32 ( 199 1h 195--l
Journul of Erh~o~ha~maco~~g~. Elsevier Scientific
Publishers Ireland
195
97
Ltd.
How and why should we standardize phytopharmaceutical drugs for clinical validation? A. Bonati
Standardized of analytical
extracts are needed to carry out reliable clinical trials. The standardization
controls: it requires also a thorough
description
items for the constancy of the quality of an extract. Another
not minor aspect in the standardization
which must include all the necessary data for a clear definition itself. Two examples of standardization (anthocyanoside
cal markers (Pygeum africunum
and the other concerning
Presented at the First International Strasbourg,
C[~rr~~~~~~~~~~e fo: Ripamonti.
99, 20141
0378-8741/$03.50
0
5-9 A.
Bonati,
Congress on Ethnophartnverni
della
Beffa,
Via
Italy.
1991 Elsevier Scientific
Published and Printed in Ireland
of the extract are known
an extract whose constituents must be regarded as chemi-
body of information and controls that are necessary to guarantee constancy of composition - hence the standardized quality - of a phytopharmdceutical drug. (b) Directives on the analytical control of a vegetable drug must take account of the fact that the material to be examined has a complex and “inconstant” composition. Because of this: the analytical limits cannot be so precise as for chemically pure products. the identity tests must be specific and include several groups of constituents so as to arrive at a cross-proof of the composition. The importance of a specific identification that includes several groups of substances is clear: it allows the detection of adulteration and degradations of the material to be used. A cross-identification also allows the “discovery” of reconstructed extracts, that is extracts obtained by diluting an extract that has a high content of a given constituent with large quantities of excipients.
June, 1990.
Milan,
composition)
active constituents
extract).
The answer to the question “Why standardize phytopharmaceutical drugs?” is obvious: to conduct reliable clinical trials one needs active principles of well-defined and constant quality, which, in the case of a phytopharmaceutical, means of standardized quality. Not so obvious is the answer to the question “How to standardize phytopharmaceutical drugs?“: on this purpose I recall that, according to the EEC directives on the quality of herbal remedies, “a vegetable drug or a preparation thereof is regarded as one active ingredient in its entirety, whether or not the constituents with therapeutic activity are known”. Some general points are in order here: (a) The standardization of a vegetable drug and of a preparation thereof is not just an analytical operation: it does not end with the identification and assay of an active principle or of a marker. Standardization signifies the
macology,
an extract whose therapeutical
process, both basic
of an extract is its nomenclature
(drug, physical state. solvent of extraction,
are described, one concerning
complex of Vaccinium myrtillus)
of an extract does not include only a whole
of the starting drug and of the whole extraction
Publishers
Ireland
Ltd.
196
(c) Vegetable drugs are inevitably “inconstant” because their composition, and hence their standardization, may be influenced by several factors: age and origin, harvesting period, method of drying and so on. To elimate some of the causes of inconstancy one should use cultivated rather than wild plants, which are often heterogeneous in respect of the above factors and consequently in their content of active principle. Cultivated plants offer several advantages in terms of standardization: can be cultivated in the plants homogeneous climatic and soil conditions; plants of known age can be harvested at the right time and quickly; plants can be dried in controlled time and temperature conditions. It should never be forgotten that a homogeneous and correct drying is often the most delicate and essential phase in the whole process of production. (d) Because of the complex composition of vegetable drugs, the process of production of a vegetable drug preparation (extracts, tinctures etc.) is crucial to the constancy of quality: it has to be kept constant because only through a constant process of extraction is it possible to obtain from a standardized vegetable drug a standardized vegetable drug preparation. As I said earlier, standardization is the body of information and controls that guarantees the constancy of composition - and therefore the constancy of activity - of a phytopharmaceutical product. Although chemical analysis is the most important part of standardization, it must be completed: in the case of the vegetable drug, with information on the place and period of harvesting, period of vegetation, treatments given during growth (e.g. pesticides), method of drying, storage (use of fumigants). in the case of the extract, with information on the method of production (particle size, type of extraction, extraction solvent, conditions
of concentration, any purification processes, any additions of excipients to bring the extract to a given level of constituents with known therapeutic activity or for any other reason) and on conditions of storage. With regard, more specifically, to the analytical controls, I will make only a few general remarks. For identity tests, chromatographic methods (TLC, GLC, HPLC) yield results that may be regarded as satisfactory. I repeat once again that the chromatographic identification should not be confined to a single product but extended to several groups of constituents in order that the composition of a phytopharmaceutical may be demonstrated as completely as possible. Further, cross-testing on several substances is a precondition for an objective study of stability. Certain chromatographic techniques, especially HPLC and GLC, are to be considered the most suitable for the assay of vegetable drugs and their derivatives. However, they are not always applicable: for a quantitative determination equally satisfactory results can be obtained by combining non-specific methods with specific chromatogra’phic procedures. As a rule, vegetable drugs contain several products with similar structure. In the absence of a specific HPLC or GLC determination, other methods have to be used (spectrophotometry, colorimetry, gravimetric determination, ester or acid or hydroxyl value): in this particular “complex“ situation these methods supply only “global“ results and have to be combined with specific chromatographic identifications. Here is the general rule of many pharmacopoeias: the assay of total alkaloids, hydroxyanthracene derivatives, total anthocyanosides etc. is always combined with specific TLC identifications. The analyses must include the determination of the physico-chemical characteristics. These are not merely routine tests: each has its value and its importance both for the stability (pH, alcoholic degree) and for the constancy of content of extractive material (total solids). Total microbial content, pesticides, results of stability tests in normal and stress conditions are supplementary data for the standardization of a phytopharmaceutical drug.
