Kratom Study

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<H3 align=left>Clinical Study - Analgesia Induced by Mitragynine</H3></TD></TR>
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INTRODUCTION

In 1932, Grewal reported that on the isolated ileum the alkaloid from Mitragyna speciosa reduced the amplitude and tone of the smooth muscle. Perry (1980) quoted a report by Field and Quisumbing which stated that Mitragynine has a local anaesthetic effect and that the side-effects include dryness of the mouth, diuresis, constipation, with stools becoming small and dark, loss of appetite and subsequently a reduction in body weight. Jansen and Prast (1988) reported that the alkaloid from M.speciosa had opiate-like effects such as analgesia, antitussive and also cause hypothermia in animals. On the other hand, its action was not reversed by Nalorphine though it suppresses the opioid withdrawal syndrome.

It was thought that this novel analgesic action from the alkaloids of M. speciosa would be interesting to study since Nalorphine did not seem able to overcome its action (Idid et al., 1992). The alkaloid may be useful in the treatment of opiate addiction as a replacement therapy without the attendant dependence if it should act as an opioid agonist-antagonist.


METHODS

Preparation of Mitragynine
Mitragynine was extracted from fresh leaves of M. speciosa according to the method described by Ikram and Houghton (1986).

Preparation of Drug Solutions
Morphine sulphate, paracetamol and mitragynine were dissolved or suspended in 4% acacia gum.

Antinociceptive Assay
Three types of antinociceptive assays were employed in this study; these were i) Writhings test, ii)
Hot tail-flick and iii) Cold tail-flick test.

(i) Writhing tests The abdominal constriction test described by Collier et al. (1968) was used to measure the antinociceptive actions of morphine, mitragynine and paracetamol. Male or female albino mice weighing between 20-25 g were fasted for 24 hours with water given ad libitum, and were pretreated with oral 4% acacia gum solution (0. 15 ml/10 g), morphine (5 mg/kg), paracetamol(100 mg/kg) and mitragynine (200 mg/kg) 30 minutes prior to intraperitoneal injection of 0. 15 ml/l 0 g of 0.6% acetic acid to cause a typical stretching response. Writhings or stretchings (abdominal constrictions) were coun-ted for a period of 5 minutes under a double blind observation. The antinociceptive effects of drugs were measured by calculating the mean reduction in the number of abdominal constrictions for each drug, as compared to gum acacia controls.

(ii) Hot tail-flick Male or female albino mice weighing between 20 - 25 g were fasted for 24 hours with water given ad libitum, maintained at room temperature and were divided into 4 groups of six mice. Mice were treated with 4% acacia gum (0.15 m1/10g), morphine (5 mg/kg), paracetamol (100 mg/kg) and mitragynine (200 mg/kg). Antinociceptive effect of the test substances was determined by the hot tail-flick method described by Sewell and Spencer (1976). One to two cm of the tail of mice was immersed in warm water kept constant at 50 °C. The reaction time was the time taken by the mice to deflect their tails. The first reading is discarded and the reaction time was taken as a mean of the next two readings. The latent period of the tail-flick response was taken as the index of antinociception and was determined before and at 15, 30, 45 and 60 min after the administration of drugs. The maximum reaction time was fixed at 15 seconds. The maximum possible analgesia (MPA) was calculated as:

Test reaction time - Saline reaction time
MPA =15- Saline reaction time

(iii) Cold tail-flick Male or female albino mice weighing between 20 - 25 g were fasted for 24 hours with water given ad libitum, maintained at room temperature and were divided into 4 groups of six mice. They were then given the same treatment as in the writhing test. The reaction time of the mice was measured at 15, 30, 45 and 60 min. The antinociceptive activity was determined by the cold tail-flick test described by Pizziketti et al. (1985). One to two cm of the tail of mice was immersed in a cold 1: 1 mixture of water and ethylene glycol kept constant at -10 °C. The reaction time was the time taken by the mice to deflect their tails. The first reading is discarded and the reaction time was taken as the mean of the last two readings.

ASEAN Review of Biodiversity and Environmental Conservation (ARBEC) May 1998
The maximum reaction time was fixed at 30 seconds . The maximum possible analgesia (MPA) was calculated as:

Test reaction time - Saline reaction time
MPA = 30- Saline reaction time


STATISTICAL ANALYSIS

The experimental data were analyzed by two-tailed impaired student's ttest. P &lt;0.05 was considered significant. Data were presented as mean ±SEM of n observations.


