Mass spectral analysis of secondary metabolites from Zingiber montanum rhizome extract using UHPLC-HR-ESI-QTOF-MS/MS.

INTRODUCTION: Zingiber montanum (J.Koenig) Link ex A.Dietr. is a popular medicinal plant in Thailand. Its rhizomes have been used as an ingredient in various Thai traditional medicine formulas. While many reports have focused on the chemical constituents and biological activities of this plant, a comprehensive study on secondary metabolite profiling using tandem mass spectrometry has, to this point, never been documented. OBJECTIVE: To analyze the chemical constituents in Z. montanum rhizomes using ultra-high performance liquid chromatography coupled with ultra-high-resolution electrospray ionization quadrupole time-of-flight tandem mass spectrometry (UHPLC-HR-ESI-QTOF-MS/MS) analyses and to utilize the characteristic fragmentation patterns of these compounds to facilitate their identification. METHODOLOGY: UHPLC-HR-ESI-QTOF-MS/MS in positive ion mode was used for chemical identification of secondary metabolites from the ethanolic extract of the plant material. MS/MS data of some known reference compounds, together with detailed fragmentation pattern information of several compounds obtained from the crude extract, were used to elucidate their chemical structures. RESULTS: In this work, one benzaldehyde, ten phenylbutenoid monomers, six curcuminoids, and nine phenylbutenoid dimers were assigned based on their characteristic fragment ions. Among these compounds, 2-(3,4-dimethoxystyryl)oxirane was tentatively suggested as a potential new compound. Several characteristic fragment ions from these compounds were assigned and the relative ion abundance of these was also used to differentiate the chemical structures of compounds having the same molecular mass. CONCLUSIONS: The results will benefit future high-throughput screening of bioactive compounds and method development for the quality control of raw materials and herbal drugs derived from Z. montanum rhizome extracts.

Phytochemicals and Bioactivities of Zingiber cassumunar Roxb.

Zingiber cassumunar Roxb. (Zingiberaceae), is an important medicinal plant known as "Plai (Phlai)" in Thailand, "Bangle" in Indonesia, and "Bulei" in China. Traditionally, this plant has been used to treat inflammation, pain, and respiratory problems. The rhizomes are the primary part of the plant that has been used for medicinal purposes due to their constituents with therapeutic properties, including phenylbutenoids, curcuminoids, and essential oils. Since the 1970s, many studies have been conducted on the phytochemicals and bioactivities of Z. cassumunar to establish fundamental scientific evidence that supports its use in traditional medicine. The accumulated biological studies on the extracts, solvent fractions, and constituents of Z. cassumunar have described their diverse medicinal properties, including antioxidant, anti-inflammatory, anticancer, neuroprotective/neurotrophic, cosmeceutical, and antifungal/antimicrobial bioactivities. In this review, we summarize information on the phytochemicals of Z. cassumunar and the bioactivities of its extracts and constituents.

Development of Microemulsion Delivery System of Essential Oil from Zingiber cassumunar Roxb. Rhizome for Improvement of Stability and Anti-Inflammatory Activity.

The present study aims to investigate the major constituents of the essential oil from Zingiber cassumunar rhizome (EO) and to develop microemulsions with enhanced chemical stability and anti-inflammatory activity of EO. The major constituents of EO were terpinen-4-ol (40.5 ± 6.6%) and sabinene (17.4 ± 1.4%) as determined by gas chromatography-mass spectrometry. These compounds were responsible for the anti-inflammatory activities of EO. Sabinene and terpinen-4-ol significantly reduced nuclear factor-kappa B (NF-kB) expression by 47 ± 5 and 78 ± 8%, respectively (p < 0.001) and significantly reduced the interleukin-6 (IL-6) secretion levels to 64 ± 4% (p < 0.05) and 50 ± 1% (p < 0.001), respectively. EO microemulsions, developed using the system of EO/Tween 20 and propylene glycol (2:1)/water, showed the internal droplet size in the range of 211.5 ± 63.3 to 366.7 ± 77.8 nm. Both EO and EO microemulsions were shown to be safe for human use since there was no apparent toxic effect on human peripheral blood mononuclear cells. Interestingly, EO microemulsion could significantly protect sabinene from the evaporation after heating-cooling stability test, which leads to a good stability and high efficacy. Moreover, EO microemulsions significantly enhanced the anti-inflammatory effect comparing to the native EO. Therefore, microemulsions were attractive delivery system for natural anti-inflammatory compounds since they could enhance both efficacy and stability of EO.

Topical Niosome Gel of Zingiber cassumunar Roxb. Extract for Anti-inflammatory Activity Enhanced Skin Permeation and Stability of Compound D.

