Optimization of fecal sample preparation for untargeted LC-HRMS based metabolomics

Introduction

Collecting feces is easy. It offers direct outcome to endogenous and microbial metabolites.

Objectives

In a context of lack of consensus about fecal sample preparation, especially in animal species, we developed a robust protocol allowing untargeted LC-HRMS fingerprinting.

Methods

The conditions of extraction (quantity, preparation, solvents, dilutions) were investigated in bovine feces.

Results

A rapid and simple protocol involving feces extraction with methanol (1/3, M/V) followed by centrifugation and a step filtration (10 kDa) was developed.

Conclusion

The workflow generated repeatable and informative fingerprints for robust metabolome characterization.

     

Iridoid and phenylethanoid/phenylpropanoid metabolite profiles of <Emphasis Type="Italic">Scrophularia</Emphasis> and <Emphasis Type="Italic">Verbascum</Emphasis> species used medicinally in North America

Introduction

Botanicals containing iridoid and phenylethanoid/phenylpropanoid glycosides are used worldwide for the treatment of inflammatory musculoskeletal conditions that are primary causes of human years lived with disability, such as arthritis and lower back pain.

Objectives

We report the analysis of candidate anti-inflammatory metabolites of several endemic Scrophularia species and Verbascum thapsus used medicinally by peoples of North America.

Methods

Leaves, stems, and roots were analyzed by ultra-performance liquid chromatography-mass spectrometry (UPLC-MS) and partial least squares-discriminant analysis (PLS-DA) was performed in MetaboAnalyst 3.0 after processing the datasets in Progenesis QI.

Results

Comparison of the datasets revealed significant and differential accumulation of iridoid and phenylethanoid/phenylpropanoid glycosides in the tissues of the endemic Scrophularia species and Verbascum thapsus.

Conclusions

Our investigation identified several species of pharmacological interest as good sources for harpagoside and other important anti-inflammatory metabolites.

     

Health seeking behavior and use of medicinal plants among the Hamer ethnic group,South Omo zone,southwestern Ethiopia

Background

Health seeking behavior of people around the globe is affected by different socio-cultural and economic factors. In Ethiopia, people living in rural areas in particular, are noted for their use of medicinal plants as a major component of their health care option. This study was conducted to document ethnopharmacological information of the Hamer semi-pastoralists ethnic group in southwestern Ethiopia.

Methods

A cross-sectional study was carried out whereby information on demographic characteristics, prevalence of perceived illnesses, factors associated with preference of health care seeking options, medicinal plants used and hoarded as well as some healers’ socio-economic characteristics were collected using two sets of semi-structured questionnaires – one for household (HH) heads and the other for traditional healers supplemented by focus group discussions (FGDs). Households were selected using a cluster sampling followed by systematic sampling techniques; whereas healers and FGD participants were purposively selected with the assistance of local leaders and elders from the community.

Results

The study revealed that the use of traditional medicine among the Hamer ethnic group is very high. Females preferred traditional medicine more than males. The main reasons for this preference include effectiveness, low cost and ease of availability. Malaria (gebeze) was the most frequently occurring illness in the area identified by all FGD participants. A total of 60 different medicinal plants were reported [34 by HH respondents, 14 by traditional healers and 12 by both]. Fifty-one medicinal plants were fully identified, 3 at generic level and 6 have not yet been identified.

Conclusion

It can be concluded that traditional medical practices, particularly herbal aspect, is widely used by the Hamer ethnic group, although health seeking behavior of the community is affected by different socio-economic and cultural factors.

     

Medicinal plants used by women from Agnalazaha littoral forest (Southeastern Madagascar)

Background

The country of Madagascar is renowned for its high level of biodiversity and endemism, as well as the overwhelming pressures and threats placed on the natural resources by a growing population and climate change. Traditional medicine plays an important role in the daily lives of the Malagasy for various reasons including limited access to healthcare, limited markets and traditional values. The objective of this study was to assess the modern utitilization of the Agnalazaha Forest by the local population in Mahabo-Mananivo, Madagascar, for medicinal plants used by women, and to establish a list of medicinal plants used by women sourced from Agnalazaha Forest.

