Protective effects of fermented honeybush (Cyclopia intermedia) extract (HU-018) against skin aging: a randomized, double-blinded, placebo-controlled study.

BACKGROUND: Oxidative stress and photodamage resulting from ultraviolet radiation exposure play key roles in skin aging. Fermented Cyclopia intermedia, which is used to brew honeybush tea, exerts antioxidant and anti-wrinkle effects by inhibiting reactive oxygen species production and downregulating matrix metalloproteinase activity. OBJECTIVES: This randomized, double-blinded, placebo-controlled study aimed to evaluate the efficacy and safety of fermented honeybush (Cyclopia intermedia) extract (HU-018) for skin rejuvenation. METHODS: 120 Korean subjects with crow's feet wrinkles were randomized to receive either low-dose extract (400 mg/day), high-dose extract (800 mg/day), or placebo (negative control, only dextran) for 12 weeks. Wrinkles were evaluated using JANUS® and PRIMO pico®. Skin elasticity, hydration and transepidermal water loss were measured. RESULTS: Global skin wrinkle grade was significantly improved in both low-dose and high-dose groups compared to placebo group, as well as for skin hydration and elasticity. Both the low- and high-dose groups showed significantly decreased TEWL compared to the placebo group. There were no adverse effects during the entire study period. CONCLUSION: Our data indicate that HU-018 is effective for improving skin wrinkles, elasticity, and hydration. Therefore, daily supplementation with fermented honeybush could be helpful for protecting against skin aging.

Comparison of the efficacy and safety of 2% lidocaine HCl with different epinephrine concentration for local anesthesia in participants undergoing surgical extraction of impacted mandibular third molars: A multicenter, randomized, double-blind, crossover, phase IV trial.

BACKGROUND: The most commonly impacted tooth is the third molar. An impacted third molar can ultimately cause acute pain, infection, tumors, cysts, caries, periodontal disease, and loss of adjacent teeth. Local anesthesia is employed for removing the third molar. This study aimed to evaluate the efficacy and safety of 2% lidocaine with 1:80,000 or 1:200,000 epinephrine for surgical extraction of bilateral impacted mandibular third molars. METHODS: Sixty-five healthy participants underwent surgical extraction of bilateral impacted mandibular third molars in 2 separate visits while under local anesthesia with 2% lidocaine with different epinephrine concentration (1:80,000 or 1:200,000) in a double-blind, randomized, crossover trial. Visual analog scale pain scores obtained immediately after surgical extraction were primarily evaluated for the 2 groups receiving different epinephrine concentrations. Visual analog scale pain scores were obtained 2, 4, and 6 hours after administering an anesthetic. Onset and duration of analgesia, onset of pain, intraoperative bleeding, operator's and participant's overall satisfaction, drug dosage, and hemodynamic parameters were evaluated for the 2 groups. RESULTS: There were no statistically significant differences between the 2 groups in any measurements except hemodynamic factors (P >.05). Changes in systolic blood pressure and heart rate following anesthetic administration were significantly greater in the group receiving 1:80,000 epinephrine than in that receiving 1:200,000 epinephrine (P ≤.01). CONCLUSION: The difference in epinephrine concentration between 1:80,000 and 1:200,000 in 2% lidocaine liquid does not affect the medical efficacy of the anesthetic. Furthermore, 2% lidocaine with 1:200,000 epinephrine has better safety with regard to hemodynamic parameters than 2% lidocaine with 1:80,000 epinephrine. Therefore, we suggest using 2% lidocaine with 1:200,000 epinephrine rather than 2% lidocaine with 1:80,000 epinephrine for surgical extraction of impacted mandibular third molars in hemodynamically unstable patients.

HS-23, a Lonicera japonica extract, reverses sepsis-induced immunosuppression by inhibiting lymphocyte apoptosis.

