A Scientometric Study to a Critical Review on Promising Anticancer and Neuroprotective Compounds: Citrus Flavonoids

Flavonoids derived from citrus plants are favored by phytomedicinal researchers due to their wide range of biological activities, and relevant studies have been sustained for 67 years (since the first paper published in 1955). In terms of a scientometric and critical review, the scientometrics of related papers, chemical structures, and pharmacological action of citrus flavonoids were comprehensively summarized. The modern pharmacological effects of citrus flavonoids are primarily focused on their anticancer activities (such as breast cancer, gastric cancer, lung cancer, and liver cancer), neuroprotective effects (such as anti-Alzheimer’s disease, Parkinson’s disease), and metabolic diseases. Furthermore, the therapeutic mechanism of cancers (including inducing apoptosis, inhibiting cell proliferation, and inhibiting cancer metastasis), neuroprotective effects (including antioxidant and anti-inflammatory), and metabolic diseases (such as non-alcoholic fatty liver disease, type 2 diabetes mellitus) were summarized and discussed. We anticipate that this review could provide an essential reference for anti-cancer and neuroprotective research of citrus flavonoids and provide researchers with a comprehensive understanding of citrus flavonoids.


Introduction
Citrus genome studies have shown that citrus plants originated in the Himalayas about 6 to 8 million years ago, and the majority of species are descendants of wild broad-hued oranges (C. reticulata), grapefruit (C. maxima), and citron (C. medica) [1]. As time went on, people found citrus fruits were not only beautiful and edible, but also fairly tasty. In a bid to improve the taste of citrus fruit, people have developed several citrus cultivation varieties, such as C. hystrix, C. japonica, C. mitis, C. aurantifolia, C. paradisi, C. junos, C. limetta, C. maximas, C. aurantium, C. limon, and C. sinensis. In recent years, citrus has become one of the most productive fruits in the world for economic cultivation [2]. Since ancient times in China, Egypt, and India, citrus fruit was not only used as a tasty fruit but also as a medicine. The dried peel of Citrus reticulata Blanco was used as an ingredient in tea and in Chinese patent medicines, which was beneficial to promote health by regulating "qi". Modern botanists are also interested in the medicinal value of citrus. Botanical medicine researchers have found that citrus fruits are rich in a variety of beneficial components, such as fibers [3], phenolic acids [4], and flavonoids [5]. Notably, citrus flavonoids, recognized as a class of substances with important nutritional value, are comprehensively investigated. Citrus flavonoids are usually classified according to their chemical structures, such as flavanone aglycones, flavone aglycones, flavanonols, flavanone-O-glucoside, polymethoxyflavonoids, other flavonoids, flavone-O-glucoside, and flavone-C-glucoside [6]. Studies have shown that citrus flavonoids possess a variety of pharmacological properties including antioxidant and and anti-inflammatory [7], among others. In view of the complex chemical composition and diverse pharmacological activities of citrus flavonoids, we believe that it is significant to summarize Citrus flavonoids by means of scientometric analysis and a critical review. comprehensively investigated. Citrus flavonoids are usually classified according to their chemical structures, such as flavanone aglycones, flavone aglycones, flavanonols, flavanone-O-glucoside, polymethoxyflavonoids, other flavonoids, flavone-O-glucoside, and flavone-C-glucoside [6]. Studies have shown that citrus flavonoids possess a variety of pharmacological properties including antioxidant and and anti-inflammatory [7], among others. In view of the complex chemical composition and diverse pharmacological activities of citrus flavonoids, we believe that it is significant to summarize Citrus flavonoids by means of scientometric analysis and a critical review.
The scientometric methods were first applied to obtain a holistic and comprehensive view based on the published studies on citrus flavonoids, which is an application of mathematical and statistical methods to perform retrospective reviews, calculate correlations in publication data, elucidate current research progress, and predict research directions [8]. Scientometric methods have played an important role in bone disease research [9], hotspots of exercise for intervening diabetes [10], COVID-19 research [11], and exosome studies [12]. The numerous published articles were summarized through the scientometric methods for providing rich reference information for researchers in need based on keywords, highlights, and important research-related information.
The research protocol of this article was shown in Figure 1. We summarized the development of scientific research on citrus flavonoids, the chemical structure, and distribution of citrus flavonoids in plants, and summarized the main pharmacological actions including antioxidation, inhibition of cancer progression, and neuroprotection. The mechanisms of these pharmacological actions were also discussed. The purpose of this article was to sort out the research history of citrus flavonoids, the plant sources of citrus flavonoid compounds, and the main pharmacological mechanisms of action of citrus flavonoids in order to provide researchers with a comprehensive understanding of citrus flavonoids.