An extract of Zingiber cassumunar Roxb. (ZC) was encapsulated in niosomes of which a topical gel was formed. (E)-4-(3',4'-dimethoxyphenyl)but-3-en-1-ol or compound D detected by a gradient HPLC was employed as the marker and its degradation determined to follow zero-order kinetics. Niosomes significantly retarded thermal-accelerated decomposition of compound D in the gel (p < 0.05) but did not change the activation energy of compound D. Niosomes enhanced in vitro permeation rate of compound D from the gel. Topical applications of ZC noisome gel gave a faster change in tail flick latency than piroxicam gel and hydrocortisone cream (p < 0.05) while there were insignificant differences in anti-inflammatory activity up to 6 h using croton oil-induced ear edema model in mice (p > 0.05). Thus, encapsulation of ZC extract in niosomes enhanced chemical stability and skin permeation with comparable topical anti-inflammatory effects to steroid and NSAID.

Zingiber cassumunar Roxb. Essential Oil-Loaded Electrospun Poly(lactic acid)/Poly(ethylene oxide) Fiber Blend Membrane for Antibacterial Wound Dressing Application.

The essential oil from Zingiber cassumunar Roxb. (Plai) has long been used in Thai herbal remedies to treat inflammation, pains, sprains, and wounds. It was therefore loaded into an electrospun fibrous membrane for use as an analgesic and antibacterial dressing for wound care. The polymer blend between poly(lactic acid) and poly(ethylene oxide) was selected as the material of choice because its wettability can be easily tuned by changing the blend ratio. Increasing the hydrophilicity and water uptake ability of the material while retaining its structural integrity and porosity provides moisture balance and removes excess exudates, thereby promoting wound healing. The effect of the blend ratio on the fiber morphology and wettability was investigated using scanning electron microscopy (SEM) and contact angle measurement, respectively. The structural determination of the prepared membranes was conducted using Fourier-transform infrared spectroscopy (FTIR). The release behavior of (E)-1-(3,4-dimethoxyphenyl) butadiene (DMPBD), a marker molecule with potent anti-inflammatory activity from the fiber blend, showed a controlled release characteristic. The essential oil-loaded electrospun membrane also showed antibacterial activity against S. aureus and E. coli. It also exhibited no toxicity to both human fibroblast and keratinocyte cells, suggesting that the prepared material is suitable for wound dressing application.

Repellent, larvicidal and adulticidal activities of essential oil from Dai medicinal plant Zingiber cassumunar against Aedes albopictus.

Zingiber cassumunar is an important plant used in traditional medicine and as a natural mosquito repellent. However, the compounds responsible for the repellent activity of the plant are still unknown. The aim of the study is to identify the components of Z. cassumunar essential oil that show repellent activity against Aedes albopictus. We also evaluated the larvicidal and adulticidal activities of Z. cassumunar essential oil against Ae. albopictus. In-cage mosquito repellent experiments showed that Z. cassumunar essential oil possessed moderate repellent activity with a minimum effective dose (MED) of 0.16 ± 0.01 mg/cm2, compared to reference standard N,N-diethyl-3-methylbenzamide (DEET, 0.03 ± 0.01 mg/cm2). Bioassay-guided fractionation identified the major active compound of Z. cassumunar essential oil as (-)-terpinen-4-ol (1) (MED: 0.19 ± 0 mg/cm2). We also found that Z. cassumunar essential oil showed moderate larvicidal activity against first instar larvae of Ae. albopictus with a LC50 (50% lethal concentration) of 44.9 µg/L after 24 h. Fumigation bioassays showed that Z. cassumunar essential oil exhibits moderate adulticidal activity against Ae. albopictus with a LC50 of 5.44%, while (-)-terpinen-4-ol showed significant adulticidal activity with a LC50 of 2.10% after 24 h. This study verifies that the Z. cassumunar essential oil has mosquito repellent activity, and that (-)-terpinen-4-ol is mainly responsible for this activity. Furthermore, this study provides scientific support for the folk usage of Z. cassumunar essential oil as mosquito repellent and indicates that Z. cassumunar essential oil and (-)-terpinen-4-ol can be used as plant-derived repellents and insecticides for mosquito control.

Antibacterial secondary metabolites from an endophytic fungus, Arthrinium sp. MFLUCC16-1053 isolated from Zingiber cassumunar.

Forty-four endophytes were isolated from Zingiber cassumunar and identified morphologically. The ethyl acetate extracts of all endophytes were obtained. The ethyl acetate extracts were subjected to study antibacterial and antioxidant activities. The ethyl acetate extract of the Arthrinium sp. MFLUCC16-1053 showed activity against both gram-positive and gram-negative bacteria. Specifically, the minimum inhibition concentration against Staphylococcus aureus and Escherichia coli was 31.25 and 7.81 µg/mL, respectively. In addition, the crude extract showed highest antioxidant activity in 2,2-diphenyl-1-picrylhydrazyl scavenging at IC50 value of 28.47 µg/mL. Gas chromatography-mass spectrometry analysis revealed that the extract of the Arthrinium sp. MFLUCC16-1053 contains various antibacterial and antioxidant compounds which are ß-cyclocitral, 3E-cembrene A, laurenan-2-one, sclareol, 2Z,6E-farnesol, cembrene, ß-isocomene and γ-curcumene. The fungus Arthrinium sp. MFLUCC16-1053 was also identified by molecular and phylogenetic methods.

