Dietary bioactives and essential oils of lemon and lime fruits

Siyu Liu, Shiming Li*, Chi-Tng Ho,*

a Department of Food Science, Rutgers University, New Brunswick, New Jersey 08901, USA

b Hubei Key Laboratory of EFGI & RCU, College of Life Sciences, Huanggang Normal University, Huanggang 438000, China

Keywords:

Lemon

Lime

Essential oils

Flavonoids

Biological activity

A B S T R A C T

Human beings have consumed lemon (Citrus limon) and lime (Citrus aurantifolia or Citrus latiflia) for thousands of years. Among the variety of citrus families, lemon and lime are originated from the hybridization of citron with primitive papeda, hence they are similar from the nutritional and organoleptic standpoints,whereas very different from other citrus species such as orange and mandarin. Except for fresh produce, a significant percentage of lemon and lime are processed and separated as juice, essential oils, pulps and other products. Lemon and lime juice or fruit itself is rich in vitamins, minerals and flavonoids which are rich sources for human nutrition. Consumption of lemon and lime fruit or juice are beneficial for human health in the scope of urinary citrate increase, oxidative stress relief, improvement in lipid profiles and inflammation markers, neuroprotective effects among others. These beneficial effects of lemon and lime are not only because of their high vitamin C content but also other bioactive micronutrients such as flavonoids. Essential oils from lemon and lime have fresh and zesty aroma for perfumery and flavor applications for centuries.Compared with orange or mandarin, the integrated review for lemon and lime dietary bioactive compounds and essential oils is scarce. Therefore, in this review, we introduced the historical cultivation, consumption and process of lemon and lime, discussed the chemical and biological activities of phytochemicals in lemon and lime fruits and juice, and summarized volatile and non-volatile components in lemon and lime oil. This review may provide a comprehensive perspective for entire lemon and lime industry as well as their scientific values.

1. Introduction

1.1 Background

Pomelo (Citrus maximaorC. grandis), citron (C. medica),mandarin (C. reticul) and one recently discovered genusC. halimii[1]are the parent species of all the citrus known today. Various crosspollination among these biotypes derived all different citrus fruits in the market. Lemon and lime are both originated through the hybridization of citron with primitive papeda (C. cavalerieiorC. ichangensis) [2]. The origin of lemon is still not very clear and some people believe it is originated from China, India or Middle East.Back to the first and second centuries AD, lemon has been cultivated in the Middle East and Greece. The earliest documentary records about lemon is Rome imported lemons from North Africa in the second century AD. Also, there is a lemon mosaic found in the ruins of Pompeii. During the following centuries, lemon was cultivated and spread throughout the Roman Empires, Iberian Peninsula and the Mediterranean [3,4]. For lime, the origin of true limes is considered at Malaysia or the East Indian archipelago. The earliest botanical remains were found in the eastern Egyptian desert dated to the 11-13thcentury AD [5].

Lemon and lime generally are very similar to each other from the nutritional perspectives. There are many different types for both lemon and lime cultivated based on regional climate and processing aids. They both have a significant market share, followed by orange,among other citrus fresh fruits or fruit processing industry. Most beverage companies, like Coke or Pepsico, have lemonades or limonade section as their main product lines. Flavor houses extracted a lot of lemon and lime essential oils to create the fresh citrus sip.Citrus fiber and pectin are considered as premium quality for the application of hydrocolloids. With the increasing demand of healthy diet, the consumption of citrus fruit is keeping increasing due to its versatile usage for flavoring as well as its health beneficial effects.In this review, we first introduce the history of consumption and current industry processing for lemon and lime, then focus on a detailed discussion of bioactive components in juice and key aroma contributing molecules of essential oils.

