TRPC6[1]

The activity of hyperforin is multidirectional. It inhibits voltage-gated channels (Ca2+, K+, and Na+) as well as ligand-gated channels (GABA, NMDA, and AMPA receptors) in conductance[2]. In vitro-cultured murine splenic γδ T cells demonstrate a reduction in IL-17A expression and secretion upon treatment with hyperforin (0.1, 1, 10 μM; 2 h)[3]. In TNF-stimulated HaCaT cells, hyperforin (0.1, 1, 10 μM; 2 h) inhibits the phosphorylation of the MAPK and STAT3 pathways[3]. Without causing any harmful consequences, hyperforin (IC50=3.7 μmol/L) suppresses the development of microvascular tubes and the proliferation of HDMEC in a dose-dependent manner[4].

Imiquimod psoriatic skin lesions in mice are improved by hyperforin (5 mg/kg; ip; once daily for 7 d); it also prevents inflammatory cell infiltration and the release of inflammatory cytokines[3].

Western Blot Analysis[3]
Cell Types: HaCaT cells
Tested Concentrations: 0.1, 1, 10 μM; with or without 10, 20 ng/mL TNF-α
Incubation Duration: 2 hrs (hours)
Experimental Results: diminished the expressions of p-p38, p-ERK, p-JNK, and p-STAT3 , especially at the dosage of 10 μM.

Animal/Disease Models: IMQ-induced psoriasis-like mice model[3]
Doses: 5 mg/kg
Route of Administration: intraperitoneal (ip) injection; one time/day for 7 days
Experimental Results: Dramatically ameliorated skin lesion throughout the treatment period, demonstrated by the decreased severity score of skin inflammation. Suppressed infiltration of CD3+ T cells and downregulated expression of Il1 , Il6, Il23, Il17a, Il22, antimicrobial peptides (AMPs) in the skin lesion.

Absorption, Distribution and Excretion
This article describes an effective analytical method for determining plasma hypericin levels after administration of a hypericin-containing extract of St. John's wort. Following oral administration of 300 mg/kg of St. John's wort extract (WS 5572, containing 5% hypericin) to rats, plasma hypericin concentrations were determined using high-performance liquid chromatography-ultraviolet detection (HPLC-UV). Peak plasma hypericin concentrations were reached after 3 hours, approximately 370 ng/ml (approximately 690 nM). The estimated half-life and clearance were 6 hours and 70 ml/min/kg, respectively. Because the therapeutic dose of St. John's wort extract is significantly lower than the experimental dose in rats, a more sensitive LC-MS/MS method was developed. The limit of quantitation for this method is 1 ng/ml. Using this method, plasma hypericin levels can be monitored for up to 24 hours after administration of a film-coated tablet containing 300 mg of St. John's wort extract (equivalent to 14.8 mg of hypericin) to healthy volunteers. Peak plasma hypericin concentrations were reached 3.5 hours after administration, at approximately 150 ng/ml (approximately 280 nM). The half-life and mean residence time were 9 hours and 12 hours, respectively. The pharmacokinetics of hypericin were linear up to 600 mg of the extract. When the extract dose was increased to 900 mg or 1200 mg, the Cmax and AUC values were lower than expected based on linear extrapolation from low-dose data. Volunteer plasma concentration curves fit well to an open two-compartment model. No accumulation of hypericin in plasma was observed in repeated-dose studies. Based on the AUC values observed in repeated-dose studies, the steady-state plasma concentration of hypericin is estimated to be approximately 100 ng/ml (approximately 180 nM) after administration of the extract at 300 mg three times daily (i.e., the normal therapeutic dose regimen).
After oral administration of 300 mg/kg of Hypericum extract (WS 5572, containing 5% hypericin) to rats, the plasma concentration reached a peak of approximately 370 ng/ml (approximately 690 nM) after 3 hours, as determined by HPLC-UV detection. The estimated half-life and clearance were 6 hours and 70 ml/min/kg, respectively. Since the therapeutic dose of Hypericum extract is much lower than that used in rats, a more sensitive LC/MS/MS method was developed. The limit of quantification for this method is 1 ng/ml. Using this method, plasma hypericin levels can be monitored for up to 24 hours after administration of a film-coated tablet containing 300 mg of Hypericum extract (equivalent to 14.8 mg of hypericin) to healthy volunteers. The peak plasma hypericin concentration was reached 3.5 hours after administration, at approximately 150 ng/ml (approximately 280 nM). The half-life and mean residence time were 9 hours and 12 hours, respectively. The pharmacokinetics of hypericin were linear until the extract dose reached 600 mg. When the extract dose was increased to 900 mg or 1200 mg, the Cmax and AUC values were lower than the expected values extrapolated linearly from the low-dose data. Volunteer plasma concentration curves fit well to the open two-compartment model. No accumulation of hypericin in plasma was observed in the repeated-dose study. Based on the AUC values observed in the repeated-dose study, the steady-state plasma concentration of hypericin after three daily doses of 300 mg of the extract (i.e., the normal therapeutic dose regimen) is estimated to be approximately 100 ng/ml (approximately 180 nM).