Clinical effects of Zingiber cassumunar (Plai): A systematic review.

Zingiber cassumunar Roxb. known locally as "Plai" in Thai, has been used for treating bruise, sprain and musculoskeletal pain. Several pre-clinical studies demonstrated the anti-inflammatory effect of Plai. However, current evidence of clinical effects of Plai is still unclear. This study aimed to determine the clinical efficacy and safety of Plai among all identified indications. Of the 808 articles identified by a systematic review, six studies were included. Four studies were randomized controlled trials, while two studies were quasi-experimental studies involving 178 patients in intervention group and 177 patients in control group. Duration of treatment ranged from 7days to 2 months. Our findings showed that 14% Plai cream had a strong trend of benefits in pain reduction for muscle pain and ankle sprain. However, evidence supporting the effects of Plai on acne vulgaris treatment and anti-histamine effect are still unclear.

Anti-inflammatory activity of niosomes entrapped with Plai oil (Zingiber cassumunar Roxb.) by therapeutic ultrasound in a rat model.

OBJECTIVE: The aim of this study was to evaluate the antioxidant and anti-inflammatory activities of Plai oil-encapsulated niosomes (Zingiber cassumunar Roxb.) on inflamed subcutaneous Wistar rat skin by therapeutic ultrasound. METHODS: Pure oil from Plai rhizomes was extracted by steam distillation, and antioxidant activities were determined by 2,2-diphenyl-1-picrylhydrazyl radical scavenging assay. Bioactive compounds were analyzed by gas chromatography-mass spectrometry. Niosome particles containing Plai oil were prepared by chloroform film method with sonication before testing for anti-inflammatory activity on locally inflamed subcutaneous rat skin after inducement from lipopolysaccharide with ultrasound once a day for 3 days. Skin temperatures and blood flow were evaluated. RESULTS: Plai oil presented antioxidant activity that inhibited 2,2-diphenyl-1-picrylhydrazyl radicals. Four active compounds found in the essential oil were sabinene, γ-terpinene, terpinene-4-ol, and (E)-1-(3,4-dimethyoxy phenyl) butadiene. Application of ultrasound (0.2 W/cm2, 20%, 3 min) with gel containing Plai oil-encapsulated niosomes decreased skin temperature and blood flow to the lowest level compared to the application of neurofen drug or gel-based control. CONCLUSION: Plai oil, which consists of four main bioactive compounds and possesses antioxidant and anti-inflammatory activities, can be applied against local subcutaneous inflammation when used with therapeutic ultrasound via entrapped niosomes.

Active compound of Zingiber cassumunar Roxb. down-regulates the expression of genes involved in joint erosion in a human synovial fibroblast cell line.

Rheumatoid arthritis (RA) is a chronic inflammatory disease of the synovium. It is involved in up-regulation of pro-inflammatory cytokines and matrix metalloproteinases (MMPs), resulting in joint inflammation and erosion. Zingiber cassumunar Roxb. has long been used to reduce joint pain and inflammation. This study aimed to investigate the inhibitory activities of an active compound of Z. cassumunar, (E)-4-(3',4'-dimethoxyphenyl)but-3-en-1-ol (compound D), against cytokine-induced up-regulation of catabolic genes involved in cartilage degradation in RA. Synovial fibroblast cell line, SW982, was cultured in media containing interleukin-1ß (IL-1ß), in the presence or absence of compound D at the concentration range of 1 to 100 µM. After 24 hours, the cells were analyzed for the expressions of MMPs, IL-1ß and interleukin-1ß-converting enzyme (ICE) by RT-PCR. MMPs activities in the culture media were analyzed by zymographic techniques. Dexamethasone was used as the positive control. It was found that compound D at the concentration of 10 - 100 µM significantly decreased the mRNA expressions of MMP-1, -2, -3, and -13 which was induced by IL-1ß (P<0.05) concomitantly with a decrease in activities of these MMPs in the culture media. An increase in the mRNA expression of IL-1ß and ICE was also suppressed by compound D. The results suggest that the potent activities of this compound may be involved in the reduction of IL-1ß protein synthesis in both pro-form and active form which played an important role in up-regulation of MMPs. This study first revealed the chondroprotective activity of Z. cassumunar in the transcriptional level by suppressing cytokine-induced catabolic genes which caused cartilage erosion in RA.