This study was aimed to investigate the chemical compositions of the essential oils from Goniothalamus macrophyllus and Goniothalamus malayanus growing in Malaysia. The essential oils were obtained by hydrodistillation and fully characterized by gas chromatography (GC-FID) and gas chromatography-mass spectrometry (GC-MS). Analyses of the essential oils from G. macrophyllus and G. malayanus resulted in 93.6 and 95.4% of the total oils, respectively. The major components of G. macrophyllus oil were germacrene D (25.1%), bicyclogermacrene (11.6%), α-copaene (6.9%) and δ-cadinene (6.4%), whereas in G. malayanus oil bicyclogermacrene (43.9%), germacrene D (21.1%) and β-elemene (8.4%) were the most abundant components.
Matched MeSH terms: Gas Chromatography-Mass Spectrometry
Polyalthia is one of the largest genera in the Annonaceae family, and has been widely used in folk medicine for the treatment of rheumatic fever, gastrointestinal ulcer, and generalized body pain. The present investigation reports on the extraction by hydrodistillation and the composition of the essential oils of four Polyalthia species (P. sumatrana, P. stenopetalla, P. cauliflora, and P. rumphii) growing in Malaysia. The chemical composition of these essential oils was determined by gas chromatography (GC-FID) and gas chromatography-mass spectrometry (GC-MS). The multivariate analysis was determined using principal component analysis (PCA) and hierarchical clustering analysis (HCA) methods. The results revealed that the studied essential oils are made up principally of bicyclogermacrene (18.8%), cis-calamenene (14.6%) and β-elemene (11.9%) for P. sumatrana; α-cadinol (13.0%) and δ-cadinene (10.2%) for P. stenopetalla; δ-elemene (38.1%) and β-cubebene (33.1%) for P. cauliflora; and finally germacrene D (33.3%) and bicyclogermacrene for P. rumphii. PCA score and HCA plots revealed that the essential oils were classified into three separated clusters of P. cauliflora (Cluster I), P. sumatrana (Cluster II), and P. stenopetalla, and P. rumphii (Cluster III) based on their characteristic chemical compositions. Our findings demonstrate that the essential oil could be useful for the characterization, pharmaceutical, and therapeutic applications of Polyalthia essential oil.
Matched MeSH terms: Gas Chromatography-Mass Spectrometry
Chemical composition and anticholinesterase activity of the essential oil of Pavetta graciliflora Wall. ex Ridl. (Rubiaceae) was examined for the first time. The essential oil was obtained by hydrodistillation and was fully characterized by gas chromatography (GC) and gas chromatography-mass spectrometry (GC-MS). A total of 20 components were identified in the essential oil, which made up 92.85% of the total oil. The essential oil is composed mainly of β-caryophyllene (42.52%), caryophyllene oxide (25.33%), β-pinene (8.67%), and α-pinene (6.52%). The essential oil showed weak inhibitory activity against acetylcholinesterase (AChE) (I%: 62.5%) and butyrylcholinesterase (BChE) (I%: 65.4%) assays. Our findings were shown to be very useful for the characterization, pharmaceutical, and therapeutic applications of the essential oil from P. graciliflora.
Matched MeSH terms: Gas Chromatography-Mass Spectrometry
A method has been developed for the determination of trace levels of alpha-endosulfan, beta-endosulfan, endosulfan sulfate, and endosulfan diol in rat plasma and tissue samples. Endosulfan and its metabolites in the plasma samples were extracted with solid-phase extraction Chromabond-end-capped C18 cartridges and analyzed by a Shimadzu QP-5050A gas chromatograph-mass spectrometer (GCMS) with quadrupole detector in selected-ion-monitoring mode. The analysis of endosulfan and its metabolites in liver and kidney samples involved solvent extraction, Florisil solid-phase-extraction cleanup, and quantitation by GCMS. Recovery experiments for the plasma and tissue samples were conducted over concentration ranges of 10-100 ng mL(-1) and 100-1000 ng mL(-1), respectively. The method was applied to the analysis of trace levels of endosulfan and its metabolites in plasma and tissue samples collected from an animal study. Trace levels of alpha-endosulfan and beta-endosulfan in the ranges of undetectable to 3.11 microg g(-1) and undetectable to 1.19 microg g(-1), respectively, were detected in the kidney samples, whereas trace levels of endosulfan sulfate in the range of 0.02-0.22 microg g(-1) were detected in the liver samples of rats. Neither endosulfan nor its metabolites was detected in any of the plasma samples.