1.2 History of consumption

Based on the historical documents, lemon was used as an ornamental plant in the early Islamic garden. In the medieval time,Egyptian made the lemonade first and then human consumption of lemon started. Moroccan Jewish people pickled lemons to make lemonade or as the garnish of drinks. By the mid-1 600s, lemonade has been popular in the Europe and then introduced to America in 18thcentury by the European immigrants. There are fewer records for the lime history, it was first cultivated in south Iraq and Persia.In 1747, James Lind [6]conducted the systemic experiment on the effect of lemon and lime as the treatment for scurvy. Nowadays, we know that the effect is due to its high content of vitamin C. Then from 19thcentury, British sailors had the lemon and lime as daily supplementation to prevent scurvy. Later on, European immigrants introduced the cultivation of lemon and lime to South America where is current main citrus growing area. Lemon and lime are very different from other popular sweet fruits. People cultivated lemon and lime to consume the juice which contains 4%–7% acids and has a very tart taste. Except for the nutritional value, the attractive zesty aroma from lemon and lime is another main reason for its popularity from the East to West. Other than making beverages, they are also used a lot in culinary. Lemon and lime juice are good to marinate fish because it can neutralize the unpleasant fishy aroma. Acids in lemon can combine with the amines inflsh and generate nonvolatile ammonium salts while the zesty lemon flavor was added into the dish.It is very common in Indian, Mexican and Thai dishes. Lemon and lime juice are also great to marinate meat before cooking because the acids can partially hydrolyze the tough collagen fibers in meat tissues so that tenderization can be achieved. The grated rind of lemon is often used as flavoring ingredients in bakery, dairy products or other dishes. Lime is the key ingredient for several highball cocktails such as margaritas and mojito.

1.3 Cultivation and processing of lemon and lime

1.3.1 Lemon

Lemon cultivation is mainly concentrated in sub-tropical areas.Argentina, US, Italy, Spain and South Africa are main countries growing lemon nowadays. There are many different types of lemon varieties. Based on different purpose and environments, different types of lemons are cultivated. For example, Eureka lemon is the most popular type and growing all over the world except for Italy and Spain because it can bear fruit year-round. Also, there is no thorn on lemon trees which helps the harvest process since hand-picking is still a favored method for lemon trees’ health. Lisbon lemon may be the second popular type. This type of lemon is very acidic and has no seeds inside. More importantly, Lisbon lemon can grow better in cooler temperature. Femminello St. Teresa lemon is mainly grown in Italy and the well-known type of Italian lemon for its high-quality fruit and high amount of lemon oil. Bearss lemon has many fruits grow on a single tree.

Most lemons harvested in US, South Africa and Italy are distributed to fresh produce market. However, Argentinian lemons are mainly processed to lemon oil, lemon juice and dried lemon peels.People have been processing lemon fruit to juice and lemon oil for centuries. Fig. 1A showed the dissection of lemon fruit. Flavedo is the dominant section for essential oil and pigments and pulp is the source of juice containing rich bioactive phytochemicals. Extraction of lemon essential oil is a delicate industrial process and more detail can be found in Akyildiz and Ağçam, 2014 [7]. There are two main extraction systems used in lemon processing industry: FMC (Food Machinery Corporation) juice extractor and Brown process. Used in orange industry first, FMC can extract and separate oil and juice from fruit in one step. There are upper and lower cups to hold a citrus fruit evenly so that juice can be squeezed without bursting. The upper and lower cutters cut the plugs to separate the peel from internal portion of fruit as well as introduce the internal portion of fruit to pre finisher tube. After the internal flesh is separated, the peel will be squeezed mechanically to release the essential oil from flavedo. In the two steps Brown process to extract the oil and juice from lemon, the flavedo of lemon will be firstly punctured by millions of needles continuously and generally on rotating rolls. Thereby the oil sacs on the flavedo will be damaged and essential oil released and washed out with water.In the second step after the oil is extracted, the de-oiled lemon is cut into half and squeezed to get the juice.

Fig. 1 Lemon and lime. (A) Dissection of lemon fruit; (B) Comparison of Persian lime and Key lime.

After the extraction and separation of oil and juice from lemon fruits, there are subsequent steps followed for quality optimization,such as winterization for wax removal from essential oil and mandatory pasteurization for juice. Based on different specifications,requirements and applications, more treatments could be conducted for essential oil and juice industry. As byproducts, lemon peels and pulps are normally dried in lemon processing facility and they are premium source to extract high-quality pectin which is considered as vegetarian gelatin. With the increasing demand for dietary fiber, citrus fiber from lemon is very promising in the market too.

1.3.2 Lime

本期木皮市场需求一般，总体销势下滑，行情也属稳中转跌，价格水平有所偏软。但进口木皮较比国产木皮销势还是要好一些。市场上麦当娜树瘤的进口天然木皮，售价高达170～200元/m2，雀眼枫木、红影、白影，以及环纹沙比利等高档树种的木皮制品标价在140元/m2上下，虽然买卖频率不高，但是大单需求几率不少，促使价位还是居高不下。本期科技木皮、染色木皮价格下落变化明显，例如桦木、杨木产品几块钱就能买到，枫木的市场售价也下到十几元钱。

Lime is mainly separated into two different varieties:1)C. latifoliaTanaka (Persian, Tahiti or Bearss lime) looks more like “green lemon” which are seedless fruits mainly cultivated in the US and Brazil; and 2)C. aurantifoliaSwingle (Key, Mexican or West Indian lime) is more sensitive to cold than the first type lime,C. latifolia, and more considered as “true lime”. It is normally cultivated in extremely hot climatic areas (e.g. Mexico and Peru) and the size of fruit is much smaller than lemon or the other lime [8]. Fig. 1B showed the different appearance of Key lime and Persian lime.

