Extraction hypericins

Another interesting development is the use of the naturally occurring herbal extract hypericin (4.35), which has the extended quinone structure (4.35), in both photodiagnosis and therapy. This is given to the patient, either orally or topically, who is then illuminated with blue hght. The cancer tumours show up as red spots, the hght from which can be recorded on a red sensitive camera, subjected to computational analysis and then converted into an image on the computer. 

It should first perhaps be mentioned that pressurized cold water has been successful in plant extraction. Hypericin, protohypericin, pseudohypericin and protopseudohypericin have been extracted from St. John s wort (Hypericum perforatum)... 

Fig. 1 Chromatography of flavonoids. 1. Extract. Solidaginis 2. Rutin — chlorogenic acid — isoquercitrin — quercitrin 3. Extract. Hyperici 4. Hyp>erosid — quercetin-3-arabinosid — hypericin — quercetin 5. Extract. Betulae...

Hydroxytryptamine 380 5-Hydroxytryptophan 240,241 Hydroxytyramine 392 Hyodesoxycholic acid 334 Hyperici, Extract. 279 Hypericin 148, 279, 280 Hyperoside 149,279,323 -, -quercetin 280... 

It is being recognized increasingly that regulation can have a positive impact on laboratory productivity.36 System suitability testing has been proposed as superior to and supplemental to calibration in the UV-VIS detector.37 Large variations in both response factor and in relative response factors were observed on different instruments. Even on the same instrument, UV-VIS spectra can be extremely dependent on solution conditions, as was observed in a separation of hypericin, the antidepressant extract of St. John s wort.38... 

Fig. 2.49. Profile of Hypericum perforatum extract with the H LC-MS attributions of the components detected. 1 = chlorogenic acid isomer 2 = 3-0- -coumaroylquinic acid 3 = chlorogenic acid 4 = rutin 5 = hyperoside 6 = isoquercitrin 7 = 3,3, , , 7-pentahydroxyflavanone 7-0-rhamnopyranoside 8 = quercitrin 9 = quercetin 10 = 13,118 tapigenin 11 = pSeudohypericin 12 = hypericin 13 = hyperforin analogue 14 = hyperform dialogue 15 = hyperforin 16 = adhyperforin. Reprinted with permission from M. Brolis eta. [ ].

Extracts of hypericum may vary considerably in terms of the quantity and ratio of their constituents based on the extraction process used. Maximum extraction of hypericin and pseudohypericin is obtained with an 80% methanol solvent at 80°C (Wagner and Bladt 1994). Hyperforin is a lipophilic constituent of hypericum that is present in the oil extract (Chatterjee et al. 1998a). It is not very stable, but its presence is sustained by hot maceration of the flowers and storage in the absence of air (Maisenbacher and Kovar 1992). 

The pharmacokinetics of hypericin and pseudohypericin piasma have been studied as weii (Brockmoiier et ai. 1997). Human subjects receiving piacebo, or 900, 1800, or 3600 mg of a standardized hypericum extract (LI 160), which contained 0, 2.81, 5.62, and 11.25 mg of totai hypericin and pseudohypericin, achieved maximum total plasma concentrations at 4 hours (0.028, 0.061, and 0.159 mg/L, respectively). The half-lives of absorption, distribution, and elimination were 0.6, 6.0, and 43.1 hours, respectively, using 750 pg of hypericin, and are slightly different for 1578 pg of pseudohypericin (1.3, 1.4, and 24.8 hours, respectively) (Kerb et ai. 1996). The systemic availability of the hypericum extract LI 160 is between 14 and 21%. Comparable results are found in another study using LI 160 (Staffeldt et ai. 1994). Long-term dosing of 3 x 300 mg per day showed that steady-state levels of hypericin are reached after 4 days. 