ETHNOPHARMACOLOGICAL RELEVANCE: Lonicera japonica Thunberg, a widely used traditional Chinese medicine, possesses antibacterial, antiviral, and antiendotoxin activities. This study investigated the molecular mechanisms of HS-23, the ethanol extract of the dried flower buds of L. japonica, on sepsis-induced immunosuppression. MATERIALS AND METHODS: Male ICR mice were intravenously administered HS-23 (10, 20, and 40mg/kg) immediately (0h) and 22h after cecal ligation and puncture (CLP). The spleen was isolated for biochemical assays 24h after CLP. RESULTS: HS-23 improved sepsis-induced mortality. CLP induced a marked decrease in the number of splenocytes, B cells, and natural killer cells, which was attenuated by HS-23. HS-23 also attenuated CLP-induced apoptosis in CD4(+) and CD8(+) T cells and inhibited both the intrinsic and extrinsic apoptotic pathway in the spleen. HS-23 attenuated the CLP-induced decrease in interleukin (IL)-17 production. CLP significantly decreased splenic production of tumor necrosis factor-α and IL-2, and these effects were attenuated by HS-23. CONCLUSION: Our findings suggest that HS-23 reverses immunosuppression during the late phase of sepsis by inhibiting lymphocyte apoptosis and enhancing Th1 cytokine production. HS-23 warrants further evaluation as a potential therapeutic agent for the treatment of sepsis.

Metabolism-mediated drug interaction potential of HS-23, a new herbal drug for the treatment of sepsis in human hepatocytes and liver microsomes.

HS-23, an extract of the dried flower buds of Lonicera japonica, is a new botanical drug currently being evaluated in a phase I clinical study in Korea for the treatment of sepsis. The in vitro induction and inhibition potentials of HS-23 on the drug-metabolizing enzymes using human hepatocytes and liver microsomes were assessed to evaluate herb-drug interaction according to botanical drug guideline and drug interaction guidance of FDA. HS-23 slightly inhibited CYP2A6, CYP2B6, CYP2C9, CYP2C19, and CYP3A4 enzyme activities in human liver microsomes with IC50 values of 80.6, 160.7, 169.5, 85.4, and 76.6 µg/mL, respectively. HS-23 showed negligible inhibition of CYP1A2, CYP2C8, CYP2D6, UGT1A1, UGT1A4, UGT1A9, and UGT2B7 activities in human liver microsomes. Based on these results, HS-23 may not inhibit the metabolism of CYP2A6, CYP2B6, CYP2C9, CYP2C19, and CYP3A4-catalyzed drugs in humans. HS-23 did not affect the mRNA expression of CYP1A2, CYP2B6, and CYP3A4 after 48 h treatment at three concentrations (0.5, 5, and 50 µg/mL) in three independent human hepatocytes, indicating that HS-23 has no effect on herb-drug interactions that up- or down-regulate CYP1A2, CYP2B6, and CYP3A4. These results indicate that the administration of HS-23 in human may not cause clinically relevant inhibition and induction of these cytochrome P450 (CYP) and UDP-glucuronosyltransferase (UGT) enzymes and HS-23 may be promising therapeutic agent for treatment of sepsis.

Metabolism of rutin and poncirin by human intestinal microbiota and cloning of their metabolizing α-L-rhamnosidase from Bifidobacterium dentium.

To understand the metabolism of flavonoid rhamnoglycosides by human intestinal microbiota, we measured the metabolic activity of rutin and poncirin (distributed in many functional foods and herbal medicine) by 100 human stool specimens. The average α-Lrhamnosidase activities on the p-nitrophenyl-α-L-rhamnopyranoside, rutin, and poncirin subtrates were 0.10 ± 0.07, 0.25 ± 0.08, and 0.15 ± 0.09 pmol/min/mg, respectively. To investigate the enzymatic properties, α-L-rhamnosidase-producing bacteria were isolated from the specimens, and the α-L-rhamnosidase gene was cloned from a selected organism, Bifidobacterium dentium, and expressed in E. coli. The cloned α-L-rhamnosidase gene contained a 2,673 bp sequcence encoding 890 amino acid residues. The cloned gene was expressed using the pET 26b(+) vector in E. coli BL21, and the expressed enzyme was purified using Ni(2+)-NTA and Q-HP column chromatography. The specific activity of the purified α-L-rhamnosidase was 23.3 µmol/min/mg. Of the tested natural product constituents, the cloned α-L-rhamnosidase hydrolyzed rutin most potently, followed by poncirin, naringin, and ginsenoside Re. However, it was unable to hydrolyze quercitrin. This is the first report describing the cloning, expression, and characterization of α-L-rhamnosidase, a flavonoid rhamnoglycosidemetabolizing enzyme, from bifidobacteria. Based on these findings, the α-L-rhamnosidase of intestinal bacteria such as B. dentium seem to be more effective in hydrolyzing (1-->6) bonds than (1-->2) bonds of rhamnoglycosides, and may play an important role in the metabolism and pharmacological effect of rhamnoglycosides.