Literature Search and Data Download
Citrus flavonoid was searched as a topic in the Web of Science core collection database as of 19 July 2022. Full records and cited references considered as raw data were downloaded from the database, and the file format was plain text.

Scientometric Analysis and Visualization
CiteSpace was initially used for bibliometric analysis; the factors include country, institute, keyword, category, reference, and cited journal, with parameter settings of time slicing (1991-2022), node type, and selection criteria (top 50 levels of the most cited or occurring items). In addition, VOSviewer was applied to optimize and provide an aesthetic map. Impact factor (IF) and Hirsh Index (H-index) were fully considered for a comprehensive and scientometric analysis.  (g) Institution citation co-occurrence view. Node size represents the total number of citations.

Journal Analysis
Twenty-five journals published the majority of articles related to citrus flavonoids. As shown in Table 1, the Journal of Agricultural and Food Chemistry had the highest publication number, followed by Food Chemistry, Molecules, Food and Function, and the Journal of Functional Foods. Interestingly, as shown in Table 2, five journals possessed fewer publications but higher citations (cited number > 600), which were the Journal of Biological Chemistry, Life Sciences, the Journal of Nutrition, PLoS ONE, and Phytochemistry.   Table 3. The most cited article reported that vitamin C and phenols were the main components of antioxidant capacity in citrus juice [13]. The second most cited article reported the extraction of polyphenols, especially flavanones from orange (Citrus sinensis L.) peel by using ethanol as a food-grade solvent [14]. Figure 3b presents a co-citation reference review. The top 15 cited references on citrus flavonoids are shown in Table 4.

Keywords Analysis
The keyword co-occurrence network is shown in Figure 3c. The weight of the circle size denotes the frequency of occurrence of keywords. The keyword population distribution is shown in Figure 3d

Chemical Structures and Sources of Citrus Flavonoids
The structures of citrus flavonoids are summarized in    types except flavanone aglycones; C. reticulata includes all structural types and co the most flavone glycosides among this type of compound; C. limetta contains on vanone aglycones and flavone glycosides. The abovementioned results indicate that plants tend to enrich flavonoids in fruits rather than roots. Naringin, narirutin, hespe and rutin are the most widely distributed compounds found in citrus plants.

Citrus Flavonoids and Cancers
According to the search results, citrus flavonoids have an obvious inhibitory effect against various cancers [18], the most studied of which is breast cancer; other cancers include rectal cancer, gastric cancer, liver cancer, lung cancer, prostate cancer, uterine cancer, ovarian cancer, epidermal cancer. Citrus flavonoids play a role in cancer therapy by inhibiting cancer cell proliferation [40], migration, angiogenesis, and inducing apoptosis [41]. We summarized the molecular mechanism of citrus flavonoids in cancer therapy and provided a reference for cancer therapy research.
It should be noted here that CYP3A4, as the most important oxidative enzyme, plays a metabolic role for most drugs [42], and the role of CYP3A4 on drug metabolism in cancer treatment has attracted more attention [43]. While grapefruit inhibits the expression of CYP3A4 [44], some studies have shown that Fructus aurantia and tangeretin induce CYP3A4 [45,46]. It is clear that some citrus flavonoids have a regulatory effect on CYP3A4.
It is suggested that we should be cautious when consuming products derived from citrus to avoid reducing the efficacy of the drug or enhancing the adverse effects.