Although the individual inhibition of either MAO or COMT may be comparatively minor in isolation, their combined inhibition along with other monoamine or nonmonoamine actions could have additive if not synergistic effects. For example, a fraction with combined hypericin and flavonoids had antidepressant effects in an animal model (Butterweck et al. 1997). Hypericum is a particular case wherein a single isolated principle may be sufficient for the desired effect, but less effective than the entire plant extract. 

Amino acid neurotransmitter Constituents of hypericum also appear to have effects on amino acid neurotransmission, particularly GABA. Hypericin and a crude extract bind to GABAA and GABAB receptors (Cott 1997). Hyperforin also inhibits synaptosomal GABA reuptake in the low micromolar range (IC50 values of 0.05-0.10 ug/ml). Activity at GABAA benzodiazepine receptors was noted in extracts of four hy-... 

Other mechanisms A few other effects of hypericin, and a crude hypericum extract have been found, including affinity for NMDA, inositol triphosphate, and adenosine receptors. However, these are not likely to be significant to its therapeutic effects because concentrations required for these interactions are not likely to be achieved by oral administration (Cott 1997). Vasoactive effects are possible because hypericum extracts blocked the vasoconstricting effects of histamine and prostaglandin F2o in porcine coronary arteries, and some vasorelaxation occurs in one particular fraction. These effects are hypothesized to be mediated by inhibition of phosphodiesterase (Melzer et al. 1991). 

Finally, a thorough receptor binding study by Raffa and colleagues (1998) showed that hypericin extracts had no effect at adrenergic (alpha or beta), adenosine, angiotensin, benzodiazepine, dopamine, bradykinin, neuropeptide Y, PCP, NMDA, opioid, cholecystokinin A, histamine HI, or nicotinic ACh receptors. Although comprehensive, this study did not look at the binding of any other hypericum constituents. 

Staffeldt B, Kerb R, Brockmoller J, Ploch M, Roots I. (1994). Pharmacokinetics of hypericin and pseudohypericin after oral intake of the hypericum perforatum extract LI 160 in healthy volunteers. J Geriatr Psychiatry Neurol. 7(suppl 1) S47-53. 

Hypericin from St. John s wort is another phenolic compound with multiple rings and multiple double bonds. This compound readily absorbs UV light and is a primary photosensitizing agent that will result in severe sunburn in species that either ingest the plant or come in contact with plant dust or leaf extracts. 

St. John s wort is commonly used at 300 mg of extract (standardized to 0.3% hypericin) three times daily for 6 weeks or longer. Short-term treatment is usually ineffective. 

Many constituents with potential biological activity have been extracted from the flowers and leaves, the parts of the plant used for medicinal purposes. These include naphthodianthrones, flavonoids, phlorogluci-nols, and xanthones. Hypericin, one of the naphthodianthrones, has traditionally been considered the main active ingredient, but it is not known whether this is the compound with antidepressant activity. Recent data suggest that a component called hyperforin may be more important than hypericin for the antidepressant activity. 

A multicenter trial comparing more appropriate doses of imipramine (75 mg twice daily, N = 167) and St. John s wort extract (250 mg twice daily standardized to 0.2% hypericin, N = 157) showed no difference in efficacy after 6 weeks of treatment. However, St. John s wort seemed to reduce anxiety symptoms more often than imipramine and was better tolerated (Woelk, 2000). A study including 240 participants compared St. John s wort with fluoxetine in mild to moderate depression and also concluded that efficacy of both treatments was comparable (Schrader, 2000). These results have been replicated in a smaller trial us-... 

It is not possible to discuss pharmacokinetics when the active compound or compounds of St. John s wort are not known. The half-life of hypericin and hyperforin have been estimated at between 6 and 9 hours, with peak plasma concentrations at about 2-3 hours after administration. Some of the ingredients of Hypericum extracts are metabolized in the liver. 

The adult dosage of St John s wort traditionally recommended for treating depression is 300 mg of plant extract orally three times daily (plant extracts are usually standardised to 0.3% hypericin). There are no data about optimal dosage in young people. Clinicians often start with half the adult dose and increase the amount up to 300 mg three times daily after 3 or 4 weeks if the herb is well tolerated and there is no improvement. Clinical experience shows this regime results in few unwanted effects in the young. 