197
Another aspect of by no means secondary importance for the standardization of a phytopharmaceutical product is its nomenclature. The problem of a rational nomenclature has been solved satisfactorily for pure products, both synthetic and extractive. In the case of vegetable drugs and preparations thereof the situation is somewhat different and, unfortunately, confused because there is no single universally accepted name for a vegetable drug or extract. This situation creates confusion and uncertainty: it prevents the comparative evaluation of pharmacological and clinical results obtained by different research teams and makes it impossible for the health authorities and for the physicians to compare phytopharmaceuticals containing the same active principles on the basis of rational and homogeneous names, thereby giving rise to errors in doses and prices. All this throws discredit on an entire field of great therapeutic and market importance. It is therefore necessary to draw up rules that will ensure clear and simple nomenclature for phytopharmaceuticals: I will make a few suggestions. Nomenclature of vegetable drugs A vegetable drug should be referred to solely by its correct botanical name followed by the name of the authority and by the part of the plant used. For example: Cinchona succirubru Pavon, bark. When a drug is used fresh, this must be specified because this factor is crucial for the composition of the vegetable drug. For example: Vaccinium m_~rtilius L., fresh fruits. If the constituents responsible for the therapeutic activity are known, their quantities must be stated. For example: Atropa belladonna L., leaves, containing . . .% of total alkaloids calculated as hyoscyamine. Extracts In the case of total extracts the name of the drug must be followed by the drug/extract ratio, by the solvent used for extraction and by the physical form (fluid, soft or dry). For example: Rhamnus purshiana DC., bark, l:l, 20%
hydroalcoholic fluid extract. Rhamnus purshiana DC., bark, 6:1, 60% hydroal~oholic dry extract. If the constituents responsible for the therapeutic activity are known, their quantity should be stated instead of the extract/drug ratio. If some other substance is added during the preparation of an extract to bring the content of active principles up to a given level, or for any other reason, the substance added must be reported as “other constituent(s)” and the extract (natural extract) as “active principle”.
-
Purified extracts The preparation of a purified extract includes phases of fractionation or purification (usually protected by patent), which are not simple operations of defatting but very important processes that increase the content of active principle appreciably beyond that of a total extract. The general nomenclature for total extracts cited earlier cannot be used for these extracts. A satisfactory approach might be to write the name of the vegetable drug followed by a term such as “anthocyanoside complex” or “liposoluble fraction (or complex)” or “total alkaloids”. Indication of the physical state may be unnecessary because these purified extracts are practically always dry extracts or, in the case of liposoluble fractions, oily products. The name of a purified extract must always be followed by the content of active principles. The trademark may be added in brackets, Examples: - Vaccinium myrtillus L., fresh fruits, anthocyanoside complex containing 35% anthocyanosides (~yrto~yan R). - Atropa belladonna L., leaves, 90% total alkaloids calculated as hyoscyamine. - Serenoa repens, fruits, liposoluble fraction, containing 90% of C6 to CzO linear chain fatty acids. These are only suggestions, certainly not 100% valid or acceptable: at all events, a rational nomenclature for phytopharmaceuticals is very important because it is the key to any comparison of activity, dosage and price.