Matched MeSH terms: Gas Chromatography-Mass Spectrometry
Among several animals, Rattus rattus (rat) lives in polluted environments and feeds on organic waste/small invertebrates, suggesting the presence of inherent mechanisms to thwart infections. In this study, we isolated gut bacteria of rats for their antibacterial activities. Using antibacterial assays, the findings showed that the conditioned media from selected bacteria exhibited bactericidal activities against Gram-negative (Escherichia coli K1, Klebsiella pneumoniae, Pseudomonas aeruginosa, Serratia marcescens, and Salmonella enterica) and Gram-positive (Bacillus cereus, methicillin-resistant Staphylococcus aureus, and Streptococcus pyogenes) pathogenic bacteria. The conditioned media retained their antibacterial properties upon heat treatment at boiling temperature for 10 min. Using MTT assays, the conditioned media showed minimal cytotoxic effects against human keratinocyte cells. Active conditioned media were subjected to tandem mass spectrometry, and the results showed that conditioned media from Bacillus subtilis produced a large repertoire of surfactin and iturin A (lipopeptides) molecules. To our knowledge, this is the first report of isolation of lipopeptides from bacteria isolated from the rat gut. In short, these findings are important and provide a platform to develop effective antibacterial drugs.
Antimicrobial irradiated seaweed-neem biocomposite films were synthesized in this study. The storage functional properties of the films were investigated. Characterization of the prepared films was conducted using SEM, FT-IR, contact angle, and antimicrobial test. The macroscopic and microscopic including the analysis of the functional group and the gas chromatography-mass spectrometry test revealed the main active constituents present in the neem extract, which was used an essential component of the fabricated films. Neem leaves' extracts with 5% w/w concentration were incorporated into the matrix of seaweed biopolymer and the seaweed-neem bio-composite film were irradiated with different dosages of gamma radiation (0.5, 1, 1.5, and 2 kGy). The tensile, thermal, and the antimicrobial properties of the films were studied. The results revealed that the irradiated films exhibited improved functional properties compared to the control film at 1.5 kGy radiation dosage. The tensile strength, tensile modulus, and toughness exhibited by the films increased, while the elongation of the irradiated bio-composite film decreased compared to the control film. The morphology of the irradiated films demonstrated a smoother surface compared to the control and provided surface intermolecular interaction of the neem-seaweed matrix. The film indicated an optimum storage stability under ambient conditions and demonstrated no significant changes in the visual appearance. However, an increase in the moisture content was exhibited by the film, and the hydrophobic properties was retained until nine months of the storage period. The study of the films antimicrobial activities against Staphylococcus aureus (SA), and Bacillus subtilis (BS) indicated improved resistance to bacterial activities after the incorporation of neem leaves extract and gamma irradiation. The fabricated irradiated seaweed-neem bio-composite film could be used as an excellent sustainable packaging material due to its effective storage stability.
Matched MeSH terms: Gas Chromatography-Mass Spectrometry
This study was designed to examine the chemical composition and anticholinesterase inhibitory activity of the essential oil of Pseuduvaria macrophylla (Oliv.) Merr. (Annonaceae) from Malaysia. The essential oil was obtained by hydrodistillation and fully analyzed by gas chromatography (GC) and gas chromatography-mass spectrometry (GC-MS). The analysis led to the identification of thirty-four chemical components that represented 87.7 ± 0.5% of the total oil. The essential oil was found to be rich in germacrene D (21.1 ± 0.4%), bicyclogermacrene (10.5 ± 0.5%), δ-cadinene (5.6 ± 0.2%), α-copaene (5.1 ± 0.3%), and α-cadinol (5.0 ± 0.3%). Anticholinesterase activity was evaluated using Ellman method. The essential oil showed weak inhibitory activity against acetylcholinesterase (I%: 32.5%) and butyrylcholinesterase (I%: 35.4%) assays. Our findings demonstrate that the essential oil could be very useful for the characterization, pharmaceutical and therapeutic applications of the essential oil from Pseuduvaria macrophylla.