RESULTS AND DISCUSSION

Acetic Acid-induced Writhings
Intraperitoneally injected acetic acid produced abdominal constrictions which is characterized by a stretching response. Mean writhings observed in control treated with vehicle (acacia gum) animals over a period of five minutes was 17.5 + 2.8 counts. Morphine (5 mg/kg) and mitragynine (200mg/kg) significantly (p&lt;0.05) reduced writhings to 7.3 + 0.6 and 9.6 + 0.6 counts, respectively (Figure 1). On the other hand, paracetamnol (100 mg/kg), did not significantly reduce writhings induced by acetic acid.
This test also showed that 5mg/kg of morphine is about equipotent to 200mg/kg of the crude extract. Other investigators have shown that the cold tail- flick method is a selective method able to screen centrally acting opiate-fike analgesic agents, and is not sensitive to analgesics acting peripherally, such as aspirin, or non-analgesic drugs acting on the central nervous system, such as chlorpromazine (Pizziketti et al., 1985).

On the other hand, the hot tail-flick method is shown here as incapable of differentiating between opiate and non-opiate analgesics, or between peripherally acting or centrally acting substances. It is possible that the induction of analgesia by the alkaloid has both a peripheral and a central component. In the report of Jansen and Prast (1988) they found that nalorphine did not antagonize the effect caused by alkaloid from M speciosa.

Hot Tail-flick Response
Throughout the 60 min observation, animals pretreated with acacia gum did not show significant effect on the latent period of tail-flick response. The antinociceptive effect of morphine was evident within 15 min following oral administration. The mean possible analgesia (MPA) increased from 3.9 ± 3.7 to 35.1 + 4.8 % which remained elevated above the basal levels throughout the observation period (Figure 2 and Table 1). Likewise, paracetamol also exhibited significant antinociception which began at 15 min following oral administration and the effect remained significant throughout the 60 min observation period (p&lt;0.05). The MPA calculated for paracetamol increased to 27.8+ 2. 1 %. Similarly, the antinociceptive effect of mitragynine was also observed at 15 mins following oral administration and the effect remained significant throughout the 60 min observation period. The MPA calculated for mitragynine increased to 42.8 + 7.2

Cold Tail-flick Response
While animal groups treated with gum acacia and paracetamol showed no significant effect on the latent period of tail-flick response, the antinociceptive effect of morphine was evident only after 30 min, reaching its peak at 45 min and this effect remained significant until the 60 min (p&lt;0.05) test period. The peak MPA value calculated at 45 min was 66.2 ± 2.4 % compared to 0.1 + 7.2 % induced by the control acacia gum (Figure 3, Table 1). On the other hand, the antinociceptive effect of mitragynine was evident after 15 min reaching its peak at 30 min and this effect remained significant until the 45 min (p&lt;0.05) test period. The peak MPA calculated at 30 min was 49.0 ± 5.9 %.

Use In Traditional Medicine
Reports of the various uses of Mitraynine in traditional medicine in the countries of the South East Asia region (Jansen & Prast, 1988; Suwanlert, 1974) spurred research into the pharmacological aspects of its chemical constituents. More than twenty-five alkaloids have been identified from this plant (Ikram, 1885) but of these the major ones (in terms of % yield) are three indoles i.e. mitragynine, speciogynine and paynanthine and two oxindoles i.e. mitraphylline and speciofoline.

In a previous study (unpublished report) we had detected the analgesic property of the crude alkaloidal extract using the hot-plate method of Ankier (1974) and the tail-flick method of Sewell and Spencer (1976). The effect showed similarities to analgesia caused by morphine but its effect was not antagonized by Naloxone in the hot-plate test. This is interesting because although paracetamol also exhibited a significant effect in the hot tail-flick test, our additional experiment using the cold tail-flick test further separated the analgesic efficacy of the alkaloidal extract and morphine from that of paracetamol. The failure of paracetamol to exhibit antinociceptive effect in the cold tail-flick test suggests that analgesia induced by morphine and the extract is by a different pathway involving other mediators than that due to paracetamol. This test also showed that 5 mg/kg of morphine is about equipotent to 200mg/kg of the crude extract. Other investigators have shown that the cold tail- flick method is a selective method able to screen centrally acting opiate-fike analgesic agents, and is not sensitive to analgesics acting peripherally, such as aspirin, or non-analgesic drugs acting on the central nervous system, such as chlorpromazine (Pizziketti et al., 1985).

On the other hand, the hot tail-flick method is shown here as incapable of differentiating between opiate and non-opiate analgesics, or between peripherally acting or centrally acting substances. It is possible that the induction of analgesia by the alkaloid has both a peripheral and a central component. In the report of Jansen and Prast (1988) they found that nalorphine did not antagonize the effect caused by alkaloid from M speciosa.

