural toxicity of crude Aconitum
products. Unprocessed Aconitum is highly toxic. Expert processing is required to reduce (but not
eliminate) toxicity, and where individual national regulatory schemes do not ban the herb, it is generally
restricted to a high-risk category, such as Schedule 2 in the Hong Kong Chinese Medicine regulations.
Also, there are sporadic cases of Aconitum alkaloid poisoning due to misuse reported all over the world.
Nonetheless, the toxicity of Aconitum herbs can be reduced dramatically with proper processing (such
as repeated boiling or steaming), prolonged decoction and dose control. However, testing standards for
Aconitum alkaloids have not yet been harmonized on an international level, and regulatory authorities
in many nations do not adequately differentiate highly toxic forms from less-toxic forms (or even non-
toxic forms) of Aconitum herbs.
Six kinds of Aconitum alkaloids [aconitine (AC), mesaconitine (MA), hypaconitine (HA), benzoylaconine
(BAC), benzoylmesaconine (BMA) and benzoylhypaconine (BHA)] are commonly used as chemical
markers for quality control of aconite, determined by the high-performance liquid chromatography
[1]
(HPLC) method . The AOAC Official Method 2008.11 also requires the determination of three Aconitum
alkaloids, AC, MA and HA, in dietary supplements and raw botanical materials by LC/UV detection
[2]
with confirmation by LC/MS/MS . Nevertheless, poisoning cases are still occasionally reported. From
[3]
1989 to 2010, 140 cases of Aconitum poisoning, including one fatal case, were reported in Hong Kong .
Additionally, 17 cases were reported in Taiwan from 1990 to 1999, 2017 cases in China from 1989 to
[4]
2008 and 121 cases in Korea from 1995 to 2007 . Multiple reasons for Aconitum poisoning exist and
include overdoses, inadequate processing, Aconitum contamination in other herbs, dispensing and
management errors and hidden risk factors. In the 17 cases reported in Hong Kong, yunaconitine (YAC),
crassicauline A (CCA) and 8-deacetyl-yunaconitine (DYA) were detected instead of AC, MA and HA in the
[3,4]
urine samples of the Aconitum poisoning patients . As a result, these alkaloids are considered to be
[5]
hidden risk factors and should be covered in laboratory screenings for toxic compounds . Therefore,
a method to simultaneously determine the levels of these nine alkaloids is needed for quality control of
[6]
the herb and its products in order to ensure the safe use of these medicinal materials .
This document aims to build a systematic and practical international standard for the determination of
Aconitum alkaloids with the goal of standardizing the global market, to ensure safe and effective use in
clinics and to reduce cases of Aconitum alkaloid poisoning.
As national implementation may differ, national standards bodies are invited to modify the limit values
of selected Aconitum alkaloids in their national standards. Examples of national and regional values are
given in Annex D.
INTERNATIONAL STANDARD ISO 23191:2020(E)
Traditional Chinese medicine — Determination of
selected Aconitum alkaloids by high-performance liquid
chromatography (HPLC)
1 Scope
This document specifies methods for the determination of the selected Aconitum alkaloids, including
aconitine, mesaconitine, hypaconitine, benzoylaconine, benzoylmesaconine, benzoylhypaconine,
yunaconitine, deacetyl-yunaconitine and crassicauline A.
2 Normative references
The following documents are referred to in the text in such a way that some or all of their content
constitutes requirements of this document. For dated references, only the edition cited applies. For
undated references, the latest edition of the referenced document (including any amendments) applies.
ISO 3696, Water for analytical laboratory use — Specification and test methods
World Health Organization, Quality control methods for herbal materials, 2011
3 Terms and definitions
No terms and definitions are listed in this document.
ISO and IEC maintain terminological databases for use in standardization at the following addresses:
— ISO Online browsing platform: available at https:// www .iso .org/ obp
— IEC Electropedia: available at http:// www .electropedia .org/
4 Abbreviated terms
For the purposes of this document, the following abbreviated terms apply.
AC aconitine
BAC benzoylaconine
BHA benzoylhypaconine
BMA benzoylmesaconine
CAS chemical abstracts service
CCA cra
...
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