Introduction: Folkloric claims have surrounded essential oils, including their enhancement of learning and memory through inhalational exposure. Few studies in humans have shown a benefit in cognition, albeit incremental. However, this benefit may not be entirely attributable to the essential oil aroma but may be confounded by psychological associations. We investigated rosemary, peppermint, lemon, and coffee aromas in a learning and memory model of Drosophila melanogaster to eliminate this confounder. Methods: We screened for concentrations of the four treatments that are non-stimulatory for altered locomotory behavior in the flies. At these concentrations, we determined if they were chemoneutral (i.e., neither chemoattractant nor chemorepellent) to the flies. Learning and memory of the flies exposed to these aromas were determined using an Aversive Phototaxis Suppression (APS) assay. Results: The aromas of rosemary, peppermint, and lemon that did not elicit altered mobility in the flies were from dilute essential oil solutions that ranged from 0.2 to 0.5% v/v; whereas for the aroma in coffee, it was at a higher concentration of 7.5% m/v. At these concentrations, the aromas used were found to be chemoneutral towards the flies. We observed no improvement in both learning and memory in the four aromas tested. While a significant reduction (p < 0.05) in learning was observed when flies were treated with the aromas of rosemary, peppermint, and coffee, a significant reduction (p < 0.05) in memory was only observed in the peppermint aroma treatment. Conclusion This study demonstrated that in the absence of psychological association, the four aromas do not enhance learning and memory
Drosophila melanogaster ; Learning ; Memory ; Rosmarinus ; Mentha piperita ; Citrus ; Coffea

Carotenoids are secondary metabolite responsible for colored pigments in plants and microbes (Li et al., 2022). They are a class of C₄₀ tetraterpenoids consisting of eight isoprenoid units, and can be classified into carotenes and xanthophylls on the basis of their functional groups (Saini et al., 2015). Carotenes can be linear (phytoene, phytofluene, and ζ‍-carotene) or branched (β‍-carotene and α‍-carotene). Xanthophylls comprise β,β‍-xanthophylls (β‍-cryptoxanthin, zeaxanthin, violaxanthins, and neoxanthin) and β,ε‍-xanthophylls (α-cryptoxanthin, α-carotene, and lutein). Citrus fruits are complex sources of carotenoids, which are the principal pigments responsible for the typical orange color of most types (Chen, 2020). The difference in total carotenoid content and the diversity of carotenoid isomer proportion also accounts for other colors of citrus fruits, such as yellow, red, and pink (Chen, 2020).
Citrus/metabolism* ; Carotenoids ; Xanthophylls ; Lutein/metabolism* ; Zeaxanthins/metabolism* ; Fruit

The present study explored the consistency of the content proportions of active components of Aurantii Fructus and analyzed the influencing factors based on three-dimensional multi-component analysis. A total of 839 Aurantii Fructus samples in 65 research articles were analyzed using the three-dimensional multi-component analysis mode. The content data of flavonoid components(naringin, hesperidin, neohesperidin, narirutin, and nobiletin), coumarin components(meranzin and gluconolactone), and alkaloid(synephrine) in 386 samples which met the criteria of 2020 edition of the Chinese Pharmacopoeia were extracted and adjusted to percentages, and the content ratios between components were calculated. The influencing factors of Aurantii Fructus quality were analyzed. The results showed content ratios of components as follows: neohesperidin∶naringin in the range of 0.4-1.2; narirutin∶naringin in the range of 0.02-0.16; hesperidin∶naringin in the range of 0.01-0.3; nobiletin∶naringin in the range of 0.000 588 3-0.069 68; synephrine∶naringin in the range of 0.02-0.042; gluconolactone∶naringin in the range of 0.001-0.01; meranzin∶naringin in the range of 0.000 4-0.035. The quality of Aurantii Fructus was closely related to the origin, variety, harvesting time, and processing method of medicinal materials. Harvesting time had a greater impact on the quality of Aurantii Fructus, and the origin and variety had a certain impact on the quality of Aurantii Fructus. The findings of this study indicated that the ratios between flavonoid components, flavonoids and coumarin components, and flavonoids and alkaloids fluctuated. The production base should optimize the varieties, harvesting period, and processing methods of Aurantii Fructus to provide a scientific basis for the production of high-quality Aurantii Fructus.
Citrus ; Flavonoids/analysis* ; Drugs, Chinese Herbal ; Fruit/chemistry* ; Coumarins/analysis* ; Chromatography, High Pressure Liquid/methods*