Methods

Ethnobotanical studies were conducted over a period of five months in 2010 to determine the diversity of medicinal plants used by women in the commune of Mahabo-Mananivo. In all, 498 people were interviewed, both male and female ranging age from 15 to over 60 years old.

Results

152 medicinal plants used by local people were collected during the ethnobotanical studies. Among the recorded species, eight native species are widely used by women. These species are known for their therapeutic properties in treating placental apposition and complications during childbirth as well as tropical illnesses such as malaria, filariasis, and sexual diseases like gonorrhea and syphilis.

Conclusions

Littoral forests are rare ecosystems that are highly threatened on the island nation of Madagascar. Our investigation into the use of medicinal plants sourced from and around the Agnalazaha Forest by the women of Mahabo-Mananivo reinforces the need for this natural resource as a first line of health care for rural families.

     

Sociocultural and ecological factors influencing management of edible and non-edible plants: the case of Ixcatlán,Mexico

Background

Identifying factors influencing plant management allows understanding how processes of domestication operate. Uncertain availability of resources is a main motivation for managing edible plants, but little is known about management motives of non-edible resources like medicinal and ceremonial plants. We hypothesized that uncertain availability of resources would be a general factor motivating their management, but other motives could operate simultaneously. Uncertainty and risk might be less important motives in medicinal than in edible plants, while for ceremonial plants, symbolic and spiritual values would be more relevant.

Methods

We inventoried edible, medicinal, and ceremonial plants in Ixcatlán, Oaxaca, Mexico, and conducted in-depth studies with 20 native and naturalized species per use type; we documented their cultural importance and abundance by interviewing 25 households and sampling vegetation in 33 sites. Consumption amounts and preferences were studied through surveys and free listings with 38 interviewees. Management intensity and risk indexes were calculated through PCA and their relation analyzed through regression analyses. Canonical methods allowed identifying the main sociocultural and ecological factors influencing management of plants per use type.

Results

Nearly 64, 63, and 55% of all ceremonial, edible, and medicinal wild plants recorded, respectively, are managed in order to maintain or increase their availability, embellishing environments, and because of ethical reasons and curiosity. Management intensity was higher in edible plants under human selection and associated with risk. Management of ceremonial and medicinal plants was not associated with indexes of risk or uncertainty in their availability. Other sociocultural and ecological factors influence management intensity, the most important being reciprocal relations and abundance perception.

Conclusions

Plant management through practices and collectively regulated strategies is strongly related to control of risk and uncertainty in edible plants, compared with medicinal and ceremonial plants, in which reciprocal interchanges, curiosity, and spiritual values are more important factors. Understanding how needs, worries, social relations, and ethical values influence management decisions is important to understand processes of constructing management strategies and how domestication could be started in the past and are operated at the present.

     

An ethnobotanical study of medicinal plants used in Kilte Awulaelo District,Tigray Region of Ethiopia

Background

The Ethiopian people have been dependent on traditional medicine, mainly medicinal plants, from time immemorial for control of human and animal health problems, and they still remain to be largely dependent on the practice. The purpose of the current study was to conduct ethnobotanical study to document medicinal plants used to treat diseases of human and domestic animals in Kilte Awulaelo District in the Tigray Region of Ethiopia.

Methods

Ethnobotanical data were collected between July and September 2011 through semi-structured interviews, ranking exercises and field observations. For the interviews, 72 knowledgeable informants were sampled using purposive sampling method. For the different ranking exercises, key informants were identified with the help of elders and local administrators from informants that were already involved in the interviews.

Results

The study revealed 114 medicinal plant species belonging to 100 genera and 53 families. The plants were used to treat 47 human and 19 livestock diseases. Of the species, the majority (74%) were obtained from the wild. Herbs were the most utilized plants, accounting for 44% of the species, followed by shrubs (29%). Leaf was the most commonly used plant part accounting for 42.98% of the plants, followed by roots (25.73%). Preference ranking exercise on selected plants used against abdominal pain indicated the highest preference of people for Solanum marginatum. Direct matrix ranking showed Cordia africana as the most preferred multipurpose plant in the community. Preference ranking of selected scarce medicinal plants indicated Myrica salicifolia as the most scarce species, followed by Boscia salicifolia and Acokanthera schimperi. According to priority ranking, drought was identified as the most destructive factor of medicinal plants, followed by overgrazing and firewood collection.