Matched MeSH terms: Gas Chromatography-Mass Spectrometry
A new sol-gel hybrid methyltrimethoxysilane-chloropropyltriethoxysilane was prepared as sorbent for solid-phase extraction. The extraction efficiency of the prepared sol-gel hybrid methyltrimethoxysilane-chloropropyltriethoxysilane was assessed by using three selected organophosphorus pesticides, namely, chlorpyrifos, profenofos, and malathion. Gas chromatography-mass spectrometry was used for detection of organophosphorus pesticides. Several vital parameters were optimized to identify the best extraction conditions. Under the optimum extraction conditions, solid-phase extraction-methyltrimethoxysilane-chloropropyltriethoxysilane method showed good linearity range (0.05-1 μg/mL) with coefficient of determination more than 0.995. The limits of detection obtained were in the range of 0.01-0.07 μg/mL and limits of quantification ranging from 0.03 to 0.21 μg/mL. The limits of detection obtained for the developed method were 2.3-6.5× lower than the limits of detection of commercial octadecyl silica sorbent. Real samples analysis was carried out by applying the developed method on red apple and purple grape samples. The developed method exhibited good recoveries (88.33-120.7%) with low relative standard deviations ranging from 1.6 to 3.3% compared to commercial octadecyl silica sorbent, which showed acceptable recoveries (70.3-100.2%) and relative standard deviations (6.3-8.8%). The solid-phase extraction-methyltrimethoxysilane-chloropropyltriethoxysilane method is presented as an alternative extraction method for determination of organophosphorus pesticides.
Matched MeSH terms: Gas Chromatography-Mass Spectrometry
Female breast cancer is one of the leading causes of female mortality worldwide. In Malaysia, breast cancer is the most commonly diagnosed cancer in women. Of the women in Malaysia, the Chinese have the highest number of breast cancer cases, followed by the Indian and the Malay. The most common type of breast cancer is infiltrating ductal carcinoma (IDC). A proteomic approach was applied in this study to identify changes in the protein profile of cancerous tissues compared with normal tissues from 18 patients; 8 Chinese, 6 Malay and 4 Indian were analysed. Twenty-four differentially expressed hydrophilic proteins were identified. We evaluated the potential of these proteins as biomarkers for infiltrating ductal carcinoma based on their ethnic-specific expressions. Three of the upregulated proteins, calreticulin, 14-3-3 protein zeta and 14-3-3 protein eta, were found to be expressed at a significantly higher level in the cancerous breast tissues when compared with the normal tissues in cases of infiltrating ductal carcinoma. The upregulation in expression was particularly dominant in the Malay cohort.
Mud lobsters are crustaceans from the genus Thalassina which are lesser known and seldom seen but are nevertheless an important organism to the mangrove ecosystem. In Malaysia and Thailand, mud lobsters are eaten by locals as treatment for asthma. It is traditionally believed that they are effective in reducing the number of asthma attacks and severity of asthma symptoms. However, the therapeutic potential of mud lobster extract remains unclear and has not been fully elucidated or reported in any scientific study. The objectives of this study are to investigate the anti-inflammatory potential of mud lobster, Thalassina anomala extracts (hexane, chloroform and methanol) in lipopolysaccharide (LPS)-stimulated RAW 264.7 macrophages, and to identify the potential bioactive compounds involved. An MTT assay was performed to determine the cytotoxicity of the T. anomala extracts on RAW 264.7 macrophages. Nitrite quantification assay and enzyme-linked immunosorbent assay (ELISA) were conducted to investigate the ability of the T. anomala extracts to suppress the secretion and expression of nitric oxide (NO), Prostaglandin E2 (PGE2) and proinflammatory cytokines (TNF-α, IL-6 and IL-1β) in LPS-stimulated macrophages. GC-MS analysis was done to identify putative metabolites. The hexane extract of T. anomala showed anti-inflammatory activity by significantly inhibiting the LPS-induced production of NO, PGE2, interleukin- (IL-) 6, IL-1β and tumour necrosis factor-alpha (TNF-α) in a concentration-dependent manner. Hexane extract treatment with 100 μg/mL has decreased the NO secretion into 37 μM. Meanwhile, hexane extract at concentration of 100 μg/mL able to significantly suppressed PGE2,TNF-α, IL-6 and IL-1β production into 2015 pg/mL, 2406 pg/mL, 460 pg/mL and 9.6 pg/mL, respectively. GC-MS analysis of the hexane extract revealed the presence of 19 putative compounds. The identified compounds were reported to have anti-inflammatory, antioxidant and antibacterial activities. These results suggest that the hexane extract of T. anomala potentially has anti-inflammatory properties and concentration dependently suppressed NO, PGE2 and proinflammatory cytokines' production in LPS-stimulated macrophages. The findings provide a rational basis of the traditional use of mud lobster for inflammation-associated ailments.