Breast Cancer
Compounds and molecular mechanisms of citrus flavonoids against breast cancer are shown in Figure 6.  Nobiletin, a natural flavonoid isolated from citrus peel, has anti-angiogenic effects [47]. Nobiletin was shown to inhibit MCF7 breast cancer cells by inducing its metabolism by up-regulating cytochrome P450 family 1 subfamily A member 1 (CYP1A1) and cytochrome P450 family 1 subfamily B member 1 (CYP1B1) [48]. Furthermore, nobiletin was shown to induce apoptotic cell death by reducing B-cell leukemia/lymphoma 2 xL (Bcl-xL) expression without affecting Bcl-2-associated x protein (Bax) levels and inhibit the activities of protein kinase B (AKT) and downstream mammalian target of rapamycin (mTOR) [49]. These targets are located in the apoptosis pathway, suggesting that the treatment of breast cancer by tangerine is mainly through inducing the apoptosis of cancer cells.
Naringenin was shown to inhibit the growth of metastases after surgery by modulating host immunity [50]. Naringin can inhibit cancer cell reproduction by inhibiting vascular endothelial factor release [51] and regulating the β-catenin pathway [52]. Hesperetin can induce apoptosis in breast cancer cells by triggering the accumulation of reactive oxygen species (ROS), activating the apoptosis signal-regulating kinase 1 (ASK1)/c-jun n-terminal kinase (JNK) pathway, and activating targets of caspase-9 and caspase-3. Hesperetin could increase the Bax: B-cell lymphoma-2 (Bcl-2) ratio in the intracellular environment [53].
The results indicate that hesperetin can inhibit cancer cells by inducing apoptosis. Polymethoxyflavonoids was shown to induce apoptosis in breast cancer cells [54] by activating a Ca (2 + )-dependent pro-apoptotic protease [55]. Retusin and Ayanin are potent inhibitors of breast cancer resistance protein (BCRP), showing only slightly lower potency than Ko143 [56].
According to the abovementioned results, it is known that the therapeutic mechanism of citrus flavonoids affecting breast cancer mainly depends on inducing apoptosis. In addition, citrus flavonoids inhibit cell proliferation pathways and slow breast cancer cell reproduction. Citrus flavonoids can also inhibit the metastasis of cancer cells. Overall, citrus flavonoids treat breast cancer in a variety of ways. In conclusion, the therapeutic effect of citrus flavonoids on breast cancer is clear. However, more comprehensive and in-depth studies are needed to make citrus flavonoids a suitable drug for the treatment of breast cancer.

Colorectal Cancer
Nobiletin showed a strong inhibitory effect on the growth of colon cancer cells [57] by inhibiting matrix metallopeptidase 7(MMP-7) (Figure 7) gene expression [58]. Nobiletin inhibited cancer invasion and metastasis by increasing tissue the tissue inhibition of metalloproteinase-1 (TIMP-1) production [59]. In addition, it was found that nobiletin could down-regulate leptin levels [60]. High levels of leptin in mice are thought to be a key factor in promoting colorectal cancer. Tangerine was metabolized in the intestine to 3 -desmethylnorchol, 4 -desmethylnorchol and 3 ,4 -didemethylnorchol. These metabolites were considered to be key compounds for the treatment of intestinal cancer [61].
Naringenin and hesperetin play a critical role in inhibiting the formation of abnormal crypt foci [64] and reducing the activity of bacterial enzymes in colon cancer [65].
Nobiletin is an important component for the treatment of colon cancer. The mechanism of citrus flavonoids on colon cancer can induce apoptosis of cancer cells, inhibit the growth of cancer cells, and regulate intestinal enzymes. It is believed that the most important mechanism is still the induction of apoptosis. Additional research is needed to clarify the mechanism of citrus flavonoids in the treatment of colon cancer.
Naringenin and hesperetin play a critical role in inhibiting the formation of abnorma crypt foci [64] and reducing the activity of bacterial enzymes in colon cancer [65].
Nobiletin is an important component for the treatment of colon cancer. The mecha nism of citrus flavonoids on colon cancer can induce apoptosis of cancer cells, inhibit the growth of cancer cells, and regulate intestinal enzymes. It is believed that the most im portant mechanism is still the induction of apoptosis. Additional research is needed to clarify the mechanism of citrus flavonoids in the treatment of colon cancer.

Gastric Cancer
Nobiletin could inhibit proliferation and induce apoptosis of gastric cancer cells [66] Nobiletin could also slow the progression of cancer by extending the cell growth cycle [67]. The preliminary effect of naringenin in treating gastric cancer has been demonstrated

Gastric Cancer
Nobiletin could inhibit proliferation and induce apoptosis of gastric cancer cells [66]. Nobiletin could also slow the progression of cancer by extending the cell growth cycle [67]. The preliminary effect of naringenin in treating gastric cancer has been demonstrated [68] by inhibiting cell proliferation, migration, and invasion [69], and by causing ASK1-induced apoptosis mediated by ROS (Figure 7) [70].
5-Demethyltangeretin inhibited human non-small-cell lung cancer cell growth by inducing G2/M cell cycle arrest and apoptosis [74].
Flavanones and 2 -OH flavanones could inhibit the growth of A549 and Lewis lung cancer cells in vivo [75].
Hesperidin produced in vitro inhibitory effects NSCLC cells by modulating immune response-related pathways that affect apoptosis [76]. These results provide scientific support for the use of flavonoids extracted and isolated from citrus plants for the treatment of human lung cancer.