The present study analyzed the effect of Citri Reticulatae Pericarpium on endogenous metabolites in spleen deficiency and phlegm dampness syndrome by metabolomics, and explored the underlying mechanism of Citri Reticulatae Pericarpium in the treatment of spleen deficiency and phlegm dampness syndrome.The model of spleen deficiency and phlegm dampness syndrome was induced in rats by the multi-factor modeling method.The intervention effects of Citri Reticulatae Pericarpium on rats with spleen deficiency and phlegm dampness syndrome were preliminarily evaluated by observing the pathological changes of rat liver tissues and measuring the plasma content of pathological and biochemical indexes such as triglyceride(TG), total cholesterol(TC), low-density lipoprotein cholesterol(LDL-C), and high-density lipoprotein cholesterol(HDL-C).Immunohistochemistry was used to detect the expression of AQP2 in the kidney, AQP3 in the colon, and AQP5 in the submandibular gland, and the effect of Citri Reticulatae Pericarpium on aquaporin expression in rats with spleen deficiency and phlegm dampness syndrome was evaluated.Furthermore, UHPLC-ESI-MS/MS was used to analyze the metabolic profiles of rat plasma samples.Multiple methods, such as principal component analysis(PCA) and orthogonal partial least squares discriminant analysis(OPLS-DA) were used for pattern recognition.Differential metabolites were screened out by t-test and variable importance in projection(VIP), followed by pathway analysis based on MetaboAnalyst 5.0.As revealed by experimental results, Citri Reticulatae Pericarpium could improve the pathological changes of liver tissues, increase the levels of HDL-C in the plasma, reduce the levels of TC, TG, and LDL-C, and enhance the expression of AQP2 in the kidney, AQP3 in the colon, and AQP5 in the submandibular gland of rats with spleen deficiency and phlegm dampness syndrome.In addition, 87 differential metabolites of spleen deficiency and phlegm dampness syndrome were screened out by UHPLC-ESI-MS/MS(the levels of 39 metabolites increased significantly and the levels of 48 metabolites decreased significantly), with the representatives of glycine, L-isoleucine, N-acetyl-L-tyrosine, xanthine, hypoxanthine, and trigonelline.The differential metabolites were mainly enriched in the pathways of steroid hormone biosynthesis, linoleic acid metabolism, and purine metabolism.This study distinguished and revealed the characteristic metabolic pattern of spleen deficiency and phlegm dampness syndrome by metabolomics.The preliminary construction of the OPLS-DA model provides an objective basis for the differentiation of spleen deficiency and phlegm dampness syndrome in traditional Chinese medi-cine(TCM), as well as ideas and methods for exploring the biological basis of TCM syndrome from the molecular level and the overall level.
Animals ; Aquaporin 2 ; Cholesterol, LDL ; Citrus/chemistry* ; Drugs, Chinese Herbal ; Metabolomics ; Rats ; Spleen ; Tandem Mass Spectrometry

The present study established the spectrum-effect relationship model of flavonoids in Citri Reticulatae Pericarpium(CRP) from 15 batches of Liujunzi Decoction and statistically analyzed the correlation between chemical peaks and efficacy to identify the main effective components. HPLC fingerprints of flavonoids in CRP from 15 batches of Liujunzi Decoction were established. HPLC analysis was carried out on the Venusil XBP C_(18)(L) column(4.6 mm×250 mm, 5 μm) at 30 ℃ with acetonitrile-water(containing 0.1% formic acid) as mobile phase for gradient elution, a flow rate of 1.0 mL·min~(-1), and detection wavelength of 300 nm to obtain chemical fingerprints. Additionally, the effects of flavonoids from CRP in 15 batches of Liujunzi Decoction on the content of GAS, MTL, and VIP, TFF3 mRNA expression, and percentage of CD3~+ T-cells of model rats with spleen deficiency were determined. The spectrum-effect relationship model was established by gray correlation analysis. The results showed that the main characteristic peaks with great contribution to the regulation of gastrointestinal tract were peak 16(vicenin-2), peak 63(sinensetin), peak 64(isosinensetin), peak 65(nobiletin), peak 67(3,5,6,7,8,3',4'-heptemthoxyflavone), peak 68(tangeretin), and peak 69(5-desmethylnobiletin). Therefore, there was a linear correlation between flavonoids from CRP in Liujunzi Decoction and the efficacy, and the medicinal effect was achieved by multi-component action. This study is expected to provide a new idea for exploring the material basis of the effect, i.e., regulating qi prior to replenishing qi, of CRP in Liujunzi Decoction.
Animals ; Citrus/chemistry* ; Drugs, Chinese Herbal ; Flavonoids/pharmacology* ; Hormones ; RNA, Messenger/genetics* ; Rats ; Spleen