Conclusion

Medicinal plants are still playing significant role in the management of various human and livestock diseases in the study area with herbs taking the lead in the number of plants used in the preparation of remedies, which may be an indication of their relatively better abundance as compared to other life forms. Recurrent drought was reported to have seriously threatened medicinal plant resources in the District. Awareness is thus needed be raised among local people on sustainable utilization and management of plant resources. Ex situ and in situ conservation measures should be taken to protect the medicinal plants of the District from further destruction and special attention should be given to the medicinal plants that were indicated by preference ranking exercise as the most threatened ones.

     

Yao herbal medicinal market during the Dragon Boat Festival in Jianghua County,China

Background

The traditional medicinal markets held during the Dragon Boat Festival are common and important in China’s countryside. In Jianghua, a Yao autonomous county in Hunan Province in China, the medicinal market also plays an important role for the application, conservation, and communication of traditional Yao medicinal knowledge.

Methods

During the Dragon Boat Festival in 2016 and 2017, ethnobotanical surveys and inventories were conducted in the medicinal market of Jianghua County, and voucher plant specimens were collected, identified, and deposited in a herbarium. Quantitative analysis included measurement of frequency of occurrence for species in the marketplace and the relative importance index for the number of uses for a given species.

Results

A total of 306 plant species (249 genera, 113 families) and their related information about the medicinal market were collected. Some major findings include the following: (1) Using the whole plant as medicine is more common than other medicinal plant parts; (2) treating rheumatism and clearing inner heat are the most frequent medicinal uses; and (3) taking a medicinal bath is the most frequent modality to administer the traditional medicine. The frequency of occurrence and the relative importance index of some medicinal plants were analyzed, as well as the demographics and the number of stalls and the status of traditional Yao medicinal knowledge in Jianghua. Based on the investigation, suggestions were proposed for better protecting the medicinal market and preserving traditional medicinal knowledge in Jianghua County.

Conclusion

The medicinal market during the Dragon Boat Festival in Jianghua County possesses an important cultural value and helps to conserve the traditional Yao medicinal knowledge. The medicinal plants sold at the market showed great diversity and unique local characteristics. The medicinal market is facing some challenges in such a rapidly developing era. Cultivation of young healers and maintaining the local biodiversity might be the key solutions for the development of local medicinal market and local Yao medicinal knowledge.

     

A lost opportunity for science: journals promote data sharing in metabolomics but do not enforce it

Introduction

Data sharing is being increasingly required by journals and has been heralded as a solution to the ‘replication crisis’.

Objectives

(i) Review data sharing policies of journals publishing the most metabolomics papers associated with open data and (ii) compare these journals’ policies to those that publish the most metabolomics papers.

Methods

A PubMed search was used to identify metabolomics papers. Metabolomics data repositories were manually searched for linked publications.

Results

Journals that support data sharing are not necessarily those with the most papers associated to open metabolomics data.

Conclusion

Further efforts are required to improve data sharing in metabolomics.

     

Collection and trade of wild-harvested orchids in Nepal

Background

Wild orchids are illegally harvested and traded in Nepal for use in local traditional medicine, horticulture, and international trade. This study aims to: 1) identify the diversity of species of wild orchids in trade in Nepal; 2) study the chain of commercialization from collector to client and/or export; 3) map traditional knowledge and medicinal use of orchids; and 4) integrate the collected data to propose a more sustainable approach to orchid conservation in Nepal.

Methods

Trade, species diversity, and traditional use of wild-harvested orchids were documented during field surveys of markets and through interviews. Trade volumes and approximate income were estimated based on surveys and current market prices. Orchid material samples were identified to species level using a combination of morphology and DNA barcoding.

Results

Orchid trade is a long tradition, and illegal export to China, India and Hong Kong is rife. Estimates show that 9.4 tons of wild orchids were illegally traded from the study sites during 2008/2009. A total of 60 species of wild orchids were reported to be used in traditional medicinal practices to cure at least 38 different ailments, including energizers, aphrodisiacs and treatments of burnt skin, fractured or dislocated bones, headaches, fever and wounds. DNA barcoding successfully identified orchid material to species level that remained sterile after culturing.