---

Metabolism/Metabolites
Hypericin is an important active component of Hypericum perforatum and is believed to be associated with the antidepressant effects and herb-drug interactions of Hypericum perforatum. In this study, the in vitro metabolic profile of hypericin was investigated using liver microsomes from male and female Sprague-Dawley rats, with or without phenobarbital or dexamethasone induction. Four major phase I metabolites were isolated by high-performance liquid chromatography and named 19-hydroxyhypericin, 24-hydroxyhypericin, 29-hydroxyhypericin, and 34-hydroxyhypericin, respectively, and identified by mass spectrometry and nuclear magnetic resonance. The results indicate that hydroxylation is a major biotransformation step in the hypericin metabolic pathway in rat liver, and inducible cytochrome P450 3A (CYP450 3A) and/or CYP2B may be the main cytochrome P450 isoenzymes catalyzing these hydroxylation reactions.
Repeated testing of the aerial parts of St. John's wort revealed a new hypericin degradation product (1), namely deoxyfuran hypericin A (2), as well as previously identified hypericin (3), hypericin (4), hypericin A (5a and 5b), pyrano[7,28-b] hypericin (6) and 3-methyl-4,6-di(3-methyl-2-butenyl)-2-(2-methyl-1-oxopropyl)-3-(4-methyl-3-pentenyl)-cyclohexanone (7)

---

Biological half-life
9 hours
After oral administration of 300 mg/kg of Hypericin extract (WS 5572, containing 5% hypericin) to rats... the estimated half-life and clearance rate were 6 hours and 70 mL/min/kg, respectively... the half-life and mean residence time were 9 hours and 12 hours, respectively.

Interactions
Concomitant use with St. John's wort can lead to decreased plasma concentrations of several drugs, including amitriptyline, cyclosporine, digoxin, indinavir, irinotecan, warfarin, phenylcoumarin, alprazolam, dexmedetomidine, simvastatin, and oral contraceptives.

[1]. TRPC6 channel-mediated neurite outgrowth in PC12 cells and hippocampal neurons involves activation of RAS/MEK/ERK, PI3K, and CAMKIV signaling. J Neurochem. 2013 Nov;127(3):303-13.

[2]. Hyperforin Potentiates Antidepressant-Like Activity of Lanicemine in Mice. Front Mol Neurosci. 2018 Dec 12;11:456.

[3]. Hyperforin Ameliorates Imiquimod-Induced Psoriasis-Like Murine Skin Inflammation by Modulating IL-17A-Producing γδ T Cells. Front Immunol. 2021 May 5;12:635076.

[4]. Biosynthesis of hyperforin in Hypericum perforatum. J Med Chem. 2002 Oct 10;45(21):4786-93.

[5]. Hyperforin: A natural lead compound with multiple pharmacological activities. Phytochemistry. 2023 Feb;206:113526.