Magnolia officinalis attenuates free fatty acid-induced lipogenesis via AMPK phosphorylation in hepatocytes.

ETHNOPHARMACOLOGICAL RELEVANCE: Magnolia officinalis (MO) is a traditional Chinese herbal medicine that has been used in clinical practice to treat liver disease. The aim of this study is to examine the effects of MO on the development of nonalcoholic fatty liver in hepatocytes. MATERIALS AND METHODS: Human hepatoma-derived HepG2 cells and mouse normal FL83B hepatocytes were exposed to 0.5mM free fatty acids (FFAs; oleate:palmitate, 2:1) for 24h to simulate conditions of nonalcoholic fatty liver in vitro. The cells were treated with a standardized MO extract 1h prior to FFA exposure. RESULTS: MO pretreatment attenuated the increases in intracellular lipid accumulation and triglyceride content in FFA-exposed hepatocytes in a dose-dependent manner. MO pretreatment significantly inhibited both sterol regulatory element-binding protein (SREBP)-1c activation and increases in fatty acid translocase, fatty acid synthase, and stearoyl CoA desaturase-1 protein expression in FFA-exposed hepatocytes in a dose-dependent manner. MO pretreatment markedly induced adenosine monophosphate-activated protein kinase (AMPK) phosphorylation in hepatocytes. Compound C, an AMPK inhibitor, blocked the inhibitory effect of MO on the increases in intracellular lipid accumulation and triglyceride content induced by FFAs. In hepatocytes pretreated with compound C, MO failed to inhibit SREBP-1c activation and the increases in fatty acid translocase, fatty acid synthase, and stearoyl-CoA desaturase-1 protein expression induced by FFAs. CONCLUSIONS: Our results indicate that MO attenuates triglyceride biosynthesis and accumulation induced by FFAs in hepatocytes, suggesting its pharmacological potential for the prevention of nonalcoholic fatty liver disease. These effects may be mediated by the inhibition of SREBP-1c via AMPK phosphorylation.

HS-23, Lonicera japonica extract, attenuates septic injury by suppressing toll-like receptor 4 signaling.

ETHNOPHARMACOLOGICAL RELEVANCE: Lonicera japonica Thunberg is a traditional herbal medicine widely used in East Asia as an anti-bacterial, anti-inflammatory, and antiviral agent. This study was designed to investigate the effects of HS-23, ethanol extract of the dried flower buds of Lonicera japonica, in experimental models of sepsis and elucidate the mechanisms of action of HS-23. MATERIALS AND METHODS: Male ICR mice were intravenously administered HS-23 (20 and 40 mg/kg) for 0 (immediately) and 24 h after cecal ligation and puncture (CLP) for survival tests, and HS-23 (40 mg/kg) immediately after CLP for biochemical assays. RESULTS: HS-23 improved sepsis-induced mortality, enhanced bacterial clearance, and attenuated multiple organ failure. The mechanisms of action of HS-23 included attenuation of increased toll-like receptor (TLR)4 protein and mRNA expression. HS-23 suppressed sepsis-induced increases in protein expression of myeloid differentiation primary response protein 88, p38 and c-Jun N-terminal kinase in both liver and lung, as well as TIR-domain-containing adapter-inducing interferon-ß and interferon regulatory transcription factor 3 protein expression in liver. CONCLUSION: The results of this study revealed that HS-23 attenuated sepsis through suppression of TLR signaling pathways. Therefore, our findings suggest that HS-23 might be useful as a potential therapeutic agent for treatment of sepsis.

Soyasaponins Ab and Bb prevent scopolamine-induced memory impairment in mice without the inhibition of acetylcholinesterase.