Similar to lemon, lime processing industry aims to separate the essential oil, lime juice and pulps or peels from lime fruits. The process of Persian lime is almost the same as lemon since they have the similar shape. Most Persian lime essential oil is cold pressed without contacting lime juice. However, unlike the extraction process of lemon oil, lime oil is sometimes extracted by steam distillation from lime juice especially for Key lime. Due to the much smaller size and thinner peel of Key lime than that of Persian lime or lemon,cold press of Key lime oil is very challenging in industrial processing system. Also in tradition, Mexican processors have been making lime oil distilled for centuries so that people, especially for American consumers, are more used to distilled lime oil aroma. The whole Key lime fruit is firstly pressed to generate the emulsion of juice and oil. Then direct steam distillation separates lime oil from juice. This process could be as long as 14 h. The unique distilled lime oil aroma is formed during this “cooking” period when the essential oil is in contact with acidic juice. Additionally, there are two types expressed lime oil extracted from Key lime without “cooking” process. Type A Key lime oil is separated from the juice oil emulsion by centrifuge and it has contacted with acidic juice. Type B is obtained by the same process as lemon without any contact with acidic juice [9]. Different processes produce different organoleptic characteristics of lime essential oil which will be discussed later.

2. Bioactive phytochemicals of lemon and lime

For lemon and lime processor, one whole lemon fruit might be separated to 5 different products: essential oil, lemon juice, frozen pulps, dried pulps and dried peel. Nowadays, the processors put the value mainly on lemon essential oil and lemon juice. Depending on different finished products, the pulps and peels will have different percent of albedo.

Historically, people started to consume lemon and lime widely since they realized these fruits can prevent the scurvy. Until today,increasing demands of fresh citrus fruits or freshly extracted juice or other beverages are also driven by its decent number of natural phytochemicals: citric acid, ascorbic acid, minerals and flavonoids among others.

There are 0.72 g/L ascorbic acid in fresh lemon juice and 0.35 g/L in fresh lime juice [10]. Other major known nutrients in lemon and lime juice are illustrated in Table 1 and Table 2. Lime juice components are similar to lemon juice. Citric acid is the most concentrated in lemon (48.0 g/L fresh juice from lemon fruit) and lime (45.8 g/L fresh juice from lemon fruit) among other fruits, it is the important index for qualifying the lemon and lime juice in compliance with Food and Drug Administration standards. Urinary citrate is a potent and important inhibitor of urinary crystallization.Hypocitraturia is a major risk factor for calcium urolithiasis because citrate retards stone formation by inhibiting the calcium oxalate nucleation process and binding with urinary calcium so that the free calcium concentration is lowered. It is critical for medical interventions to increase urinary citrate to manage the urolithiasis [11].In addition, the predominant mineral in lemon and lime juice is potassium while the content of sodium is very low [12,13]. This implied that they can play an important role in our common highsodium and low-potassium diet nowadays. Together with abundant amount of citric acid, lemon juice was studied to replace potassium citrate as the treatment of urinary calcium stones for hypocitraturia patients. It is suggested that fresh lemon juice (85 mL) or lemonade made from two middle size lemons (1 oz or about 140 g/middle size lemon) as the daily intake is effective to treat the hypocitraturia patients [14].

Table 1Most common nutrients from fresh juice of lemon [12].

Table 2Most common nutrients from fresh juice of lime [15].

From last century, more and more studies focused on distinct flavonoids in citrus fruits. Flavonoids are important secondary plant metabolites and are mainly present in plant tissues in relatively high concentrations as sugar conjugates. Citrus fruits are rich source of flavonoids. Traditionally, however, different types of lemon pulps and peels are considered as valueless by-products and either sold to pectin company or discarded, until decades ago, a decent amount of lemon flavonoids was recovered from residues after filtration process in lemon juice production and pectin extraction. Recently, Ledesma-Escobar et al. [16]compared different pretreatment and extraction methods on lemon to determine the extracted flavonoids contents. The results showed that sample dehydration prior to flavonoids extraction which provided better results than fresh samples. Lyophilization is more desirable due to the protection from high temperature. For extraction, ultrasound-assisted extraction is the best method to isolate the compounds, followed by microwave-assisted extraction and shaking extraction. Unsurprisingly, superheated liquid extraction is the least favorable due to the degradation under high temperature.