Matched MeSH terms: Gas Chromatography-Mass Spectrometry
Nutritional quality of the hepatopancreas and gonads of orange portunid mud crab, Scylla olivacea was evaluated for each gender under four treatment of different water velocities (0, 20, 40 and 60 cm s-1), in terms of nutrient reserve and nutrient for reproduction. About 56 crabs were used in this study in which fatty acids composition was analysed using gas chromatography mass-spectrometry (GC-MS). For hepatopancreas analysis, monounsaturated fatty acids (MUFAs) were present in the highest fatty acids concentration, followed by polyunsaturated fatty acids (PUFAs) and, saturated fatty acids (SFAs). However, long-chain polyunsaturated fatty acids (LC-PUFAs) were displayed in low concentration in the hepatopancreas. Total fatty acid (TFAs) composition was significantly higher at moderate velocity of 20 cm s-1 compared to other water velocity treatments. For gonad analysis, 20 cm s-1 showed the highest TFA concentration of 93.34 mg g-1 while, the lowest concentration of 3.90 mg g-1 occurred at 0 cm s-1. There were significant differences in male and female crab's fatty acids contents of gonads at all flow velocities challenged (p < 0.05). PUFAs and MUFAs were dominant while, SFAs were observed at low concentration. This study revealed that, concentration of PUFAs increased as gonad maturation increased. The decreasing concentration of hepatopancreas fatty acids over the culture period indicated that nutrient was shifted from the hepatopancreas, to be used as energy reserved to gonads for further growth of eggs and offspring. The linkages between water flow strength, hepatopancreas, and gonad fatty acids concentrations, is fundamental knowledge useful in establishing efficient habitat velocities selection which will improve aquaculture production of mud crabs with high quality broodstock.
Matched MeSH terms: Gas Chromatography-Mass Spectrometry
Mycotoxins are naturally occurring food toxins worldwide that can cause serious health effects. The measurement of mycotoxin biomarkers in biological fluids is needed to assess individuals' exposure. The aim of this study was to investigate the incidence of mycotoxins in the Qatari population. Serum samples from 412 adults and urinary samples from 559 adults were analyzed for the presence of mycotoxin biomarkers. Multimycotoxin approaches have been applied, using liquid chromatography mass spectrometry methods. Samples were further analyzed for the oxidative stress markers and compared with regard to the incidence of mycotoxins. The presence of mycotoxins was identified in 37% of serum samples and in less than 20% of urine samples. It was found that 88% of positive of the samples were positive for only one mycotoxin, while 12% of positive samples had two or more mycotoxins. Trichothecenes and zearalenone metabolites were most commonly detected mycotoxins, followed by aflatoxins, roquefortine C and mycophenolic acid. The presence of mycotoxins was found to positively correlate with oxidative stress markers. The obtained results illustrate the importance of mycotoxin biomonitoring studies in humans and the need to elucidate the underlying mechanisms of mycotoxin-induced toxicity.
The study aimed to establish the detection method for bound 3-, 2-MCPD, and glycidol using accelerated solvent extraction (ASE) and gas chromatography mass spectrometry (GC-MS). The ASE was modified for reduced solvent volume and process time to extract lipid from the chocolate spread, infant formula, potato chips, and sweetened creamer. The solvent selected for ASE was a mixture of iso-hexane and acetone at 100°C with the lipid and analyte recovery ranging from 96.9% to 98.6% and 84.1% to 107.5%, respectively. The derivatisation of analytes was adopted from the AOCS method Cd29a-13 for GC-MS analysis. The results showed that the coefficient of determination (R2) of all analytes was >0.99. The limit of detection (LOD) was 0.1 mg kg-1 expressed in lipid basis for both bound 3- and 2-MCPD and 0.2 mg kg-1 expressed in lipid basis for bound glycidol. The limit of quantitation (LOQ) was 0.3 mg kg-1 expressed in lipid basis for both bound 3- and 2-MCPD and 0.6 mg kg-1 expressed in lipid basis for bound glycidol. A blank spiked with 3-monochloropropanediols fatty acid esters (MCPDE) and 2-MCPDE (0.3, 2.1, and 7.2 mg kg-1) and glycidol esters (0.6, 4.7, and 16.6 mg kg-1) were chosen for accuracy and precision tests. The recoveries were 91.7% to 105.9%. Both repeatability and within-laboratory reproducibility of the analysis were within the acceptable level of precision ranging from 1.7% to 16%. This is the first time that a full validation procedure extending to both accuracy and precision tests has been carried out for sweetened creamer and chocolate spread. Overall, the combined protocol of ASE and AOCS Cd29a-13 was successfully validated for both solid and liquid food samples with lipid content from 10% to 30%.