Conclusions

Collection of wild orchids was found to be widespread in Nepal, but illegal trade is threatening many species in the wild. Establishment of small-scale sustainable orchid breeding enterprises could be a valuable alternative for the production of medicinal orchids for local communities. Critically endangered species should be placed on CITES Appendix I to provide extra protection to those species. DNA barcoding is an effective method for species identification and monitoring of illegal cross-border trade.

     

Factors affecting the use of medicinal plants by migrants from rural areas of Brazilian Northeast after moving to a metropolitan region in Southeast of Brazil

Background

Ethnopharmacological studies about migrants reveal a dynamic process of knowledge and use of medicinal plants. In this study, we sought to elucidate quantitative and qualitatively the main factors influencing the use of medicinal plants by migrants from rural areas to an urban region in Brazil with traces of remnant natural vegetation.

Methods

Seven Northeastern individuals who migrated to the Southeastern Region of Brazil (Bororé Peninsula, in the city of São Paulo) were selected to participate in semi-structured interviews regarding the use of medicinal plants throughout their lives, and indicated an inhabitant in their hometown that would be able to accompany the field collections in each area. Socioeconomic, educational, family structure, and use of Western medicine data were provided during interviews with the individuals from their hometowns. Plant samples cited by the interviewees were collected both at the current place of residence and in their hometowns.

Results

The participants cited 131 plants and 315 recipes, being the main indications related to the gastrointestinal system, respiratory problems, and pain and inflammatory processes. We observed that most plant uses were maintained after migration. Higher percentages of maintenances and incorporations in plant uses occurred to exotic species, while replacements happen mainly to native plants. The introduction of new species into the migrants’ therapeutics occurred mainly by observations of organoleptic similarities between the substituted plant and the incorporated species, conversations with neighbors, and contact with the television and print media. In addition, the public health system allowed the interviewees access to prophylactic drugs, leading to the discontinuation of certain recipes used in endemic diseases.

Conclusion

Migrants were exposed to information about new plants and their uses, new diseases, and socioeconomic and cultural differences that impacted their use of medicinal plants. Although migration to a more developed city facilitated access to public health and education, on the other hand, it made access to fresh medicinal plants difficult, causing some medicinal plants to be replaced or ceased to be used.

     

Pseudo current density maps of electrophysiological heart,nerve or brain function and their physical basis

Background

In recent years the visualization of biomagnetic measurement data by so-called pseudo current density maps or Hosaka-Cohen (HC) transformations became popular.

Methods

The physical basis of these intuitive maps is clarified by means of analytically solvable problems.

Results

Examples in magnetocardiography, magnetoencephalography and magnetoneurography demonstrate the usefulness of this method.

Conclusion

Hardware realizations of the HC-transformation and some similar transformations are discussed which could advantageously support cross-platform comparability of biomagnetic measurements.

     

Untargeted metabolomics suffers from incomplete raw data processing

Introduction

Untargeted metabolomics is a powerful tool for biological discoveries. To analyze the complex raw data, significant advances in computational approaches have been made, yet it is not clear how exhaustive and reliable the data analysis results are.

Objectives

Assessment of the quality of raw data processing in untargeted metabolomics.

Methods

Five published untargeted metabolomics studies, were reanalyzed.

Results

Omissions of at least 50 relevant compounds from the original results as well as examples of representative mistakes were reported for each study.

Conclusion

Incomplete raw data processing shows unexplored potential of current and legacy data.

     

Metabolomic profiling of maternal hair suggests rapid development of intrahepatic cholestasis of pregnancy

Introduction

Intrahepatic cholestasis of pregnancy (ICP) is a common maternal liver disease; development can result in devastating consequences, including sudden fetal death and stillbirth. Currently, recognition of ICP only occurs following onset of clinical symptoms.

Objective

Investigate the maternal hair metabolome for predictive biomarkers of ICP.

Methods

The maternal hair metabolome (gestational age of sampling between 17 and 41 weeks) of 38 Chinese women with ICP and 46 pregnant controls was analysed using gas chromatography–mass spectrometry.