Journul of Erh~o~ha~maco~~g~. Elsevier Scientific
Publishers Ireland
195
97
Ltd.
How and why should we standardize phytopharmaceutical drugs for clinical validation? A. Bonati
Standardized of analytical
extracts are needed to carry out reliable clinical trials. The standardization
controls: it requires also a thorough
description
items for the constancy of the quality of an extract. Another
not minor aspect in the standardization
which must include all the necessary data for a clear definition itself. Two examples of standardization (anthocyanoside
cal markers (Pygeum africunum
and the other concerning
Presented at the First International Strasbourg,
C[~rr~~~~~~~~~~e fo: Ripamonti.
99, 20141
0378-8741/$03.50
0
5-9 A.
Bonati,
Congress on Ethnophartnverni
della
Beffa,
Via
Italy.
1991 Elsevier Scientific
Published and Printed in Ireland
of the extract are known
an extract whose constituents must be regarded as chemi-
body of information and controls that are necessary to guarantee constancy of composition - hence the standardized quality - of a phytopharmdceutical drug. (b) Directives on the analytical control of a vegetable drug must take account of the fact that the material to be examined has a complex and “inconstant” composition. Because of this: the analytical limits cannot be so precise as for chemically pure products. the identity tests must be specific and include several groups of constituents so as to arrive at a cross-proof of the composition. The importance of a specific identification that includes several groups of substances is clear: it allows the detection of adulteration and degradations of the material to be used. A cross-identification also allows the “discovery” of reconstructed extracts, that is extracts obtained by diluting an extract that has a high content of a given constituent with large quantities of excipients.
June, 1990.
Milan,
composition)
active constituents
extract).
The answer to the question “Why standardize phytopharmaceutical drugs?” is obvious: to conduct reliable clinical trials one needs active principles of well-defined and constant quality, which, in the case of a phytopharmaceutical, means of standardized quality. Not so obvious is the answer to the question “How to standardize phytopharmaceutical drugs?“: on this purpose I recall that, according to the EEC directives on the quality of herbal remedies, “a vegetable drug or a preparation thereof is regarded as one active ingredient in its entirety, whether or not the constituents with therapeutic activity are known”. Some general points are in order here: (a) The standardization of a vegetable drug and of a preparation thereof is not just an analytical operation: it does not end with the identification and assay of an active principle or of a marker. Standardization signifies the
macology,
an extract whose therapeutical
process, both basic
of an extract is its nomenclature
(drug, physical state. solvent of extraction,
are described, one concerning
complex of Vaccinium myrtillus)
of an extract does not include only a whole
of the starting drug and of the whole extraction
Publishers
Ireland
Ltd.
196
(c) Vegetable drugs are inevitably “inconstant” because their composition, and hence their standardization, may be influenced by several factors: age and origin, harvesting period, method of drying and so on. To elimate some of the causes of inconstancy one should use cultivated rather than wild plants, which are often heterogeneous in respect of the above factors and consequently in their content of active principle. Cultivated plants offer several advantages in terms of standardization: can be cultivated in the plants homogeneous climatic and soil conditions; plants of known age can be harvested at the right time and quickly; plants can be dried in controlled time and temperature conditions. It should never be forgotten that a homogeneous and correct drying is often the most delicate and essential phase in the whole process of production. (d) Because of the complex composition of vegetable drugs, the process of production of a vegetable drug preparation (extracts, tinctures etc.) is crucial to the constancy of quality: it has to be kept constant because only through a constant process of extraction is it possible to obtain from a standardized vegetable drug a standardized vegetable drug preparation. As I said earlier, standardization is the body of information and controls that guarantees the constancy of composition - and therefore the constancy of activity - of a phytopharmaceutical product. Although chemical analysis is the most important part of standardization, it must be completed: in the case of the vegetable drug, with information on the place and period of harvesting, period of vegetation, treatments given during growth (e.g. pesticides), method of drying, storage (use of fumigants). in the case of the extract, with information on the method of production (particle size, type of extraction, extraction solvent, conditions
of concentration, any purification processes, any additions of excipients to bring the extract to a given level of constituents with known therapeutic activity or for any other reason) and on conditions of storage. With regard, more specifically, to the analytical controls, I will make only a few general remarks. For identity tests, chromatographic methods (TLC, GLC, HPLC) yield results that may be regarded as satisfactory. I repeat once again that the chromatographic identification should not be confined to a single product but extended to several groups of constituents in order that the composition of a phytopharmaceutical may be demonstrated as completely as possible. Further, cross-testing on several substances is a precondition for an objective study of stability. Certain chromatographic techniques, especially HPLC and GLC, are to be considered the most suitable for the assay of vegetable drugs and their derivatives. However, they are not always applicable: for a quantitative determination equally satisfactory results can be obtained by combining non-specific methods with specific chromatogra’phic procedures. As a rule, vegetable drugs contain several products with similar structure. In the absence of a specific HPLC or GLC determination, other methods have to be used (spectrophotometry, colorimetry, gravimetric determination, ester or acid or hydroxyl value): in this particular “complex“ situation these methods supply only “global“ results and have to be combined with specific chromatographic identifications. Here is the general rule of many pharmacopoeias: the assay of total alkaloids, hydroxyanthracene derivatives, total anthocyanosides etc. is always combined with specific TLC identifications. The analyses must include the determination of the physico-chemical characteristics. These are not merely routine tests: each has its value and its importance both for the stability (pH, alcoholic degree) and for the constancy of content of extractive material (total solids). Total microbial content, pesticides, results of stability tests in normal and stress conditions are supplementary data for the standardization of a phytopharmaceutical drug.