Soy (Glycine max, family Leguminosae), which contains isoflavones and saponins as main constituents, is known to exhibit memory-enhancing effects. Therefore, to investigate the role of soyasaponins in memory impairments, we isolated soyasaponins Ab (SA) and Bb (SB) from soybean and measured their protective effects against scopolamine-induced memory impairment in mice. SA and SB significantly prevented scopolamine-induced memory impairment in passive avoidance and Y-maze tasks. Compared to SA, SB rescued memory impairment more potently. Treatment with SB (10 mg/kg, p.o.) protected memory impairment in passive avoidance and Y-maze tasks to 97% (F = 68.10, P < 0.05) and 78% (F = 35.57, P < 0.05) of untreated normal control level, respectively. SA and SB (10 mg/kg) also rescued scopolamine-induced memory impairment in Morris water maze task (F = 14.51, P < 0.05). In addition, soyasaponins preserved brain-derived neurotrophic factor (BNDF) expression (F = 33.69, P < 0.05) and cAMP response element-binding (CREB) protein phosphorylation (F = 91.62, P < 0.05) in the hippocampus of scopolamine-treated mice. However, SA and SB did not inhibit acetylcholinesterase in vitro and ex vivo. On the basis of these findings, we suggest that soybean, particularly soyasaponins, may protect memory impairment by increasing BDNF expression and CREB phosphorylation.

Soyasaponin I improved neuroprotection and regeneration in memory deficient model rats.

Soy (Glycine Max Merr, family Leguminosae) has been reported to possess anti-cancer, anti-lipidemic, estrogen-like, and memory-enhancing effects. We investigated the memory-enhancing effects and the underlying mechanisms of soyasaponin I (soya-I), a major constituent of soy. Impaired learning and memory were induced by injecting ibotenic acid into the entorhinal cortex of adult rat brains. The effects of soya-I were evaluated by measuring behavioral tasks and neuronal regeneration of memory-deficient rats. Oral administration of soya-I exhibited significant memory-enhancing effects in the passive avoidance, Y-maze, and Morris water maze tests. Soya-Ι also increased BrdU incorporation into the dentate gyrus and the number of cell types (GAD67, ChAT, and VGluT1) in the hippocampal region of memory-deficient rats, whereas the number of reactive microglia (OX42) decreased. The mechanism underlying memory improvement was assessed by detecting the differentiation and proliferation of neural precursor cells (NPCs) prepared from the embryonic hippocampus (E16) of timed-pregnant Sprague-Dawley rats using immunocytochemical staining and immunoblotting analysis. Addition of soya-Ι in the cultured NPCs significantly elevated the markers for cell proliferation (Ki-67) and neuronal differentiation (NeuN, TUJ1, and MAP2). Finally, soya-I increased neurite lengthening and the number of neurites during the differentiation of NPCs. Soya-Ι may improve hippocampal learning and memory impairment by promoting proliferation and differentiation of NPCs in the hippocampus through facilitation of neuronal regeneration and minimization of neuro-inflammation.

Effect of honokiol on cytochrome P450 and UDP-glucuronosyltransferase enzyme activities in human liver microsomes.

Honokiol is a bioactive component isolated from the medicinal herbs Magnolia officinalis and Magnolia grandiflora that has antioxidative, anti-inflammatory, antithrombotic, and antitumor activities. The inhibitory potentials of honokiol on eight major human cytochrome P450 (CYP) enzymes 1A2, 2A6, 2B6, 2C8, 2C9, 2C19, 2D6, and 3A4, and four UDP-glucuronosyltransferases (UGTs) 1A1, 1A4, 1A9, and 2B7 in human liver microsomes were investigated using liquid chromatography-tandem mass spectrometry. Honokiol strongly inhibited CYP1A2-mediated phenacetin O-deethylation, CYP2C8-mediated amodiaquine N-deethylation, CYP2C9-mediated diclofenac 4-hydroxylation, CYP2C19-mediated [S]-mephenytoin 4-hydroxylation, and UGT1A9-mediated propofol glucuronidation with K(i) values of 1.2, 4.9, 0.54, 0.57, and 0.3 µM, respectively. Honokiol also moderately inhibited CYP2B6-mediated bupropion hydroxylation and CYP2D6-mediated bufuralol 1'-hydroxylation with K(i) values of 17.5 and 12.0 µM, respectively. These in vitro results indicate that honokiol has the potential to cause pharmacokinetic drug interactions with other co-administered drugs metabolized by CYP1A2, CYP2C8, CYP2C9, CYP2C19, and UGT1A9.

