The physical and chemical features of Cannabis plants grown in the United Kingdom of Great Britain and Northern Ireland from seeds of known origin

Sections

ABSTRACT
Introduction
Experimental
Conclusions

Details

Author: P. B. BAKER, T. A. GOUGH, B. J. TAYLOR
Pages: 27 to 36
Creation Date: 1982/01/01

The physical and chemical features of Cannabis plants grown in the United Kingdom of Great Britain and Northern Ireland from seeds of known origin

P. B. BAKER
T. A. GOUGH
B. J. TAYLOR
Laboratory of the Government Chemist, Cornwall House, London, United Kingdom of Great Britain and Northern Ireland

ABSTRACT

Cannabis plants have been grown in the United Kingdom of Great Britain and Northern Ireland from seeds taken from seizures of cannabis of known geographical origin and chemistry. The gross physical appearance and cannabinoid patterns of many of the cannabis samples produced in the United Kingdom were closely related to those of the parents. However, some notable exceptions were recorded. There were wide variations in actual tetrahydrocannabinol content between plants grown from different seedstock and rather smaller variations within the groups grown from the same seedstock. Cannabis produced in the United Kingdom had higher tetra-hydrocannabinolic acid/tetrahyrocannabinol ratios than imported material.

Introduction

In a study of the physical and chemical features of imported Cannabis 1products, Baker and others [ 1] found that the gross physical appearance and the cannabinoid pattern as revealed by thin-layer chromatography (TLC) correlated with the geographical origin for most of the samples examined. Consequently, if these features of a sample of unknown origin were compared with those of samples of known origin, an opinion as to the geographical origin of the unknown sample could be formed. Such information may be useful in criminal investigations relating to illicit Cannabisproduct importations and may aid the international control of the substance [ 2-5] . It may also be of considerable importance to know whether a seizure made within the United Kingdom was of foreign origin or whether the material was grown within the United Kingdom. It is thus necessary to study the physical and chemical features of Cannabis grown in the United Kingdom and in particular, to examine the cannabinoid patterns of Cannabis plants grown in the United Kingdom from seeds which are not only of known geographical origin but which are also derived from imported cannabis samples of known cannabinoid pattern and content.

Seeds from which Cannabis can be grown in the United Kingdom may be derived from three principal sources: ( a) from seed sold within the United Kingdom for fish bait or bird seed; ( b) from imported cannabis samples; and ( c) from fertile seeds produced from plants cultivated in the United Kingdom. Seeds sold legally within the United Kingdom have been shown to have poor germination rates [ 6] and are therefore unlikely to be used widely by those engaged in illicit Cannabis cultivation. The majority of seizures of cannabis examined at the Laboratory of the Government Chemist, London, contained fertile seeds. These were, however, unlikely to have been incorporated into cigarettes as they had negligible or zero cannabinoid content [ [ 6] , [ 7] ] and they thus provided a ready source of fertile seeds for cultivation. It was also necessary to study the physical and chemical features of plants grown from fertile seeds which themselves were produced in the United Kingdom.

The term Cannabis in this paper refers to Cannabis sativaL.; cannabis to marijuana; and cannabis resin to hashish.

A limited number of studies of Cannabis plants grown in the United Kingdom from seeds of known origin have been made. De Faubert Maunder [ 8] made a comparative study, using TLC, of the tetrahydrocannabinol (THC) content of an imported Nigerian cannabis seizure with plants grown in the United Kingdom from seeds contained therein. The original seizure and the siblings all showed the same basic cannabinoid pattern, namely absence of cannabidiol (CBD) and the presence of THC. Although TLC is essentially non-quantitative, de Faubert Maunder estimated that the siblings all contained less THC than the parent. A second study of plants grown from seeds derived from a seizure of South African cannabis gave similar results. Fairbairm and others [ 9] grew plants from seeds of known origin (although the cannabinoid pattern was not published) and examined the cannabinoid contents of the resulting plants. A Nepalese strain grown in different years showed different CBD and THC contents, but very similar CBD/THC ratios. A Turkish strain contained principally CBD with no THC content reported, and a Thai strain contained principally THC with a small amount of CBD. The authors recorded that the cannabinoids occurred in the plants principally as their acids. These same authors also observed [ 10] that conditions of growth affected the cannabinoid content by showing that plants grown in greenhouses had different THC content from plants grown from the same seed in the open. Care was taken in sampling as previous work had shown [ 9] that different parts of the same plant had differing canna-binoid contents.