197
Another aspect of by no means secondary importance for the standardization of a phytopharmaceutical product is its nomenclature. The problem of a rational nomenclature has been solved satisfactorily for pure products, both synthetic and extractive. In the case of vegetable drugs and preparations thereof the situation is somewhat different and, unfortunately, confused because there is no single universally accepted name for a vegetable drug or extract. This situation creates confusion and uncertainty: it prevents the comparative evaluation of pharmacological and clinical results obtained by different research teams and makes it impossible for the health authorities and for the physicians to compare phytopharmaceuticals containing the same active principles on the basis of rational and homogeneous names, thereby giving rise to errors in doses and prices. All this throws discredit on an entire field of great therapeutic and market importance. It is therefore necessary to draw up rules that will ensure clear and simple nomenclature for phytopharmaceuticals: I will make a few suggestions. Nomenclature of vegetable drugs A vegetable drug should be referred to solely by its correct botanical name followed by the name of the authority and by the part of the plant used. For example: Cinchona succirubru Pavon, bark. When a drug is used fresh, this must be specified because this factor is crucial for the composition of the vegetable drug. For example: Vaccinium m_~rtilius L., fresh fruits. If the constituents responsible for the therapeutic activity are known, their quantities must be stated. For example: Atropa belladonna L., leaves, containing . . .% of total alkaloids calculated as hyoscyamine. Extracts In the case of total extracts the name of the drug must be followed by the drug/extract ratio, by the solvent used for extraction and by the physical form (fluid, soft or dry). For example: Rhamnus purshiana DC., bark, l:l, 20%
hydroalcoholic fluid extract. Rhamnus purshiana DC., bark, 6:1, 60% hydroal~oholic dry extract. If the constituents responsible for the therapeutic activity are known, their quantity should be stated instead of the extract/drug ratio. If some other substance is added during the preparation of an extract to bring the content of active principles up to a given level, or for any other reason, the substance added must be reported as “other constituent(s)” and the extract (natural extract) as “active principle”.
-
Purified extracts The preparation of a purified extract includes phases of fractionation or purification (usually protected by patent), which are not simple operations of defatting but very important processes that increase the content of active principle appreciably beyond that of a total extract. The general nomenclature for total extracts cited earlier cannot be used for these extracts. A satisfactory approach might be to write the name of the vegetable drug followed by a term such as “anthocyanoside complex” or “liposoluble fraction (or complex)” or “total alkaloids”. Indication of the physical state may be unnecessary because these purified extracts are practically always dry extracts or, in the case of liposoluble fractions, oily products. The name of a purified extract must always be followed by the content of active principles. The trademark may be added in brackets, Examples: - Vaccinium myrtillus L., fresh fruits, anthocyanoside complex containing 35% anthocyanosides (~yrto~yan R). - Atropa belladonna L., leaves, 90% total alkaloids calculated as hyoscyamine. - Serenoa repens, fruits, liposoluble fraction, containing 90% of C6 to CzO linear chain fatty acids. These are only suggestions, certainly not 100% valid or acceptable: at all events, a rational nomenclature for phytopharmaceuticals is very important because it is the key to any comparison of activity, dosage and price.