Flavonoids are mainly present in plant tissues with relatively high concentrations as sugar conjugates. There is one study tested hesperidin, eriocitrin and diosmin amount in different parts of mature lemon fruit tissues (Table 3). Hesperedin and eriocitrin are much more abundant in albedo than flavedo and pulp. Diosmin is richer inflavedo while very trace amount found in pulp which may due to its lower hydrophilicity [17]. As the very promising nutraceutical compounds, lemon flavonoids have a huge potential to be extracted and applied in different functional foods. Citrus flavonoids are special nutrients in citrus juice because they are rare in other type of fruits [18]. Epidemiological studies showed that flavonoids consumption is related to lower cancer risk and preventive effects of many other chronic disease such as inflammation and oxidative stress [17,19]. Unlike orange or mandarin, flavanone glycosides are high in lemon and lime but much fewer studies have performed in these citrus flavanones than orange flavones. Rutinose glycosides of citrus flavanones are dominant in lemon and lime. Different from the neohesperidose flavanone in grapefruit which impart the bitterness,rutinose flavanones give a neutral taste for lemon and lime. Table 4 listed the flavanones in lemon and lime and Fig. 2 showed chemical structures for both flavanone aglycones and glycosides [20]. Lemon contains more flavanones (about 27 mg aglycones/100 g edible fruit or juice) than lime (about 17 mg/100 g) in total. Hesperidin is the primary flavanone in both lemon (15.78 mg/100 g) and lime(15.64 mg/100 g). Followed is eriocitrin while there is much lower level contained in lime (1.38 mg/100 g) compared with lemon(9.46 mg/100 g). Both lemon and lime contain small amount of narirutin. Didymin and naringin are found in lemon but absent in lime. Neoeriocitrin is present in lime but not in lemon.

Fig. 2 Chemical structures for mainflavanone aglycones and glycosides in lemon and lime.

Table 3Distribution of hesperidin eriocitrin and diosmin in different mature fruit tissues.

Table 4Flavanones in lemon and lime (mg aglycone/100 g fresh fruit or juice) [17].

Hesperidin can influence vascular permeability, increase capillary resistance and has analgesic as well as anti-inflammatory properties [21,22]. It showed a significant impact to increase high density lipoprotein (HDL) and lower cholesterol, low density lipoprotein (LDL), total lipid as well as triglyceride levels in plasma [23]. It can quench oxygen free radicals which are important for cancer initiation and formation [24,25]. Diosmin, another lemon flavonoid, which is biosynthesized by dehydrogenation of hesperidin and has applied in pharmaceutical industry as an active pharmaceutical ingredient in non-prescription drugs. Diosmin can be used to treat circulatory system illness, chronic venous insufficiency and rheumatic arthritis [17]. It also showed blood lipid lowering ability and significant ameliorative effects for type 2 diabetes biochemical parameters [26,27]. Only lemon and lime have significant amount of eriocitrin among all other citrus fruits.Eriocitrin was reported to possess the best antioxidant activity among all other glycoside flavonoids in lemon and have the similar antioxidant activity as (–)-epigallocatechin gallate (EGCG) [28]. Due to its high antioxidant ability, it can maintain capillary integrity and peripheral circulation. Additionally, it was found to be stable even after heat treatment and storage of juices which implied its potential to be used in other nutritional products processing [17,29]. Lemon flavonoids, in general, can mitigate the oxidative stress for diabetic rats. Streptozotocin-induced diabetic rats were given the diet which contained 0.2% crude flavonoids, 0.2% eriocitrin and 0.2% hesperidin respectively. After 28 days period, all 3 groups of rats showed a significant decrease of diabetes-induced oxidative stress compared to control group [29]. Another study revealed that after 6 months consumption of aronia-enriched lemon juice, metabolic syndrome patients showed a significant improvement for cardiovascular risk factors (oxidized LDL, C-reactive protein and homocysteine) [30,31].Since this century, citrus flavonoids have been studied for their neuroprotective effects. First of all, bothin vitroandin vivostudies showed that main citrus flavonoids (e.g. hesperidin) can traverse the brain-blood barrier. Despite of their potent antioxidant activity,it has been proposed that flavonoids are more likely to exert their neuroprotective activities by: 1) modulation of intracellular signaling cascades which control the survival, death and differentiation of neurons; 2) affecting gene expression; and 3) interactions with mitochondria [32]. Research showed that hesperitin can trigger multiple prosurvival intracellular signaling pathways as well as induce proteins that promote cognition, prevent antioxidant stress or are resistant to Aβ associated neurotoxicity [33,34].