Fairbairn and Liebmann [ 11] grew 12 strains of Cannabis in the open in southern England and found considerable variations between plants of the same strain grown together and harvested at the same time. Considerable variations within plants were once again noted. However, the basic cannabinoid ratios remained the same in plants of the same strain despite these quantitative variations. Results from other groups of workers who had grown plants from the same batch of seeds in different parts of the world indicated that the same qualitative picture of cannabinoid pattern was always exhibited [ [ 7] , [ 12] , [ 13] ]. On the basis of these observations, Fairbairn and Liebmann [ 11] suggested that there may be two chemical races of Cannabis plants, one being THC-rich and the other CBD-rich. They considered that this ratio was independent of environmental conditions, but qualified their opinion by suggesting that several generations of plants should be grown to confirm this.

Rowan and Fairbairn [ [ 6] , [ 14] ] presented further evidence for the existence of two chemical races of Cannabis in a study of the cannabinoid patterns of Cannabis seedlings. THC-rich and CBD-rich patterns were shown to be established when plants were small. In the THC-rich strain, cannabichromene (CBC) was present, whereas in the CBD-rich strain, CBC was absent.

There have been a number of studies of Cannabis plants grown from seeds of known origin in other parts of the world. Fetterman and others [ 7] in a study of Mexican and Turkish seeds grown in Mississippi, United States of America, proposed that Cannabisplants fell into two groups which were termed phenotypes, corresponding to fibre-producing and to drug-producing plants. These authors suggested that the phenotypes should be classified as phenotype I (drug-type) if the ratio (% THC + % CBN)/(% CBD) was greater than 1.0 and phenotype II (fibre-type) if this ratio was less than 1.0. All plants of known origin and history could be classified using this system; only plants of unknown history could not be so classified and the authors suggested that these samples could have contained mixtures of different types of cannabis. From their data, the authors concluded, the phenotype of one variety of cannabis remained the same regardless of the year of planting or place of growth of the sample analysed. In a related study, Doorenbos and others [ 15] found most samples to have a phenotype ratio greater than 5 or less than 0.2. They concluded, as a result of their studies of cannabinoid content of plants of the same variant grown in different countries and plants grown through three generations, that environmental factors were not as important as heredity in determining cannabinoid patterns of Cannabis. The authors noted the different cannabinoid contents in plants of the same variant grown under different conditions and that most of the cannabinoids were present as their acids. Ohlsson and others [ 16] , in a study of Cannabis plants grown in Sweden, concluded that the type of cannabinoid produced by the plant depended on the seed and that the influence of climate was limited. Turner and Hadley [ 17] grew three generations of plants from seeds originating in South Africa and noted some fluctuation in cannabinoid content, particularly an increase in cannabigerol monomethyl ether (CBGM) percentages. The authors point out that no major attempt was made to avoid cross-pollination and so the significance of the increase in CBGM content (a cannabinoid in plants from north-east Asia [ 18] ) cannot be properly assessed. Small and Beckstead [ 12] grew plants from 350 different seeds at Ottawa, Canada, and increased the number of phenotypes by the addition of a third group where CBD/THC = 1.0 and a fourth group from north-east Asia containing CBGM. These authors correlated phenotype with the latitude of the seed source as follows: phenotype I from south of 30°N had THC > 0.3% and CBD < 0.5% and female plants similar in cannabinoid content to male plants; phenotype II had THC > 0.3% (in female plants) and CBD > 0.5% and originated from north of 30°N with female plants of much higher cannabinoid content than males; phenotype III had THC < 0.3% and CBD > 0.5%, but was otherwise similar to phenotype II, and phenotype IV contained CBGM. The authors considered that climate might affect the actual amount of THC in a plant, with a more southerly location meaning a longer growing season resulting in most high-THC (drug-type) plants being produced from seeds originating from south of 30°N. Krejci and others [ 19] demonstrated that seeds from Turkish fibre-producing plants, even when grown under favourable conditions did not produce appreciable amounts of THC and that seeds derived from South Africa and Thailand (drug-type), even when grown under unfavourable conditions, still produced relatively large amounts of THC. Boucher and others [ 20] grew South African Cannabis in France and observed that there were two types, either tetrahydrocannabinolic acid (THCA)-rich or tetrahydrocannabinvarinic acid (THVA)-rich, an observation also recorded by Field and Arndt [ 21] and Baker and others [ 1] . After three generations, however, all the THVA-rich plants had become THCA-rich, indicating that THCA/THVA ratios might be under environmental control. Both Novotny and others [ 22] and Turner and others [ 23] suggested that minor cannabinoids might be liable to variations when plants were grown in different environments.