The role of intestinal microflora in anti-inflammatory effect of baicalin in mice.

Baicalin, a main constituent of the rhizome of Scutellaria baicalensis, is metabolized to baicalein and oroxylin A in the intestine before its absorption. To understand the role of intestinal microflora in the pharmacological activities of baicalin, we investigated its anti-inflammatory effect in mice treated with and without antibiotics. Orally administered baicalin showed the anti-inflammatory effect in mice than intraperitoneally treated one, apart from intraperitoneally administered its metabolites, baicalein and oroxylin A, which potently inhibited LPS-induced inflammation. Of these metabolites, oroxylin A showed more potent anti-inflammatory effect. However, treatment with the mixture of cefadroxil, oxytetracycline and erythromycin (COE) significantly attenuated the anti-inflammatory effect of orally administered baicalin in mice. Treatment with COE also reduced intestinal bacterial fecal ß-glucuronidase activity. The metabolic activity of human stools is significantly different between individuals, but neither between ages nor between male and female. Baicalin was metabolized to baicalein and oroxylin A, with metabolic activities of 1.427 ± 0.818 and 1.025 ± 0.603 pmol/min/mg wet weight, respectively. Baicalin and its metabolites also inhibited the expression of pro-inflammatory cytokines, TNF-α and IL-1ß, and the activation of NF-κB in LPS-stimulated peritoneal macrophages. Of them, oroxylin A showed the most potent inhibition. Based on these findings, baicalin may be metabolized to baicalein and oroxylin A by intestinal microflora, which enhance its anti-inflammatory effect by inhibiting NF-κB activation.

Gypenoside TN-2 ameliorates scopolamine-induced learning deficit in mice.

ETHNOPHARMACOLOGICAL RELEVANCE: Gynostemma pentaphyllum (Thunb.) Makino (GP, family Cucurbitaceae), which contains dammarane saponins as its main constituents, is used in China, Japan, and Korea as a traditional medicine to treat cancer, obesity, arteriosclerosis, asthma and senility. AIM OF THE STUDY: To investigate the memory-enhancing effects of GP, Gypenoside TN-2 (TN-2) was isolated by activity-guided fractionation and administered to scopolamine-induced memory-deficient mice. MATERIALS AND METHODS: The memory-enhancing effects of TN-2 were evaluated using passive avoidance, Y-maze, and Morris water maze tests, and the protein expressions of brain-derived neurotrophic factor (BDNF), cAMP element binding protein (CREB), and p-CREB were determined by immunoblotting. RESULTS: TN-2 inhibited memory and learning deficits in scopolamine treated mice in the passive avoidance test. TN-2 (10, 20, and 40 mg/kg, p.o.) significantly inhibited memory and learning deficits in the passive avoidance test by 40%, 96% and 78%, respectively, and exhibited significant memory-enhancing effects on the Y-maze test and the Morris water maze test. TN-2 also markedly increased BNDF expression and activated the transcription factor CREB in the hippocampi of scopolamine-treated mice. CONCLUSIONS: TN-2 may ameliorate memory and learning deficits by activating the CREB-BDNF pathway.

Metabolic activities of ginseng and its constituents, ginsenoside rb1 and rg1, by human intestinal microflora.