It is important to note that amounts used vary considerably and there are no systematic studies on the minimum therapeutic dosage. Furthermore, the quantity of active substances can change depending on factors such as the extraction process, season, and plant part used. Quality control studies show that the amount of active ingredients (e.g., hypericin) is often very different from those advertised in the label. 

This herbal product has the most data available to support its usefulness as an antidepressant. Nevertheless, only minimal information is available about its pharmacology and its relative risk-benefit ratio. At least seven different biologically active chemicals have been isolated from crude extracts of hypericum. Several are ubiquitous in the plant kingdom. The exceptions are hypericin and pseudohypericin, which have been assumed to be responsible for any antidepressant activity of this product. Nevertheless, there is the potential for one or more of these seven compounds and their metabolites to mediate desired or undesired effects, particularly when used in combination with other medications (i.e., herb-drug interactions). 

As with drugs and purified biomarkers, thermal- and photostability of botanical products are the factors that must be considered. Commercial dried extract and capsules of SJW were evaluated under harmonized test conditions (25). Photostability testing showed all the constituents to be photosensitive in the tested conditions. However, different opacity agents and pigments influenced the stability of the constituents. Amber containers had little effect on the photostability of the investigated constituents. Long-term thermal stability testing showed a shelf life of less than four months for hyperforins and hypericins, even when ascorbic and citric acids were added to the formulation. 

Thiede HM, Walper A. Inhibition of MAO and COMT by hypericum extracts and hypericin. J Geriatr Psychiatr Neurol 1994 7(suppl 1) S54-S56. 

The pharmacological activity of SJW extracts has recently been reviewed (55-58). Recent reports have shown that the antidepressant activity of Hypericum extracts can be attributed to the phloroglucinol derivative hyperforin (59-62), to the naphthodianthrones hypericin and pseudohypericin (18,63-65), and to several flavonoids (66-69). The role and the mechanisms of action of these different compounds are still a matter of debate. But, taking these previous findings together, it is likely that several constituents are responsible for the clinically observed antidepressant efficacy of SJW. 

Single- and multiple-dose pharmacokinetic studies with extracts of SJW were performed in rats and humans, which focused on the determination of plasma levels of the naphthodianthrones hypericin and pseudohypericin and the phloroglucinol derivative hyperforin. Results from pharmacokinetic... 

A placebo-controlled, randomized clinical trial with monitoring of hypericin and pseudohypericin plasma concentrations was performed to evaluate the increase in dermal photosensitivity in humans after application of high doses of SJW extract (Table 2) (73). The study was divided into a single-dose and a multiple-dose part. In the single dose crossover study, each of the 13 volunteers received either placebo or 900, 1800, or 3600 mg of the SJW extract LI 160. Maximum total hypericin plasma concentrations were observed about four hours after dosage and were 0, 28, 61, and 159ng/mL, respectively. Pharmacokinetic parameters had a dose relationship that appeared to follow linear kinetics (73). 

LI 160 tablets containing 300mg extract (0.25mg hypericin and 0.52mg pseudohypericin per tablet). 

Pharmacokinetics, safety, and antiviral effects of hypericin were studied in patients with chronic hepatitis C infection (Fig. 4) (71). The patients received an eight-weeks course of 0.05 and 0.10 mg/kg hypericin orally once a day. The pharmacokinetic data revealed a long elimination half-life (mean values of 36.1 and 33.8 hours, respectively, for the doses of 0.05 and 0.10 mg/ kg) and mean AUC determinations of 1.5 and 3.1 pg/mL/hr, respectively. Because relatively high doses of 0.05 and O.lOmg/kg/day were given, which will probably be not reached after oral intake of recommended doses of SJW extract preparations, it is not surprising that hypericin caused a considerable phototoxicity in this study. 

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