As a result of the studies described above, the majority of workers have concluded that although actual THC content, and possibly minor cannabinoid content, may be under environmental control, the ratios of the principal cannabinoids CBD and THC and the absence of the former in some Cannabissamples [ 1] were controlled by the genetic material in the seed. However, there was a need for multigeneration growing trials before this opinion could be confirmed [ 11] .

Although the geographical origin of the seeds was known in many of the studies described, in no case had the parent material (as produced before export) been adequately analysed in order that changes (if any) produced by growing seeds in a totally alien environment could be observed. It was therefore considered to be both necessary and useful to grow plants from seeds whose origin was known and whose parent chemistry could be studied in detail. The Laboratory of the Government Chemist was fortunate in possessing a large number of cannabis samples of known origin containing fertile seeds which were used as a basis for a study of Cannabis grown in the United Kingdom.

Experimental

Seed selection

Seeds were taken from illicitly imported samples of cannabis of known origin. The country of origin was assigned by taking into account information from the carrier, from Officers of Her Majesty's Customs and Excise and from the route of importation. The cannabis samples were chosen for having particularly characteristic cannabinoid patterns or content which might be easily followed through subsequent generations. Although homogeneity of seeds within a seizure could not be completely guaranteed, the homogeneity of seizures as regards cannabinoid pattern made it a reasonable assumption that seeds would also be homogeneous.

Growing conditions

The first generation of United Kingdom grown plants of Indian and Thai origin were cultivated indoors in a poorly lighted and unheated room in 1979. The second generation of the above plants, together with the first generation of the others listed in table 1 were

Table 1

Country of origin and chemical features of seedstock Cannabis

Country of origin	Cannabinoid content	CBD % w/w	THCA % w/w	THC % w/w	THCA/THC
Morocco	CBD, THC and low THV b	0.8	1.14	0.37	3.1
Sri Lanka	High CBD, THC and low THV	1.1	0.33	0.74	0.5
Swaziland	THC > THV, no CBD	ND c	0.84	0.87	1.0
Thailand a	THC and THCA the only major components in both parent and first	ND	0.9	<0.1	>9.0
India a	generation plants	ND	1.46	<0.1	>15.0
Zambia	THV > THC, no CBD	ND	1.37	1.1	1.2
Zimbabwe	THC = THV, no CBD	ND	5.1	0.63	8.1

^aSecond generation

^bTHV = tetrahydrocannabivarin

^cND = not detected.

grown in greenhouses in south-east England during 1980. The greenhouses were gently heated during October and November. As far as possible conditions for growing each plant were identical as attention had been drawn to such factors as plant pot size [ 24] , cultivation conditions [ 25] , light intensity [ 11] , plant maturity [ 24] and other local environmental factors. In as far as it was possible, all plants in this study were grown under identical conditions in order that any differences observed between the parent cannabis sample and the sibling plants might be ascribed to the environmental differences between the United Kingdom and the country of origin of the seedstock. In this study the term seedstock means the batch of seeds taken from a particular importation of cannabis.