Results

Of 105 metabolites detected in hair, none were significantly associated with ICP.

Conclusion

Hair samples represent accumulative environmental exposure over time. Samples collected at the onset of ICP did not reveal any metabolic shifts, suggesting rapid development of the disease.

     

Quantitative analysis of tetrahydrofolate metabolites from <Emphasis Type="Italic">clostridium autoethanogenum</Emphasis>

Introduction

Quantification of tetrahydrofolates (THFs), important metabolites in the Wood–Ljungdahl pathway (WLP) of acetogens, is challenging given their sensitivity to oxygen.

Objective

To develop a simple anaerobic protocol to enable reliable THFs quantification from bioreactors.

Methods

Anaerobic cultures were mixed with anaerobic acetonitrile for extraction. Targeted LC–MS/MS was used for quantification.

Results

Tetrahydrofolates can only be quantified if sampled anaerobically. THF levels showed a strong correlation to acetyl-CoA, the end product of the WLP.

Conclusion

Our method is useful for relative quantification of THFs across different growth conditions. Absolute quantification of THFs requires the use of labelled standards.

     

Investigation of species and environmental effects on rhubarb roots metabolome using <Superscript>1</Superscript>H NMR combined with high performance thin layer chromatography

Introduction

The pharmacological activities of medicinal plants are reported to be due to a wide range of metabolites, therein, the concentrations of which are greatly affected by many genetic and/or environmental factors. In this context, a metabolomics approach has been applied to reveal these relationships. The investigation of such complex networks that involve the correlation between multiple biotic and abiotic factors and the metabolome, requires the input of information acquired by more than one analytical platform. Thus, development of new metabolomics techniques or hyphenations is continuously needed.

Objectives

Feasibility of high performance thin-layer chromatography (HPTLC) were investigated as a supplementary tool for medicinal plants metabolomics supporting ¹H nuclear magnetic resonance (¹H NMR) spectroscopy.

Method

The overall metabolic difference of plant material collected from two species (Rheum palmatum and Rheum tanguticum) in different geographical locations and altitudes were analyzed by ¹H NMR- and HPTLC-based metabolic profiling. Both NMR and HPTLC data were submitted to multivariate data analysis including principal component analysis and orthogonal partial least square analysis.

Results

The NMR and HPTLC profiles showed that while chemical variations of rhubarb are in some degree affected by all the factors tested in this study, the most influential factor was altitude of growth. The metabolites responsible for altitude differentiation were chrysophanol, emodin and sennoside A, whereas aloe emodin, catechin, and rhein were the key species-specific markers.

Conclusion

These results demonstrated the potential of HTPLC as a supporting tool for metabolomics due to its high profiling capacity of targeted metabolic groups and preparative capability.

     

KniMet: a pipeline for the processing of chromatography–mass spectrometry metabolomics data

Introduction

Data processing is one of the biggest problems in metabolomics, given the high number of samples analyzed and the need of multiple software packages for each step of the processing workflow.

Objectives

Merge in the same platform the steps required for metabolomics data processing.

Methods

KniMet is a workflow for the processing of mass spectrometry-metabolomics data based on the KNIME Analytics platform.

Results

The approach includes key steps to follow in metabolomics data processing: feature filtering, missing value imputation, normalization, batch correction and annotation.

Conclusion

KniMet provides the user with a local, modular and customizable workflow for the processing of both GC–MS and LC–MS open profiling data.

     

A methodology for elucidating regulatory mechanisms leading to changes in lipid profiles

Introduction

It is difficult to elucidate the metabolic and regulatory factors causing lipidome perturbations.

Objectives

This work simplifies this process.

Methods

A method has been developed to query an online holistic lipid metabolic network (of 7923 metabolites) to extract the pathways that connect the input list of lipids.

Results

The output enables pathway visualisation and the querying of other databases to identify potential regulators. When used to a study a plasma lipidome dataset of polycystic ovary syndrome, 14 enzymes were identified, of which 3 are linked to ELAVL1—an mRNA stabiliser.

Conclusion

This method provides a simplified approach to identifying potential regulators causing lipid-profile perturbations.