3. Volatile and nonvolatile compounds in lemon and lime peels

Although citrus flesh and juice are the main reason that people start cultivating and consuming citrus produce, essential oil of citrus fruits is currently much more valuable than juice or fresh fruits due to limited quantity and increasing demand. For example, there is only approximately 0.2% essential oil in one lemon fruit and about 60%–80% citrus fruit sold as fresh produce. Hence there is very limited quantity of natural citrus essential oils that can be obtained. Lemon and lime flavor are always the favorite starting point to touch for product developers in foods and other household items, because of the refreshing and sweetness-enhancing aroma the essential oils for human sensory feeling. Unlike other type of essential oils which are normally distilled from plants, citrus oil is mainly from cold press processing, except for lime, to preserve the authentic citrusy note. Due to its high value and various flavor application needs, many studies [35-37]have focused on the determination of volatile components and aroma active compounds for citrus fruits. Besides volatile chemicals, the oxygen heterocyclic compounds in lemon and lime cold-pressed essential oil are attracting more and more attention. Some studies [38-41]have revealed their antioxidant and anti-tumor effects for potential pharmaceutical purpose and also they are contributing the bodies of essential oil in sensory.

3.1 Lemon essential oil

Citral (neral and geranial) is almost the synonyms for lemon aroma. But, like other citrus oil,D-limonene is the predominant component (60%–70%) in single fold lemon essential oil. The following areβ-pinene,γ-terpinene,α-pinene, sabinene, myrcene,α-thujene and others (Table 5). Monoterpenes constitute 85%–95% of lemon oil, while the most active aroma chemical, citral is only 1.8%–2.5% of lemon oil. Other active aroma compounds are neryl acetate, geranyl acetate and citronellal among others. Different types of lemon may provide different aroma characteristics. For example,Meyer lemon is attracting more attention for its distinct aroma because it is the hybrid of citron and mandarin/pomelo. This species has significantly higher amount of citronellal, thymol and other sesquiterpenes while lower amount of citral and esters than regular lemons. Table 5 shows the main aroma components in Meyer lemon oil and regular lemon oil. Same as other crops, different location and season will also affect volatile compounds in lemon oil [35].For example, Italian or Spanish lemon oil could have 20% more sesquiterpenes (e.g.β-bisabolene) than Argentinian and California lemon oil. California lemon oil considered as with higher amount of citral compared to Argentinian lemon oil. Argentinian lemon has herniarin (7-methoxycourmarin), which is mostly found in lime, in the early season oil. Also, more monoterpens and alcohols (e.g. nerol and geraniol) detected in early season lemon oil.

Table 5Main aroma components comparison in Meyer lemon oil and Argentinian lemon oil.

Citral, as the key aroma contributing chemical in lemon oil, is the secondary metabolites of plants in order to protect the fruit and plant themselves from bacterial and pests. It has strong antibacterial,antifungal and antiparasitic effects [42,43]. Citral has also been reported for its anti-inflammatory effects. For example, citral can inhibit lipopolysaccharide (LPS)-induced acute lung injury by activating peroxisome proliferators-activated receptor γ and then inhibiting LPS-induced inflammatory response [44]. Additionally,citral can induce phase II xenobiotic metabolizing enzyme, glutathioneS-transferase (GST), to suppress the oxidative stress so that act as a cancer chemopreventive agent targeted on inflammation-related carcinogenesis such as skin cancer and colon cancer [43,45]. But citral has also been listed as fragrance allergen and irritant chemical,its dosage or formulation for skin cancer as a chemopreventive agent need to take extra cautions [46].