As a local variety of medicinal material, Citri Trifoliatae Fructus is widely used in many places, whereas its harvest time remains unclear. Therefore, studying its harvest time can make more reasonable use of this medicinal material. In this study, we determined the flavonoids content and compared the color of Citri Trifoliatae Fructus harvested in different time, aiming to guide the harvest of this medicinal material. The fresh fruits of Citrus trifoliata were collected from Xinxiang city, Henan province, graded according to the diameter range, and then dried. The contents of isonaringin, naringen, and poncirin in Citri Trifoliatae Fructus were determined by HPLC, and the color values of the samples were detected by electronic eye. The correlation analysis of the obtained data was carried out to explore the relationships of color and diameter with quality. The results showed that the contents of isonaringin, naringen, and poncirin varied significantly in different harvest time, within the ranges of 0.21-1.20, 2.21-11.59, and 3.73-23.16 mg·g~(-1), respectively. With the delay of harvest time, Citri Trifoliatae Fructus showed the color changing from green to yellow, gradually increased diameter, and gradually decreased contents of isonaringin, naringen, and poncirin. The contents of isonaringin, naringen, and poncirin were negatively correlated with the degree of red and green(a~*) and positively correlated with the degree of yellow and blue(b~*). The contents of naringen and poncirin had significantly negative correlations with the diameter. This study indicates that the quality of Citri Trifoliatae Fructus can be judged by its diameter and skin color, which provides a theoretical basis for the rational harvest of this medicinal material.
Chromatography, High Pressure Liquid ; Citrus ; Drugs, Chinese Herbal ; Electronics ; Fruit ; Technology

Aurantii Fructus is a commonly used qi-regulating medicinal herb in China. Both traditional Chinese medicine theory and modern experimental research demonstrate that Aurantii Fructus has dryness effect, the material basis of which remains unclear. In recent years, spectrum-effect relationship has been widely employed in the study of active ingredients in Chinese medicinal herbs, the research ideas and methods of which have been constantly improved. Based on the idea of spectrum-effect study, the ultra-high perfor-mance liquid chromatography-quadrupole-time of flight mass spectrometry(UHPLC-Q-TOF-MS) fingerprints of different fractions of Aurantii Fructus extract were established for the identification of total components. Then, the dryness effects of the fractions on normal mice and gastrointestinal motility disorder(GMD) rats were systematically compared. Finally, principal component analysis(PCA), Pearson bivariate correlation analysis and orthogonal partial least squares analysis(OPLS) were integrated to identify the dryness components of Aurantii Fructusextract. The results showed that narirutin, naringin, naringenin, poncirin, oxypeucedanin, and eriodictyol-7-O-glucoside had significant correlations with and contributed to the expression of AQP2 in kidney, AQP3 in colon, and AQP5 in submandibular gland, which were the main dryness components in Aurantii Fructus.
Animals ; Aquaporin 2 ; Chromatography, High Pressure Liquid ; Citrus ; Drugs, Chinese Herbal ; Gastrointestinal Motility ; Medicine, Chinese Traditional ; Mice ; Rats

The ripe dried fruit of citron(Citrus medica) is one of the important sources of Chinese herb Citri Fructus. At the same time, it is also grown for edible and ornamental uses. There are many species and abundant genetic variation. To clarify the intraspecific variation and resource distribution of citron, this study investigated the variation in 11 citron fruits, basically covering the main species in China, including Xiaoguo citron(C. medica var. ethrog), Goucheng(C. medica var. yunnanensis), Muli citron(C.medica var. muliensis), Dehong citron(C.medica×Citrus spp.), Fuzhou citron(C.medica×C.grandis?), Mawu(C.medica×C.grandis?), Cangyuan citron, Binchuan citron, Sweet citron, Big citron, and Small citron. The natural communities of citron were proved to be mainly distributed in the southwestern and western Yunnan and southeastern Tibet of China, with Yunnan, Sichuan, Guangxi, Chongqing, Hubei, and Zhejiang identified as the main production areas. Citron has also been widely grown in India, the Mediterranean region, and the Caribbean coast countries. The field investigation revealed the large-scale intraspecific variation of citron fruits. Most of the fruits are oval-like or sphere-like in shape. The fruits are green when raw and yellow when ripe, with oil cell dots on the skin, stripe-likes running from top to bottom, and bulge at the top. Usually, in the smaller citron fruits, the pulp and juice vesicles are better developed and the central columella is tighter. By contrast, the juice vesicles and central columella in larger fruits became more vacant, with carpels visible, and the apex segregation and development of the carpels is one of the reasons for variation. These variations should be given top priority in the future variety selection and breeding, and the quality differences of different citron species and their mechanisms should be further studied. In particular, variety selection and classification management according to their medicinal or edible purposes will provide scientific and technological supports for the orderly, safe, and effective production of citron products consumed as food and medicine.
China ; Citrus ; Fruit ; Taste ; Tibet