Therapeutic Uses
Hypericin (Hyp) is a polyphenol derivative of Hypericum perforatum. It is not only a key component of the plant's antidepressant activity but also inhibits the proliferation of bacteria, lymphocytes, and tumor cells, as well as the activity of matrix proteases. We tested whether Hyp, in addition to inhibiting leukocyte elastase (LE) activity, could effectively control the recruitment of polymorphonuclear neutrophils (PMNs) and the resulting adverse tissue responses. The results showed that Hyp (in its stable dicyclohexylammonium form) inhibited the chemotaxis and chemical invasiveness of human PMN cells in a dose-dependent manner without affecting their viability and chemokine receptor expression in vitro (IC50 for both was 1 μM). This effect is associated with decreased expression of the neutrophil adhesion molecule CD11b stimulated by formyl-Met-Leu-Phe and inhibited activation of the gelatinase matrix metalloproteinase-9 (MMP-9) triggered by LE. In an interleukin-8 induced mouse model, both local injection and daily intraperitoneal injection of Hyp salt blocked PMN-triggered angiogenesis. Furthermore, intraperitoneal injection of Hyp reduced acute PMN recruitment and promoted inflammation resolution in a bleomycin-induced lung inflammation model, significantly reducing subsequent fibrosis. These results demonstrate that Hyp is a potent anti-inflammatory compound with therapeutic potential and elucidate its mechanism of action.
/EXPL THER/ ... Hypericin (HF) is a natural phloroglucinol that stimulates apoptosis in B-cell chronic lymphocytic leukemia (CLL) cells and exhibits anti-angiogenic properties. This study investigated the effect of hypericin on P-gp/MDR1 activity, an ABC (ATP-binding cassette) transporter that may be involved in multidrug resistance (MDR). In vitro experiments showed that hypericin treatment of chronic lymphocytic leukemia (CLL) cells significantly inhibited P-gp activity, manifested as a decreased ability to efflux the rhodamine 123 probe after treatment. Furthermore, most CLL cells express breast cancer resistance protein (BCRP), another ABC transporter. Hypericin also inhibited BCRP activity, manifested as a decreased ability of CLL cells to efflux the specific probe mitoxantrone after treatment. In myeloid leukemia cell lines, particularly HL-60/DNR cells resistant to daunorubicin and overexpressing P-gp, hypericin was confirmed to reverse the activity of both P-gp and BCRP. Therefore, the results indicate that hypericin (HF), in addition to its pro-apoptotic properties, may also have the potential to reverse multidrug resistance (MDR), thus potentially offering therapeutic value for chronic lymphocytic leukemia (CLL) and other hematologic malignancies. Hypericin (Hyp), an active compound in the extract of St. John's wort, is known for its antidepressant activity. However, studies have found that Hyp also possesses various other biological properties, including inhibition of tumor invasion, angiogenesis, and inflammation. In this study, we found that Hyp treatment inhibited IFN-γ production and downregulated T-box (T-bet; a Th1 gene expression marker) and upregulated GATA-3 (a Th2 gene marker) on IL-2/PHA-activated T cells. Simultaneously, we also found a significant downregulation of the chemokine receptor CXCR3 expression on activated T cells. This latter effect, along with the downregulation of matrix metalloproteinase 9 expression, may ultimately lead to an inhibition of the ability of activated lymphocytes to migrate toward chemokine CXCL10 and cross the matrix, consistent with our in vitro observations. Therefore, we evaluated the role of Hyp in an experimental allergic encephalomyelitis (EAE) animal model. EAE is a classic Th1-mediated autoimmune disease of the central nervous system. We observed that Hyp significantly reduced the severity of disease symptoms. In summary, these properties make Hyp a potential therapeutic molecule for treating Th1 cell-mediated autoimmune inflammatory diseases, including EAE. Premature ejaculation is the most common sexual dysfunction in men, but there are currently no approved effective treatments. …The in vivo efficacy of hypericin (HF), a concentrated extract of Hypericum, was investigated in an anesthetized rat ejaculation model. In this study, a male rat was anesthetized (subcutaneously injected with 1.2 g/kg ethyl carbamate), and physiological saline was injected into the urethra at a rate of 116 μL/s for 2 seconds to briefly increase urethral pressure, inducing rhythmic contractions of the bulbospongiosus muscle (BS). Electrodes within the bladder muscle recorded electrical activity during contraction, which appeared as clusters of pulses on electromyography. Subcutaneous injection of the 5-hydroxytryptamine 1A receptor agonist 8-hydroxy-2-(di-n-propylamino)tetrahydronaphthalene (8-OH-DPAT) (0.4 mg/kg) enhanced bladder muscle contraction induced by increased urethral pressure. In the carrier control rats, administration of 8-OH-DPAT significantly accelerated ejaculation, with increases in electrical discharge amplitude and pulse duration associated with increased urethral pressure of 203.2% ± 32.9% and 178.1% ± 22.9% from baseline, respectively. Within a dose range of 5 to 80 mg/kg, the HF extract reduced the effect of 8-OH-DPAT on ejaculation. The effect of the HF extract in reducing the electrical pulse amplitude persisted after mid-thoracic spinal cord transection, suggesting that the site of action of HF may be the spinal ejaculatory mechanism or it may act directly on neurons innervating the bladder muscles. This is the first report of the role of HF in a rat ejaculation model. HF can be considered a novel treatment for premature ejaculation. For more complete data on the therapeutic uses of hypericin (6 types), please visit the HSDB record page. Drug Warnings: Concomitant use with St. John's wort can reduce plasma concentrations of several drugs, including amitriptyline, cyclosporine, digoxin, indinavir, irinotecan, warfarin, phenylcoumarin, alprazolam, dexmedetomidine, simvastatin, and oral contraceptives. Pharmacodynamics: Hypericin is considered the main active ingredient in St. John's wort extract that exerts its antidepressant and anti-anxiety effects. It is a reuptake inhibitor of monoamine neurotransmitters (including serotonin, norepinephrine, and dopamine) as well as gamma-aminobutyric acid (GABA) and glutamate. Except for glutamate (IC50 values in the range of 0.5 mcg/ml), the IC50 values of all compounds are 0.05–0.10 mcg/ml. It appears to exert these effects by activating the transient receptor potential ion channel TRPC6. Activation of TRPC6 induces sodium and calcium ions to enter the cell, thereby inhibiting the reuptake of monoamine neurotransmitters. Hypericin is also thought to induce the expression of cytochrome P450 enzymes CYP3A4 and CYP2C9 by binding to and activating the pregnane X receptor (PXR).