Although citral is the most important volatile chemicals for lemon essential oil, it might be one of the most unstable chemicals in the essential oil. In particular, under the acidic environment, the degradation of citral is rapid and expedited. Therefore, it is very important to separate the extracted lemon juice and oil during the processing. Unfortunately, soft drink (e.g. soda and lemonade) which is the main application of citrus scent are acidic solution (pH 2.5–4.0).Scientists have delved into its oxidation and degradation mechanisms since last century and various experiments were conducted to look for the inhibition of these reactions due to the strong unpleasant off-note chemicals generated from citral degradation such asp-cymene (gasolinic and smoky),p-cresol (smoky charcoal) andp-methylacetophenoen (cherriesh and powdery). The reaction is complicated and has different mechanisms (Fig. 3). It has been proposed that the linear chain of citral molecules will undergo an acid-catalyzed cyclization to form intermediate monoterpene alcohols:p-menthadien-8-ol andp-mentadien-4-ol. One pathway is that monoterpene alcohols can be oxidized top-cymene-8-ol followed its dehydration productsα,p-dimethylstyrene which can be oxidized top-methylacetophenone. Another pathway is that the monoterpene alcohols will be dehydrated top-mentha-1,4(8),5-triene which will be oxidized top-cymene under acid catalysis or to 8-hydroperoxyp-cymene to formp-cresol. Monoterpene alcohols can also formα-terpineol by disproportionation [47-50].

Fig. 3 Proposed degradation mechanisms of citral.

There is no experimental data that can significantly prove the inhibition of citral degradation in acidic environment under shelf-life study (typical beverage pH range is 2.5-4.0) [47-51]. Fortunately,some citral-like chemicals have been found and as long as their biological safety profiles have been supported by data, they may be the solution for citrus flavoring in the future. Cannon et al. [52]discovered some thiophenecarbaldehydes from fried chicken which have the similar sensory properties as citral or citrus flavor. 3-Butyl-2-thiophenecarbaldehyde is the most attractive compound among a total of 35 discussed thiophenecarbaldehyde derivatives. In terms of odor, it is described as citral-like, lemon and fatty. In terms of taste,it is described as lemon, citral-like, green and fatty. Although the odor threshold (151 ng/L) of 3-butyl-2-thiophenecarbaldehyde is higher than neral (8.8 ng/L) and geranial (12.0 ng/L), its stability is significantly better than citral. More importantly, the thiophenes show fewer off-notes and less profile change over time in comparison to citral in typical citrus beverage solutions (pH of 2.8).

Besides the well-known citral, volatile sulfur compounds have also been considered to contribute fresh lemon fruit aroma [36].Although the amount of volatile sulfur phyochemicals in lemon is too minute to be identified and quantified, Shaw et al. [53]has noticed the presence of hydrogen sul fide in the headspace of Eureka lemon juice as early as 1982. In 2015, Cannon et al. [54,55]has identified 445 volatile compounds extracted from Lisbon lemon peels which include 13 volatile sulfur components. There are two components showed odor activity by aroma extract dilution analysis (AEDA) in the experiment: 3-methylbut-2-ene-1-thiol (sulfurous, skunk-like)and 3-mercapto-3,7-dimethyl-6-octenyl acetate (sulfurous). The mechanism was hypothesized as direct addition of hydrogen sul fide to neryl acetate and geranyl acetate, or the adduct ofL-cystein orL-glutathion with neryl acetate and geranyl acetate followed by a specific enzyme to release the thiol.

3.2 Lime essential oil

As discussed previously, there are different types of lime essential oils based on different lime fruit species and processing methods.Table 6 showed some major volatile components for distilled lime oil and cold pressed lime oil. In cold pressed lime oil, citral is about 3.5%–5%, which is much higher than lemon oil. The analytical profile of volatile compounds is similar to lemon oil but with lower amount of monoterpenes, higher amount of sesquiterpenes and aldehydes as well as esters. For cold pressed lime oil (Key lime), there are two different processes: physical separation of oil phase from oil/juice emulsion made expressed type A of Key lime oil and oil extraction from lime fruit skin without contacting lime juice called expressed type B of Key lime oil. Along with lime oil expressed Persian, there are at least 3 different types of cold pressed lime oils. Dugo et al. [56]has compared these 3 different cold pressed lime oils. The total monoterpenes in Persian lime oil is higher than Key lime oil. There are 50%–60% of limonene in Persian oil, whereas less than 50% in Key lime oil. There is much higher amount ofγ-terpinene in Persian lime (about 13%) than Key lime (about 8%), but more pinenes in Key lime oil than in Persian lime. Non-volatile residue shows higher values for Key lime oil (about 13%) than Persian lime (about 9%).But the ultraviolet-absorbance values for 2 different lime oils are similar, which revealed the quantity of coumarins and psoralens in different species of lime oils are similar. Noticeably, esters in Persian lime oil are 3 to 4 times higher than Key lime oil. And for Key lime oil, type A oil showed a higher content of monoterpene alcohols(e.g. terpineols) than type B oil which can be explained by its process in which type A oil contacts low pH juice and causes the hydration of monoterpene hydrocarbons with formation of alcohols [56,57].