Matched MeSH terms: Gas Chromatography-Mass Spectrometry
The chemical composition of the essential oil of Knema kunstleri Warb. (Myristicaceae) was investigated for the first time. The essential oil was obtained by hydrodistillation and fully characterized by gas chromatography (GC) and gas chromatography-mass spectrometry (GC-MS). In total, 36 components were identified in the essential oil, which made up 91.7% of the total oil. The essential oil is composed mainly of β-caryophyllene (23.2%), bicyclogermacrene (9.6%), δ-cadinene (7.3%), α-humulene (5.7%), and germacrene D (4.3%). The essential oil showed moderate activity towards DPPH free-radical scavenging and lipoxygenase inhibition. To the best of our knowledge, this is the first study of the composition and bioactivities of the essential oil report concerning the genus Knema.
Matched MeSH terms: Gas Chromatography-Mass Spectrometry
The hump-nosed pit viper, Hypanle hypnale, contributes to snakebite mortality and morbidity in Sri Lanka. Studies showed that the venom is hemotoxic and nephrotoxic, with some biochemical and antigenic properties similar to the venom of Calloselasma rhodostoma (Malayan pit viper). To further characterize the complexity composition of the venom, we investigated the proteome of a pooled venom sample from >10 Sri Lankan H. hypnale with reverse-phase high performance liquid chromatography (rp-HPLC), sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and peptide sequencing (tandem mass-spectrometry and/or N-terminal sequencing). The findings ascertained that two phospholipase A2 subtypes (E6-PLA2, W6-PLA2) dominate the toxin composition by 40.1%, followed by snake venom metalloproteases (36.9%), l-amino acid oxidase (11.9%), C-type lectins (5.5%), serine proteases (3.3%) and others (2.3%). The presence of the major toxins correlates with the venom's major pathogenic effects, indicating these to be the principal target toxins for antivenom neutralization. This study supports the previous finding of PLA2 dominance in the venom but diverges from the view that H. hypnale venom has low expression of large enzymatic toxins. The knowledge of the composition and abundance of toxins is essential to elucidate the pathophysiology of H. hypnale envenomation and to optimize antivenom formulation in the future.
Screening of the occurrence of potentially toxic diatoms was carried out at two sites of cage cultures in Tebrau Straits, Johor. Phytoplankton samples from Sungai Pendas and Teluk Sengat were collected using a 20 μm mesh plankton net and salinity was recorded in-situ. Nitzschia and Pseudo-nitzschia cells were isolated and established into clonal cultures. All cultures were tested for domoic acid using HPLC-UV analysis and verified by LC-MS analysis. Three Nitzschia spp. and one Pseudo-nitzschia sp. were identified from these locations. Toxic and non-toxic strains of Nitzschia navis-varingica are found at the cage culture areas. Cellular toxin content in the toxic strain of N. navis-varingica is 1.8 pg cell(-1). This is a new record from Malaysia and this species was isolated from estuarine water with salinity 28 PSU. The discovery of toxic Nitzschia species in Tebrau Straits indicates the potential for domoic acid accumulation in seafood.
The protein profile of serum samples from patients with amoebic liver abscess (ALA) was compared to those of normal individuals to determine their expression levels and to identify potential surrogate disease markers. Serum samples were resolved by two dimensional electrophoresis (2-DE) followed by image analysis. The up and down-regulated protein spots were excised from the gels and analysed by MS/MS. The concentration of three clusters of proteins i.e. haptoglobin (HP), α1-antitrypsin (AAT) and transferrin in serum samples of ALA patients and healthy controls were compared using competitive ELISA. In addition, serum concentrations of HP and transferrin in samples of patients with ALA and pyogenic liver abscess (PLA) were also compared. The results of the protein 2-DE expression analysis showed that HP cluster, AAT cluster, one spot each from unknown spots no. 1 and 2 were significantly up-regulated and transferrin cluster was significantly down-regulated in ALA patients' sera (p<0.05). The MS/MS analysis identified the unknown protein spot no.1 as human transcript and haptoglobin and spot no. 2 as albumin. Competitive ELISA which compared concentrations of selected proteins in sera of ALA and healthy controls verified the up-regulated expression (p<0.05) of HP and the down-regulated expression (p<0.01) of transferrin in the former, while there was no significant difference in AAT expression (p> 0.05). However, when ALA and PLA samples were compared, competitive ELISA showed significant increased concentration of HP (p<0.05) while transferrin levels were not different. In conclusion, this study showed that HP is a potential surrogate disease marker for ALA.