C35H52O4

536.78

536.386

11079-53-1

Hyperforin dicyclohexylammonium salt;238074-03-8

441298

Typically exists as solid at room temperature

1.0±0.1 g/cm3

616.8±55.0 °C at 760 mmHg

79-80ºC

340.9±28.0 °C

0.0±4.0 mmHg at 25°C

1.518

12.3

1

4

11

39

1140

4

C/C(=C/CC[C@@]1([C@@H](C/C=C(\C)/C)C[C@]2(C(=C(C(=O)C1(C2=O)C(C(C)C)=O)C/C=C(\C)/C)O)C/C=C(/C)\C)C)/C

KGSZHKRKHXOAMG-HQKKAZOISA-N

InChI=1S/C35H52O4/c1-22(2)13-12-19-33(11)27(16-14-23(3)4)21-34(20-18-25(7)8)30(37)28(17-15-24(5)6)31(38)35(33,32(34)39)29(36)26(9)10/h13-15,18,26-27,37H,12,16-17,19-21H2,1-11H3/t27-,33+,34+,35-/m0/s1

(1R,5R,7S,8R)-4-hydroxy-8-methyl-3,5,7-tris(3-methylbut-2-enyl)-8-(4-methylpent-3-enyl)-1-(2-methylpropanoyl)bicyclo[3.3.1]non-3-ene-2,9-dione

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

May dissolve in DMSO (in most cases), if not, try other solvents such as H2O, Ethanol, or DMF with a minute amount of products to avoid loss of samples

Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples.

---

Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)
*Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution.
Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)
Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil)
Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals).

View More

Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)]
*Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.
Injection Formulation 5: 2-Hydroxypropyl-β-cyclodextrin : Saline = 50 : 50 (i.e. 500 μL 2-Hydroxypropyl-β-cyclodextrin → 500 μL Saline)
Injection Formulation 6: DMSO : PEG300 : castor oil : Saline = 5 : 10 : 20 : 65 (i.e. 50 μL DMSO → 100 μLPEG300 → 200 μL castor oil → 650 μL Saline)
Injection Formulation 7: Ethanol : Cremophor : Saline = 10： 10 : 80 (i.e. 100 μL Ethanol → 100 μL Cremophor → 800 μL Saline)
Injection Formulation 8: Dissolve in Cremophor/Ethanol (50 : 50), then diluted by Saline
Injection Formulation 9: EtOH : Corn oil = 10 : 90 (i.e. 100 μL EtOH → 900 μL Corn oil)
Injection Formulation 10: EtOH : PEG300：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL EtOH → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)

---

Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium)
Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose
Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals).

View More

Oral Formulation 3: Dissolved in PEG400
Oral Formulation 4: Suspend in 0.2% Carboxymethyl cellulose
Oral Formulation 5: Dissolve in 0.25% Tween 80 and 0.5% Carboxymethyl cellulose
Oral Formulation 6: Mixing with food powders

---

Note: Please be aware that the above formulations are for reference only. InvivoChem strongly recommends customers to read literature methods/protocols carefully before determining which formulation you should use for in vivo studies, as different compounds have different solubility properties and have to be formulated differently.

---

 (Please use freshly prepared in vivo formulations for optimal results.)