Although it is recognized that the acetic acid writhing test is a useful measure of analgesic activity, the method often shows positive responses even to non-analgesic compounds such as central nervous system depressants (Hendershot and Forsaith, 1959). Consistent anti-nociceptive activity of the M.speciosa extract in all three methods used in the present study is in contrast with the typical antinocipceptive activity exhibited by paracetamol, which shows significant activity only in the hot tail-flick test. The failure of paracetamol to inhibit acetic acid induced writhings may imply that suppression of writhings may only be achieved by analgesic agents of higher efficacy or at higher doses of paracetamol. Antinociceptive effects of morphine and the alkaloidal extract were evident from the fact that both of these substances were markedly effective in reducing the responses in all three tests viz the writhing and the tail-flick tests. Thus the alkaloid from M.speciosa has definite analgesic properties but further study must be carried out to determine in which particular alkaloid this effect resides.

Our present study indicates that the alkaloidal extract has analgesic properties similar to opioid and, similar to morphine is effective in the acetic-acid writhing test, the hot tail-flick and the cold tail-flick tests. We have also shown that aracetamol is only effective in the hot tail-flick test.


ACKNOWLEDGEMENT

We wish to thank Miss Christina Gertrude Yap from the department of Pharmacology, UKM and Miss Nancy Yeap from the department of Pharmacology, Universiti Malaya for their assistance.


REFERENCES

Ankier, S.I. 1974. New hot-plate test to quantify antinociceptive and narcotic antagonist. Eur. J Pharmacol. 27: 1-4
Collier, H.O.J., L.C. Dinneen, C.A. Johnson and C. Schneider 1968. The abdominal constriction response and its suppression by analgesic drugs in the mouse. Br. J Pharmacol. Chemother. 32: 295.
Grewal, K.S. 1932 Observations on the pharmacology of Mitragyna. In: Pharmacology of mitragynine. J Pharmacol. Exp. Ther. Vol XLVI(3): 251-271.
Hendershot, L.C. and Forsaith, J. 1959 Antagonism of the frequency of phenylquinone-induced writhing in mouse by peak analgesics and nonanalgesics. J Pharmacol Exp. Ther., 125: 23 7-24 1.
Houghton, P.J & 1kram M. Said. 1986. 3- Dehydromitragynine: an alkaloid from Mitragyna speciosa. Phytochemistry 25: 2910-2912.
Idid, S.Z.,, K. Norehan and A. Roslan. 1992. The involvement of the noradrenergic system in analgesia induced by the alkaloidal extract of Mitragyna speciosa in the rat. Proc. 3rd Medical Colloquim, UKM. pp 337-340.
Ikram, M.S. 1985. Studies on the components of fresh leaves of Mitragyna speciosa. In: M.S.
lkram & Z.Zakaria (Eds.) Proceedings of 2nd Meeting of the Natural Products Research Group, Chemistry Dept., UKMalaysia. pp 123-127.
Jansen K.L.R. and C.J.Prast, 1988. Ethnopharmacology of Kratom and the Mitragyna alkaloids. J Ethnopharmacology 23: 115-119,
Perry, L.M. 1980. Medicinal plants of East and South East Asia, MIT Press, U.S.A.
Pizziketti, R.J., N.S. Pressman, E. B. Geller and M.W. Adler 1985. Rat cold water tail-flick: A novel analgesic test that distinguishes opioid agonist from mixed agonist-antagonist. Eur J Pharm. 119: 23-29.
Sewell, R.D.E. and P.S.J. Spencer 1976. Antinociceptive activity of narcotic agonist and partial agonist analgesics and other agents in the tail-immersion test in mice and rats.
Neuropharmacol. 15, 23-29.
Suwanlert, 1974. A study of kratom eaters in Thailand. Thai Bull. on Narcotics. 26: 21-27.</TD></TR></T></TABLE></TD></TR></T></TABLE>

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The leaves of a tropical plant, Mitragyna speciosa KORTH. (Rubiaceae), have been traditionally used as a substitute for opium. By phytochemical studies on the constituents of the plant growing in Thailand as well as in Malaysia, several 9-methoxy-Corynanthe-type monoterpenoid indole alkaloids including new natural products were isolated. The structures of these new compounds were elucidated by the modern spectroscopic methods and/or chiral-total syntheses. The chiral total synthesis of (−)-mitragynine, a major component of this plant, was achieved. Potent opioid agonistic properties of mitragynine, which acts on ì- and ä-opioid subtype receptors, and of mitragynine pseudoindoxyl, whose analgesic activity is more potent than that of morphine, were clarified in in vitro experiments. The essential structural features in mitragynine for revealing the analgesic activity were elucidated by pharmacological evaluation of the natural and synthetic mitragynine derivatives.