Liver Cancer
Naringenin induced cell cycle arrest and inhibited the growth of human hepatocellular carcinoma cells [77].
Hesperidin induced apoptosis of human hepatocellular carcinoma (HepG2) cells through mitochondrial and death receptor pathways [78].
Naringenin could also promote deoxyribonucleic acid (DNA) repair and prevent carcinogenesis caused by oxidative damage [80].

Cervical Cancer
Hesperetin exhibited potential anticancer activity in vitro against human cervical cancer cell lines by reducing cell viability and inducing apoptosis [81].

Epidermal Carcinoma
Studies have shown that citrus flavonoids have anti-proliferative effects in inhibiting human squamous cell carcinoma in vitro [85], and the relevant studies proved that naringenin exerts anti-proliferative effects by inducing ROS generation and cell cycle arrest [86].

Neuroprotective Effects of Citrus Flavonoids
Studies have shown that fruits rich in flavonoids could protect the nervous system [87]. Citrus flavonoids inhibited Alzheimer's disease by reducing Presenilin 1 (PS1) phosphorylation-dependent amyloid production [88], and hesperidin, hesperetin, and neohesperidin exhibited neuroprotective effects [89]. Eriodictyol induced nuclear translocation of nuclear factor erythroid-2 related factor 2 (Nrf2), enhanced heme oxygenase 1 (HO-1) and NAD(P)H quinone dehydrogenase 1 (NQO-1) expression, and increased intracellular glutathione levels against oxidative stress-induced cell death [90]. We summarized studies on the neuroprotective effects of citrus flavonoid compounds to elucidate their mechanisms of action, as shown in Figure 8.
Nobiletin could stimulate protein kinase A (PKA)-mediated phosphorylation of glutamate receptor 1 (GluR1) receptors in the hippocampus to upregulate synaptic propagation through postsynaptic AMPA receptors [91]. It could also rescue cholinergic neurodegeneration and improve memory impairment in olfactory bulbectomy (OBX) mice by reducing the acetyl cholinesterase (AChE) staining and choline acetyltransferase (ChAT) expression density in the hippocampus [92]. Furthermore, nobiletin improved memory impairment and amyloid beta disease in a transgenic mouse model of Alzheimer's disease [93]. Additionally, triple transgenic (3xTg)-AD mice were orally administrated with 30 mg/kg nobiletin for 3 months, and the results showed that nobiletin reversed the impairment of short-term memory and recognition memory by reducing soluble amyloid beta 1-40 (Aβ1-40) and ROS levels in the mouse brains [94]. Oral administration of nobiletin reduced tau phosphorylation in the hippocampus of senescence-accelerated P8 (SAMP8) mice [95]. Nobiletin rescued 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced Parkinson's in a mouse model, reducing the dopamine level in the striatum and hippocampal CA1 region to prevent motor and cognitive dysfunction [96]. Nobiletin reversed learning disabilities associated with the n-methyl-d-aspartate receptor by enhancing cAMP/PKA/extracellular-regulated protein kinase (ERK) signaling in hippocampal neurons and PC12D cells [97]. These studies suggest that nobiletin has great potential in the study of neuropathic diseases. Studies have shown that fruits rich in flavonoids could protect the nervous system [87]. Citrus flavonoids inhibited Alzheimerʹs disease by reducing Presenilin 1 (PS1) phosphorylation-dependent amyloid production [88], and hesperidin, hesperetin, and neohesperidin exhibited neuroprotective effects [89]. Eriodictyol induced nuclear translocation of nuclear factor erythroid-2 related factor 2 (Nrf2), enhanced heme oxygenase 1 (HO-1) and NAD(P)H quinone dehydrogenase 1 (NQO-1) expression, and increased intracellular glutathione levels against oxidative stress-induced cell death [90]. We summarized studies on the neuroprotective effects of citrus flavonoid compounds to elucidate their mechanisms of action, as shown in Figure 8. Naringin protected nigrostriatal nigrothymic dopaminergic (DA) projections from 6-hydroxydopamine (6-OHDA)-induced neurotoxicity [98]. Naringin conferred an important capacity for DA neurons to produce the glial cell-derived neurotrophic factor (GDNF) [99]. In addition, naringin could improve cognitive performance and attenuate oxidative damage [100].
Naringenin exerted anti-inflammatory effects due to its interaction with the p38 signaling cascade and signal transducer and activator of the transcription 1 (STAT-1) transcription factor [101]. Naringenin can also inhibit the release of idiopathic oxide (NO) and pro-inflammatory cytokines in microglia [102].
Hesperidin protected against cognitive impairment by inhibiting the overexpression of inflammatory markers such as NF-κB, nitric oxide synthase (NOS), and cyclooxygenase-2 (COX-2) [103]. Furthermore, hesperidin significantly restored the deficits in non-cognitive nesting abilities and social interaction by attenuating amyloid beta deposition in the brain [104].
Citrus flavonoids protect the nervous system by fighting against inflammation and protecting the function of nerve cells. It is promising to develop citrus flavonoids as an adjuvant treatment for neurodegenerative diseases such as Alzheimer's disease, Parkinson's disease, etc. It is believed that the protective effects of citrus flavonoids on the nervous system is worthy of further and in-depth research.