     

Extraction of natural products from bark of <Emphasis Type="Italic">Betula pendula</Emphasis> using ionic liquids

Introduction

Aqueous–methanol mixtures have successfully been applied to extract a broad range of metabolites from plant tissue. However, a certain amount of material remains insoluble.

Objectives

To enlarge the metabolic compendium, two ionic liquids were selected to extract the methanol insoluble part of trunk from Betula pendula.

Methods

The extracted compounds were analyzed by LC/MS and GC/MS.

Results

The results show that 1-butyl-3-methylimidazolium acetate (IL-Ac) predominantly resulted in fatty acids, whereas 1-ethyl-3-methylimidazolium tosylate (IL-Tos) mostly yielded phenolic structures. Interestingly, bark yielded more ionic liquid soluble metabolites compared to interior wood.

Conclusion

From this one can conclude that the application of ionic liquids may expand the metabolic snapshot.

     

Variation of the microbiota and metabolome along the canine gastrointestinal tract

Introduction

The fecal microbiota are relevant to the health and disease of many species. The importance of the fecal metabolome has more recently been appreciated, but our knowledge of the microbiota and metabolome at other sites along the gastrointestinal tract remains deficient.

Objective

To analyze the gastrointestinal microbiota and metabolome of healthy domestic dogs at four anatomical sites.

Methods

Samples of the duodenal, ileal, colonic, and rectal contents were collected from six adult dogs after humane euthanasia for an unrelated study. The microbiota were characterized using Illumina sequencing of 16S rRNA genes. The metabolome was characterized by mass spectrometry-based methods.

Results

Prevalent phyla throughout the samples were Proteobacteria, Firmicutes, Fusobacteria, and Bacteroidetes, consistent with previous findings in dogs and other species. A total of 530 unique metabolites were detected; 199 of these were identified as previously named compounds, but 141 of them had at least one significantly different site-pair comparison. Noteworthy examples include relative concentrations of amino acids, which decreased from the small to large intestine; pyruvate, which peaked in the ileum; and several phenol-containing carboxylic acid compounds that increased in the large intestine.

Conclusion

The microbiota and metabolome vary significantly at different sites along the canine gastrointestinal tract.

     

Traditional medicine practitioners’ knowledge and views on treatment of pregnant women in three regions of Mali

Background

Despite the widespread use of medicinal plants in Mali, knowledge about how traditional practitioners (TPs) treat pregnant and lactating women is lacking.

Aim of the study

The aim of this study was to investigate how traditional practitioners in Mali treat common diseases and ailments during pregnancy.

Methods

Data was collected through structured interviews of traditional practitioners in one urban (Bamako) and two rural areas (Siby and Dioila) in Mali. The TPs were interviewed about how they treat common diseases and ailments during pregnancy. They were also asked to name harmful plants in pregnancy and plants that could affect breast milk production. In addition, we asked about nine specific medicinal plants commonly used in Mali; Opilia amentacea (syn. Opilia celtidifolia), Ximenia americana, Cola cordifolia, Combretum glutinosum, Parkia biglobosa, Trichilia emetica, Combretum micranthum, Lippia chevalieri and Vepris heterophylla.

Results

A total of 72 traditional practitioners (64% women, age: 34 to 90 years) were interviewed during an eight week period October 2011 to December 2011. They treated between 1 and 30 pregnant women with medicinal plants per months. We found a relatively high consensus for treatment of pregnant women with common diseases and ailments like nausea and dermatitis. The highest informer consensus was found for the treatment of malaria during pregnancy. TPs generally recommended pregnant women to avoid medicinal plants with bitter tastes like stem and root bark of Khaya senegalensis and Opilia amentacea (syn. Opilia celtidifolia). TPs distinguished between oral (potentially unsafe) and dermal use (safe) of Opilia amentacea (syn. Opilia celtidifolia). Cola cordifolia was used to facilitate labor.

Conclusion

Experience and knowledge about treatment of pregnant women with medicinal plants was broad among the traditional practitioners in the three investigated regions in Mali. Collaborating with traditional practitioners on the safe use of medicinal plants in pregnancy may promote safer pregnancies and better health for mothers and their unborn infants in Mali.