Mitragynine (1) is a major alkaloidal component in the Thai traditional medicinal herb, Mitragyna speciosa, and has been proven to exhibit analgesic activity mediated by opioid receptors. By utilizing this natural product as a lead compound, synthesis of some derivatives, evaluations of the structure-activity relationship, and surveys of the intrinsic activities and potencies on opioid receptors were performed with guinea pig ileum. The affinities of some compounds for mu-, delta-, and kappa-receptors were determined in a receptor binding assay. The essential structural moieties in the Corynanthe type indole alkaloids for inducing the opioid agonistic activity were also clarified. The oxidative derivatives of mitragynine, i.e., mitragynine pseudoindoxyl (2) and 7-hydroxymitragynine (12), were found as opioid agonists with higher potency than morphine in the experiment with guinea pig ileum. In addition, 2 induced an analgesic activity in the tail flick test in mice.
Mitragynine, an indole alkaloid from Thai folk medicine Mitragyna speciosa, exerts agonistic effects on opioid receptors. Gastric acid secretion is proposed to be regulated by opioid receptors in the central nervous system (CNS). Previously, we reported the dual roles (inhibition via micro-opioid receptors and stimulation via kappa-opioid receptors) of the opioid system in the central control of gastric acid secretion. We investigated whether mitragynine affects gastric acid secretion via opioid receptors in the CNS. Injection of mitragynine (30 microg) alone into the lateral cerebroventricle did not have a significant effect on basal gastric acid secretion in the perfused stomach of anesthetized rats. Injection of mitragynine (3-30 microg) into the fourth cerebroventricle, like morphine, inhibited 2-deoxy-D-glucose-stimulated gastric acid secretion. The inhibitory effect of mitragynine (30 microg) was reversed by naloxone (100 microg). These results suggest that mitragynine has a morphine-like action on gastric acid secretion in the CNS.
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This recipe makes enough tea for eight to ten mild doses, four to six medium strength doses, two moderately strong doses, or one very strong dose.


STEP 1: Take 50 grams of dried, crushed kratom leaves and put into a pot. To this add 1 liter of water.


STEP 2: Boil gently for 15 minutes.


STEP 3: Pour the tea through a strainer into a bowl and reserve the liquid. Squeeze the leaves in the strainer to get most of the liquid out.


STEP 4: Put the leaves back in the pot and add another liter of fresh water. Repeat steps 2 and 3.


STEP: 5: After the leaves have been strained a second time, they can be discarded. Put the combined liquid from both boilings back into the pot and boil until the volume is reduced to about 100 ml.


Kratom tea can be safely stored in the refrigerator for about 5 days. It is probably okay to keep it a bit longer, but it's better to play it safe and not drink it after 5 days. It can be stored for many months if you add some alcohol to it. Adding about 10% alcohol will preserve it for many months (in the refrigerator). That is 1 part 80 proof vodka (or a similar spirit) to 3 parts kratom tea.


The same general preparation method can of course be used with larger or smaller amounts of herb by simply adjusting the volume of water used. Some people experience mild nausea when using doses above 25 grams, so when taking larger doses, it is best to do so on an empty stomach (i.e. wait about 3 hours after eating).</TD></TR></T></TABLE></TD></TR></T></TABLE>

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Kratom is indigenous to the rain forests of South East Asia, mainly Thailand and Malaysia. It is a large tree that can reach 15 meters. Over 25 alkaloids have been isolated from kratom, the most important being mitragynine.
In Thailand it was used in many dishes, before it was banned by the governement.
In addition to being used as a narcotic drug in its own right, it is often used as a substitute for opium when opium is unavailable, or to moderate opium addiction. As mitragynine suppresses opiate withdrawal, kratom is said to be a (temporary) cure for opium addiction. A small minority of users use kratom to prolong sexual intercourse.
Effects come on within five to ten minutes after use, and last for several hours. Kratom has both stimulating and depressant qualities as if chewing coca leaves and smoking opium simultaneously. Some sources say that it is a stimulant in lower doses, becoming sedative in higher doses. The dominant effects seem to be similar to opiate drugs, including analgesia, roughly comparable in strength to codeine. Unlike opiates, mitragynine does not appear to cause nausea or vomiting. The feeling has been described as happy, strong, and active, with a strong desire to do work. Other effects of mitragynine are local anesthesia and central nervous system depression. Heavy use can result in a prolonged sleep.
The leaves of kratom are grinded and eaten, usually followed by drinking warm water, tea or coffee. Leaves can also be made into a tea, or a crude resin extraction can be made. This resin extract is made by preparing a water extract of the leaves, boiling it down, and then shaping it into small ball.

While new users may only need 5-10 grams of leaves to obtain the desired effects, some users find with time they need to increase doses, up to 50 grams leaves per day for a strong effect. It is best to take the leaves on an empty stomach.
Usage of kratom in high dosages may be mildly addictive. Acute side effects include dry mouth, loss of appetite and constipation. Side effects from long term use include anorexia and weight loss, insomnia, and a darkening of the skin, particularly on the cheeks.
Do not combine wit MAO-inhibitors.