Citrus Flavonoid and Metabolic Disease
Oral administration of bergamot extract (150 mg containing 16% neohesperidin, 47% neohesperidin, and 37% naringin) for 6 months reduced moderate hypercholesterolemia, low-density lipoprotein, and blood lipids in patients with atherosclerosis [110]. Neohesperidin activated the AMPK pathway for hypoglycemic and exhibited lipid-lowering effects [111]. It was obvious that citrus flavonoids were beneficial in the treatment of metabolic diseases. We summarized the regulatory mechanisms of other citrus flavonoids on metabolic diseases.
Nobiletin attenuated dyslipidemia by preventing hepatic triglyceride (TG) accumulation, reducing very low-density lipoprotein (VLDL) and TG secretion [112], while increasing hepatic and peripheral insulin sensitivity and glucose tolerance, and significantly attenuating atherosclerosis in the aortic sinus hardening [113]. In addition, nobiletin could enhance the circadian rhythm to combat metabolic diseases by intervening in the circadian rhythm network [114].
Polymethoxyflavonoids are a novel flavonoid with cholesterol and triacylglycerollowering potential, and elevated levels of polymethoxyflavonoid metabolites in the liver may directly lead to its hypolipidemic effect in vivo [129]. Hesperidin stimulated nitric oxide production in endothelial cells while improving endothelial function and reducing inflammatory markers in patients with metabolic syndrome [130].
After oral administration of bergamot extract (BPF) to rats and patients for 30 days, BPF significantly reduced triglyceride levels and blood glucose. Meanwhile, BPF inhibited 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase activity and enhanced reactive vasodilation [113].
After diosmin was administered orally (100 mg/kg/day) for 45 days, both histological and biochemical parameters demonstrated antidiabetic effects in type 2 diabetic rats [133].
Based on the above-provided summary, the effect of citrus flavonoids on blood lipids was confirmed. We believe that further research on citrus flavonoids in metabolic diseases is valuable.

Conclusions
In conclusion, digesting more citrus fruits in our daily diet is likely to be beneficial to our health. Additionally, Citrus flavonoids play a certain role in anti-cancer effects, neuroprotection, and metabolic regulation. Among the many citrus flavonoids, nobiletin is a promising anti-angiogenic agent with great potential for cancer prevention and treatment. In terms of neuroprotection, nobiletin could improve learning and memory deficits, suggesting its anti-Alzheimer's disease and Parkinson's disease potential. Moreover, hesperidin and naringin could prevent Parkinson's disease-related dopaminergic neuron (DA) degeneration, oxidative damage, and cognitive impairment. In short, citrus flavonoids possess high nutritional and medicinal value.
The production of citrus fruits is abundant around the world, and there is still the problem of insufficient utilization of citrus fruits in juice. Therefore, it is important to study and develop the residues of the industrial production of citrus juice. The existing underutilization problem can in turn provide supplements for human health. Moreover, systematic and in-depth research on the neuroprotective effect of citrus flavonoids should be carried out to comprehensively, deeply, and accurately evaluate the neuroprotective effect of citrus flavonoids. The nutritional value of citrus plants as food is also worth of attention, and the content of citrus flavonoids may be used as an indicator to evaluate the nutritional value. Finally, the effects of the long-term use of citrus flavonoids on our health are worthy of further investigation.