Seeds were planted at the end of February and the beginning of March in 75 mm pots filled with compost (Levington's Fisons Limited, Levington, Ipswich, Suffolk, England). Seedlings were transplanted to 200 mm pots filled with the same compost in early April. From the beginning of July until harvesting, plants were given a weekly liquid feed (Bio Plant Food, Pan Britannica Industries Limited, Britannica House, Waltham Cross, Hertfordshire, England). As soon as flowering commenced, plants were separated according to sex and were carefully pollinated by hand to ensure that both parents of fertile seeds were from the same original seedstock. Plants were harvested at maturity, the dates are listed in table 2, except for those which showed signs of early distress. The dates of maturity fell into two groups, July and August, and November. Plants from Moroccan seed matured much earlier than all others. There seemed to be little correlation between the longitude of the plant source and the date of maturity in the United Kingdom. Nordal and Braenden [ 13] remarked on the variations in the size and shapes of plants raised from different seeds in Norway. We were also able to note several consistencies in appearance within each country as well as certain notable differences. Moroccan plants were, on the whole, shorter and bushier than other plants, their height being limited by their short growing period before maturity. Thai plants were consistently tallest while Swazi plants were initially rather slow growing. The central stems of Zambian plants were very thick and red stems were notable on some Zimbabwean plants. The plants were harvested and the leaves and flowering parts of the plants were separated from the woody parts of the plants and air-dried. Any remaining large stalks were removed before weighing the product from each plant. Female plants, in general, yielded a greater weight of cannabis within countries, although one Indian male plant which did not mature until November produced more cannabis than its female and male or female counterparts which matured much earlier. The yields of cannabis and the sex of the plants reared to maturity are recorded in table 2.

Analysis

Samples of both the parent cannabis and that grown from its seedstock in the United Kingdom were analysed by TLC [ 26] , gas-liquid chromatography (GLC) [ 27] and high performance liquid chromatography (HPLC) [ 28] . Parent cannabis seizures were quantitatively analysed when fresh, and as received, except that large stems and stalks were removed before the extraction stage of the analyses. The United Kingdom grown samples were analysed after removal from the plant as described above. TLC provided a simple and rapid method of assessing the relative proportions of the principal cannabinoids in each sample. In view of the importance of cannabinoid pattern (as distinct from actual content) in the assessment of origin [ [ 1] , [ 27] ], quantitative data may only confuse the picture. However GLC was used to determine "total" THC (i.e. the amount of THC together with its acid), and HPLC was used to determine "actual" THC.

Table 2

Data on Cannabis plants grown in 1980

Country of origin	plant number	Harvesting date	Sex	Weight of cannabis (g)	Cannabinoid content	CBD % w/w	THCA % w/w	THC % w/w	THCA/THC
India	1	August	F a	21.1	As parent	ND f	5.8	0.03	194
	2	November	M b	39.3	As parent	ND	4.5	0.08	57
	3	August	F	19.0	As parent	ND	3.3	0.16	21
	4	August	M/F	16.4	As parent	ND	3.9	0.43	9
	5	July	M/F	17.1	CBD, CBDA d,THC, low THV	1.58	0.7	0.02	37
Morocco	1	July	F	46.2	THC, THCA a, low THV, CBC, no CBD	ND	2.9	0.52	6
	2	June	M	11.4	As parent	0.47	0.3	0.03	9
	3	July		75.0	As Morocco 1	ND	5.1	0.25	21
Sri Lanka	1	August	F	47.9	As parent	2.76	0.1	0.05	3
	2	July	M	37.3	THC, THCA, low THV, CBC	ND	3.0	0.09	33
	3	August	F	52.0	As parent	1.79	0.1	0.02	7
Swaziland	1	August	M	28.4	As parent	ND	0.6	0.05	13
	2	July	M	27.7	THC, THCA, orange spot at THV Rf	ND	1.9	0.05	39
	3	August	M	59.1	As parent	ND	1.5	0.05	31
Thailand	1	November	F	18.1	As parent	ND	7.1	0.06	117
	2	November	F	32.7	As parent	ND	4.3	0.05	85
	3	July	Imm c	23.6	As parent	ND	3.5	0.09	39
	4	November	F	24.6	As parent	ND	3.2	0.05	63
	5	July	Imm	14.5	As parent	ND	3.5	0.06	58
Zambia	1	November	F	27.7	As parent	ND	0.6	0.01	60
	2	November	M	20.1	As parent	ND	1.1	0.01	110
	3	November	F	26.7	As parent	ND	0.8	0.01	80
Zimbabwe	1	August	Imm	28.9	As parent	ND	0.7	0.15	4
	2	November	F	27.6	As parent	ND	1.3	0.13	10
	3	November	M	10.9	As parent	ND	1.1	0.09	13

^aF = female plant

^b M = male plant

^c Imm = Immature plant

^d CBDA = cannabidiolic acid

^e THCA = tetrahydrocannabinolic acid

^f ND = not detected.