Table 6Comparison of major volatile components (%) for distilled lime oil and cold pressed lime oil.

Distilled lime oil has very different aroma and amounts of components compared with cold-pressed lime oil because reactions occur during long-time “cooking” process for oil and juice emulsion.Almost all the citral will be decomposed by this distillation process and high amount of terpineols (piney) existing in distilled lime oil.Together with terpinolene, cineols,p-cymene, fenchol, borneol and a lot of other degradation or oxidation products will give the distilled lime oil distinct sweet limey aroma. This is because monoterpenes are also inclined to be oxidized under acidic and heating environment although the reaction is not as active as citral.For example,D-limonene will also generateα-terpineol under acidcatalyzed oxidation reactions. Reaction pathways include hydrations of double bonds, and dehydrations, rearrangements, cyclizations and hydrolyses of esters in diverse combinations [58]. However, aqueous acid-catalyzed reactions do not produce “off-notes” flavor like citral.Also, it could be that consumers are already very well adapted these flavor notes since the lime flavors are dominated by distilled lime oil in some popular commercial beverages such as Sprite or SevenUp beverages in US.

Besides the aldehydes and esters that are considered as potent aroma contributors, Clark et al. [59]has pointed out that germacrene B, one of the sesquiterpenes, is very important to lime peel oil aroma,and the amount of germacrene B is quantified to be about 0.35% of the whole oil. Also in this study, germacrenes has been synthesized and described as potent, warm, sweet, woody spicy, geraniumlike odor that is very important to the fresh lime peel oil character.Another study showed that the heat treatment for distillation will also affect the lime sesquiterpenes a lot. Feger et al. [8]revealed that regular gas chromatography (GC) condition is not good for analyzing the sesquiterpenes of cold pressed lime oil because germacrene A and C are thermally and chemically sensitive compounds. Fig. 4 illustrates the structures of germacrenes. Similar to germacrene B, germacrene A, C and D are also important to fresh lime peel aroma and the total of germacrene A, B, C and D makes up about 25% expressed lime oil sesquiterpenes. In the study, only under unusually mild GC conditions with a temperature at or below 100 °C throughout whole GC system,the germacrene A and C can be determined due to their sensitivity to higher temperature. Five sesquiterpens, namelyα-santalene,γ-curcumene,β-selinene,β-sesquiphellandrene and 7-epi-α-selinene were detected under regular GC conditions as well as in distilled lime oil, which also has been treated with high temperature. Moreover,bothα-amorphene andδ-cadinene are the acidic transformations of germacrene D and observed only in distilled lime oil [8].

Fig. 4 Chemical structures of germacrene isomers.

3.3 Oxygen heterocyclic compounds in lemon and lime essential oil

Except for volatile chemicals, there are about 2% of non-volatile chemicals in lemon essential oil, and which is approximately 20% in cold pressed lime essential oil. Besides wax and colorants, they are mainly coumarin and psoralene derivatives. Table 7 shows the main oxygen heterocyclic compounds in lemon and lime oil. There are 14 oxygen heterocyclic compounds identified in general lemon oil and 16 compounds in cold-pressed lime oil (both Key lime and Persian lime).The total amount of oxygen heterocyclic compounds in lime oil is more than 10 times than that in lemon oil which is in compliant with non-volatile quantities. For distilled lime oil, there is no non-volatile determined since it is made by distillation process. In lemon, the main oxygen heterocyclic compound is citroptene. In lime, bergamottin and 5-geranyloxy-7-methoxycoumarin are the most abundant compounds. Herniarin, isopimpinellin, bergapten and 5-geranyloxy-8-methoxycoumarin have also been identified in cold-pressed lime oil [56]. Different processing methods will affect the oxygen heterocyclic compounds too. Oxypeucedanin has been reported absent in type A Key lime oil while it is present in type B and Persian oil. It is proposed that the epoxy ring in oxypeucedanin will be hydrolyzed if water and juice exist and the formed oxypeucedanin hydrate is water-soluble [57].

Table 7Main oxygen heterocyclic compounds in cold pressed lemon and lime oil.