The study was aimed to investigate the expression of cytosolic and thiolated proteins of Musca domestica larvae under oxidative stress. Proteins from acute treatment of hydrogen peroxide (LC50 = 21.52% (v/v)) on 3rd stage larvae of housefly were extracted and purified using an activated Thiol Sepharose® for thiolated protein purification. Two dimensional gel electrophoresis was used for visualizing and analyzing expression of cytosolic and thiolated proteins. Protein spots with more than 5 fold of expression change were identified using liquid chromatography- tandem mass spectrometry (LC-MS/MS). The cytosolic proteins were actin, tropomyosin, ubiquitin, arginine kinase, pheromone binding protein/general odorant binding protein, and ATP: guanidino phosphotransferase. The thiolated proteins with more than 5 fold change in expression as an effect to the acute treatment were fructose bisphosphate aldolase, short chain dehydrogenase and lactate/malate dehydrogenase. The proteins identified in the study should provide vital information for future reference in oxidative stress defence and response occurring in houseflies.
The bioactivity of R. nasutus leaf extracts was assessed on Bacillus cereus, Bacillus subtilis, Staphylococcus aureus, Streptococcus pyogenes, Vibrio parahaemolyticus, Enterobacter aerogenes, Proteus mirabilis, and Klebsiella pneumoniae. Crude chloroform, petroleum ether, ethyl acetate, ethanol and methanol extracts were screened by disc diffusion method. Promising crude extract was further subjected to the column fractionation followed by the screening of the antibacterial activity of individual fractions. Biologically active pure fraction was subjected to the advanced analytical studies like HPLC, LC-MS, IR and NMR for characterisation of the bioactive compound. Ethanolic extract exhibited the maximum antibacterial activity against Klebsiella pneumoniae with the maximum of 35±0.42 mm zone of inhibition. The biologically potent column fraction from ethanol extract with 40±0.42 mm zone of inhibition upon subject to the HPLC, LC-MS, IR and NMR revealed that the active compound is rhinacanthin-C, a naphthoquinone.
Livestock industries have maintained a keen interest in pasture legumes because of the high protein content and nutritive value. Leguminous Indigofera plant species have been considered as having high feeding values to be utilized as pasture, but the occurrence of the toxic constituent indospicine in some species has restricted this utility. Indospicine has caused both primary and secondary hepatotoxicosis and also reproductive losses, but has only previously been determined in a small number of Indigofera species. This paper validates a high-throughput ultraperformance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method to determine the indospicine content of various Indigofera species found in Australian pasture. Twelve species of Indigofera together with Indigastrum parviflorum plants were collected and analyzed. Of the 84 samples analyzed, *I. spicata (the asterisk indicates a naturalized species) contained the highest indospicine level (1003 ± 328 mg/kg DM, n = 4) followed by I. linnaei (755 ± 490 mg/kg DM, n = 51). Indospicine was not detected in 9 of the remaining 11 species and at only low levels (<10 mg/kg DM) in 2 of 8 I. colutea specimens and in 1 of 5 I. linifolia specimens. Indospicine concentrations were below quantitation levels for other Indigofera spp. (I. adesmiifolia, I. georgei, I. hirsuta, I. leucotricha, *I. oblongifolia, I. australis, and I. trita) and Indigastrum parviflorum. One of the more significant findings to emerge from this study is that the indospicine content of I. linnaei is highly variable (from 159 to 2128 mg/kg DM, n = 51) and differs across both regions and seasons. Its first regrowth after spring rain has a higher (p < 0.01) indospicine content than growth following more substantial summer rain. The species collected include the predominant Indigofera in Australia pasture, and of these, only *I. spicata and I. linnaei contain high enough levels of indospicine to pose a potential toxic threat to grazing herbivores.