The amount of THCA was then determined as previously described [ 29] . All samples were analysed when fresh (i.e. upon seizure or immediately after harvesting).

Results and discussion

The United Kingdom products after harvesting were in all cases green or yellow-green friable materials. This was not in some cases dissimilar (and without hindsight not distinguishable) from the original imported cannabis. However, imported material was generally darker in colour and the samples from which Indian and Zimbabwean seeds were selected were quite brown.

Most of the cannabinoids were present as their parent acids. TLC indicated that the majority of United Kingdom grown samples were, at least in cannabinoid pattern, very similar to the sample from which their seeds were taken. However, 5 of the 25 harvested plants showed very different cannabinoid patterns from their parent cannabis samples; results for all the plants are listed in table 2. All groups produced at least one plant which was similar to the parent material and all the Indian, Zambian and Zimbabwean plants were similar to their parents. Although all seeds for a given country came from the same seizure of cannabis, it is possible that these seizures, however small, were not homogeneous. This might explain the patterns found in Sri Lanka 2 and Swaziland 2, the pattern of the latter being characteristic of some samples of South African cannabis [ 1] . In four of the five plants which were very dissimilar to their parents, CBD present in the parent did not occur in the sibling plant or vice versa. In the case of India 5, CBD did not occur in any of the plants grown in 1979 in the United Kingdom nor in the original imported material.

Quantitative data

In general, and as expected from the TLC data, HPLC patterns were similar for plants from one source, other than the major exceptions discussed above. Tables 1 and 2 record the quantitative data for the seedstock cannabis and the United Kingdom grown material respectively. It is clear from these data that good quality cannabis can be produced in the United Kingdom, a result in accordance with that of Fairbairn and Liebmann [ 11] , much of it as good as fresh imported material [ 27] . When it is considered that only 15% of all seizures of Cannabis products examined in this Laboratory are fresh at time of analysis [ 27] , the quality of United Kingdom cannabis becomes of much greater significance. Although there is a broad similarity in the actual amount of THC in plants of the same country, there are quite wide variations. Those plants (India 5, Morocco 2 and Sri Lanka 1, 3) which contain CBD have lower amounts of THC than other plants from the same seedstock. It is of interest that both the Indian and Thai first generation plants had low THC contents (see table 1) whereas the second generation plants had much higher THC content. It may be that the low content of the first generation plants is a reflection of the cold and rather dark conditions under which these first generation plants were grown.

A study of the ratio of THCA/THC in the harvested plants showed that, in general for the United Kingdom material, this had a much higher value than for imported material [ 29] . Over all major supplies of fresh cannabis to the United Kingdom in 1979 (64 examined), the average THCA/THC ratio was 2.4 (s = 2.2). However, for the 22 samples grown in the United Kingdom, this ratio averaged 40 (s = 46). Mechoulam [ 30] noted that Cannabis grown in northern countries possessed little if any neutral cannabinoids, whereas that grown in tropical or subtropical countries underwent at least some decarboxylation in the plant. Mechoulam further considered that the method of illicit preparation caused further decarboxylation.

Conclusions

From these studies, it was clear that cannabis prepared from at least the first generation of plants grown in the United Kingdom from imported Cannabis seeds could bear a close physical resemblance to the original material. Further, the cannabinoid patterns of seedstock plant material and siblings were similar in most cases, although there were some notable exceptions. The "total" THC content showed no distinct trends, but there was no doubt that it was possible to produce good quality cannabis in the United Kingdom. The ratio THCA/THC was significantly higher in the samples of United Kingdom cannabis than in imported samples: determination of this ratio for samples of unknown provenance might provide the analyst with an indication as to whether a sample had been imported, although it would be necessary to study a much greater range of plants before such a generalization could be substantiated. Further generation studies are being undertaken to monitor any gradual changes in plant characteristics.

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