Coumarins are reported to possess biological activities such as antibacterial [60], antioxidant, anti-platelet-aggregating, antimutagenic and anti-tumor effects [38,61]. Citropten, the most abundant coumarin in lemon essential oil, showed a strong inhibitory effect on the expression of interleukin 8 (IL-8), demonstrating its anti-inflammatory ability. Study proposed citropten as potential anti-inflammatory molecule in reducing lung inflammation in cystic fibrosis patients [39]. 5-Geranyloxy-7-methoxycoumarin showed the greatest inhibitory activity on the proliferation of human colon cancer (SW-480) cell compared with citropten and isopimpinellin.The mechanism was proposed that 5-geranyloxy-7-methoxycoumarin can induce cell apoptosis by activating tumor suppressor genep53andcaspase8/3while negatively regulating the phosphorylation of Bcl2 and p38 MAPK [38]. 5-Geranyloxy-7-methoxycoumarin and bergamottin are considered as bioactive compounds for antiproliferative effects on SH-SY5Y human neuroblastoma cell growth [56]. Oxypeucedanin, the unique furocoumarin for lemon and lime compared with other citrus fruits, has been found to have anticancer effect by inducing G2-M cell cycle arrest and apoptosis in human prostate carcinoma DU145 cells [41]. Furacoumarins are known for their toxic effects [62]. Epidemiological study showed that the risk of skin cancer is potentially related to relatively high level of dietary exposure of furocoumarins due to their phototoxic,photomutagenic and photocarcinogenic properties [63]. But bergamottin itself does not show any significant photomutagenicity [64].Studies has showed that bergamottin has significant anti-cancer activity in skin, myeloma, leukemia, lung cancer and other cancer cells. These underlying mechanisms of anti-cancer activities is that bergamottin is able to regulate several cancer-related pathways including chemical detoxification, cell cycle arrest, apoptosis,migration, invasion, and angiogenesis [65]. Although bergamottin is very promising as a natural anti-cancer agent, it can also inhibit the cytochrome P450 (CYP) enzyme (CYP3A4) [61]. The clinical trial for bergamottin need to take extra caution since it could interfere with the metabolism of other drugs [66].

Most flavor and perfumery companies relied on GC for their quality control tests. With the high economic motivation, coldpressed essential oil has been adulterated with other inexpensive materials such as orange terpenes or orange oil for a long time.Nowadays, these non-volatile oxygen heterocyclic compounds have been routinely tested on HPLC as quality control (QC) to identify the authenticity of natural citrus essential oil instead of the traditional total UV absorbance method. However, as part of essential oil that imparts the body note of diverse citrus aroma, the sensory profiles of these components have not been thoroughly studied. For example, if the orange residue was added to lemon oil to boost its UV absorbance value, the adulterated oil will taste more orange even with the same volatile components based on GC. On the other hand, citropten and herniarin has been described as sweet lactone-like, vanilla-like.This may be another reason why these citrus flavors can enhance sweetness. Therefore, it will be interesting to know the contribution of different oxygen heterocyclic components for the citrus aroma as well as their ability to enhance or mask the flavor.

4. Summary

Lemon and lime are main fruits cultivated and consumed among the giant citrus family. They provide us the great source of dietary bioactive compounds such as vitamin C, potassium, citric acid and flavonoids. Epidemiological studies revealed that lemon or lime are very promising to provide natural bioactives and phytochemicals such as hesperedin and eriocitrin to positively regulate oxidative stress, lipid profiles as well as inflammatory cascades especially for people with metabolic syndromes. Moreover, they are also potentially to exert the neuroprotective effects and become general dietary brain foods due to their abundant flavonoids contents. However,the research on effective dosage of lemon or lime consumption to reduce the risk of cardiovascular disease or promote brain health is very rare. Lemon and lime are very versatile in culinary or drinks. It will be very interesting to know how many lemon or lime need to be consumed as daily diet from the nutritional standpoint. Besides fresh fruit or juice, essential oils extracted from peels are very valuable goods for flavoring and perfumery purpose currently. Therefore, the processing industry for lemon and lime has been established and optimized for centuries to separate and transport the nutrients as well as zests across the equator without the compromises on their stability.Evidence has revealed that the coumarins or furocoumarins are potent anti-tumor and anti-cancer candidates especially for bergamottin and 5-geranyloxy-7-methoxycoumarin. But there are very few studies focused on the bioactivity and bioavailability of 5-geranyloxy-7-methoxycoumarin as natural anti-tumor and anti-cancer compound.Since it is very abundant in lemon and lime peel, future study should be focused on the biological effects on 5-geranyloxy-7-methoxycoumarin. In summary, this review provides the full-scale image of lemon and lime about their unique nutrients, aroma as well as the industrialized products which are ubiquitous in our daily life nowadays.

Conflict of interest

The authors declare that there is no conflict of interests.

Acknowledgement

This work was supported by Hubei Science and Technology Plan Key Project (G2019ABA100).