This study was designed to predict the Q-markers of Citri Reticulatae Pericarpium volatile oil and conduct quantitative analysis by GC-MS. The common components of Citri Reticulatae Pericarpium volatile oil were detected by GC-MS. The network pharmacology approaches were utilized for constructing the component-target network and protein-protein interaction(PPI) network, followed by the GO and KEGG pathway enrichment analysis to clarify the pharmacological effects of common components. Molecular docking was conducted to observe the biological activities of common components, thus identifying the Q-markers of Citri Reticulatae Pericarpium volatile oil. The obtained Q-markers were subjected to quantitative analysis by GC-MS. The GC-MS analysis of 19 batches of Citri Reticulatae Pericarpium volatile oil revealed three common components, namely, D-limonene, γ-terpinene, and myrcene. The common components were analyzed based on network pharmacology, and the results showed that Citri Reticulatae Pericarpium volatile oil mainly acted on the core targets GABRA1, GABRA6, GABRA5, GABRA3, and GABRA2 through D-limonene and γ-terpinene, with five important pathways such as nicotine addiction and GABAergic synapse involved. The core targets were mainly distributed in olfactory region, cerebral cortex, cerebellum, basal ganglia, hippocampus, and amygdala to exert the pharmacological effects. As revealed by molecular docking, D-limonene and γ-terpinene exhibited good biological activities, so they were identified as the Q-markers of Citri Reticulatae Pericarpium volatile oil. The results of quantitative analysis showed that the volume fraction of D-limonene was within the range of 0.77-1.03 μL·mL~(-1), and that of γ-terpinene within the range of 0.04-0.13 μL·mL~(-1). The prediction of D-limonene and γ-terpinene as the Q-markers of Citri Reticulatae Pericarpium volatile oil has laid an experimental foundation for the establishment of the quality evaluation standard for Citri Reticulatae Pericarpium volatile oil.
Citrus ; Drugs, Chinese Herbal/pharmacology* ; Gas Chromatography-Mass Spectrometry ; Molecular Docking Simulation ; Network Pharmacology ; Oils, Volatile/pharmacology*

Twenty batches of Aurantii Fructus Immaturus(AFI) were collected, with their peel and pulp taken as research objects. Ultra-high performance liquid chromatography(UPLC) fingerprints of peel and pulp of AFI were established with 17 common peaks in peel and 10 in pulp. Six kinds of flavonoids were identified, i.e., narirutin, naringin, rhoifolin, hesperidin, neohesperidin and nobiletin. The Similarity Evaluation System for Chromatographic Fingerprint of Traditional Chinese Medicine was employed for similarity analysis, which showed that the chromatographic peaks of peel and pulp were basically similar to their respective reference fingerprints, with all similarities greater than 0.90. The similarity between peel and pulp of the same batch of AFI ranged from 0.850 to 0.983. Cluster analysis(CA), principal component analysis(PCA), and orthogonal partial least squares discriminant analysis(OPLS-DA) were conducted on the common peaks of peel and pulp of AFI with SPSS 17.0 and SIMCA 14.1. Combined with the reference fingerprints, these analyses revealed 12 differential components regarding peel and pulp. Further, the content of the 6 flavonoids and synephrine was determined. The proposed method integrating UPLC fingerprint and multicomponent quantitative analysis is applicable to the quality evaluation of AFI. The results provide a certain basis for the scientific connotation about the appearance characteristic of AFI.
Chromatography, High Pressure Liquid ; Citrus ; Drugs, Chinese Herbal ; Synephrine