To evaluate the difference in expressing pharmacological effects of ginseng by intestinal microflora between Koreans, metabolic activities of ginseng, ginsenoside Rb1 and Rg1 by 100 fecal specimens were measured. The ß-glucosidase activity for p-nitrophenyl- ß-D-glucopyranoside was 0 to 0.42 µmol/min/mg and its average activity (mean±SD) was 0.10±0.07 µmol/min/mg. The metabolic activities of ginsenosides Rb1 and Rg1 were 0.01 to 0.42 and 0.01 to 0.38 pmol/min/mg, respectively. Their average activities were 0.25±0.08 and 0.15±0.09 pmol/min/mg, respectively. The compound K-forming activities from ginsenoside Rb1 and ginseng extract were 0 to 0.11 and 0 to 0.02 pmol/min/mg, respectively. Their average compound K-forming activities were 0.24±0.09 pmol/min/ mg and 2.14±3.66 fmol/min/mg, respectively. These activities all were not different between males and females, or between ages. Although compound K-forming activity from the aqueous extract of ginseng was low compared to that from ginenoside Rb1, their profiles were similar to those of isolated compounds. Based on these findings, we believe that the intestinal bacterial metabolic activities of ginseng components are variable in individuals and may be used as selection markers for responders to ginseng.

Purification and characterization of novel salt-active acharan sulfate lyase from Bacteroides stercoris HJ-15.

Salt-active acharan sulfate lyase (no EC number) has been purified from Bacteroides stercoris HJ-15, which was isolated from human intestinal bacteria with GAG degrading enzymes. The enzyme was purified to apparent homogeneity by a combination of QAE-cellulose, diethylaminoethyl (DEAE)-cellulose, CM-Sephadex C-50, HA ultrogel and phosphocellulose column chromatography with the final specific activity of 81.33 micro mol x min-1 x mg-1. The purified salt-active acharan sulfate lyase was activated to 5.3-fold by salts (KCl and NaCl). The molecular weight of salt-active acharan sulfate lyase was 94 kDa by SDS/PAGE and gel filtration. The salt-active acharan sulfate lyase showed optimal activity at pH 7.2 and 40 degrees C. Salt-active acharan sulfate lyase activity was potently inhibited by Cu2+, Ni2+ and Zn2+. This enzyme was inhibited by some agents, butanediol and p-chloromercuric sulfonic acid, which modify arginine and cysteine residues. The purified Bacteroidal salt-active acharan sulfate lyase acted to the greatest extent on acharan sulfate, to a lesser extent on heparan sulfate and heparin. The biochemical properties of the purified salt-active acharan sulfate lyase are different from those of the previously purified heparin lyases. However, these findings suggest that the purified salt-active acharan sulfate lyase may belong to heparin lyase II.

Purification and characterization of novel chondroitin ABC and AC lyases from Bacteroides stercoris HJ-15, a human intestinal anaerobic bacterium.

Two novel chondroitinases, chondroitin ABC lyase (EC 4.2.2.4) and chondroitin AC lyase (EC 4.2.2.5), have been purified from Bacteroides stercoris HJ-15, which was isolated from human intestinal bacteria with glycosaminoglycan degrading enzymes. Chondroitin ABC lyase was purified to apparent homogeneity by a combination of QAE-cellulose, CM-Sephadex C-50, hydroxyapatite and Sephacryl S-300 column chromatography with a final specific activity of 45.7 micromol.min-1.mg-1. Chondroitin AC lyase was purified to apparent homogeneity by a combination of QAE-cellulose, CM-Sephadex C-50, hydroxyapatite and phosphocellulose column chromatography with a final specific activity of 57.03 micromol.min-1.mg-1. Chondroitin ABC lyase is a single subunit of 116 kDa by SDS/PAGE and gel filtration. Chondroitin AC lyase is composed of two identical subunits of 84 kDa by SDS/PAGE and gel filtration. Chondroitin ABC and AC lyases showed optimal activity at pH 7.0 and 40 degrees C, and 5.7-6.0 and 45-50 degrees C, respectively. Both chondroitin lyases were potently inhibited by Cu2+, Zn2+, and p-chloromercuriphenyl sulfonic acid. The purified Bacteroidal chondroitin ABC lyase acted to the greatest extent on chondroitin sulfate A (chondroitin 4-sulfate), to a lesser extent on chondroitin sulfate B (dermatan sulfate) and C (chondroitin 6-sulfate). The purified chondroitin AC lyase acted to the greatest extent on chondroitin sulfate A, and to a lesser extent on chondroitin C and hyaluronic acid. They did not act on heparin and heparan sulfate. These findings suggest that the biochemical properties of these purified chondroitin lyases are different from those of the previously purified chondroitin lyases.