Plant-based insect repellents: a review of their efficacy, development and testing

  • Marta Ferreira Maia1, 2 and

    Affiliated with

    • Sarah J Moore1, 2Email author

      Affiliated with

      Malaria Journal201110(Suppl 1):S11

      DOI: 10.1186/1475-2875-10-S1-S11

      Published: 15 March 2011

      Abstract

      Plant-based repellents have been used for generations in traditional practice as a personal protection measure against host-seeking mosquitoes. Knowledge on traditional repellent plants obtained through ethnobotanical studies is a valuable resource for the development of new natural products. Recently, commercial repellent products containing plant-based ingredients have gained increasing popularity among consumers, as these are commonly perceived as “safe” in comparison to long-established synthetic repellents although this is sometimes a misconception. To date insufficient studies have followed standard WHO Pesticide Evaluation Scheme guidelines for repellent testing. There is a need for further standardized studies in order to better evaluate repellent compounds and develop new products that offer high repellency as well as good consumer safety. This paper presents a summary of recent information on testing, efficacy and safety of plant-based repellents as well as promising new developments in the field.

      Background

      Most plants contain compounds that they use in preventing attack from phytophagous (plant eating) insects. These chemicals fall into several categories, including repellents, feeding deterrents, toxins, and growth regulators. Most can be grouped into five major chemical categories: (1) nitrogen compounds (primarily alkaloids), (2) terpenoids, (3) phenolics, (4) proteinase inhibitors, and (5) growth regulators. Although the primary functions of these compounds is defence against phytophagous insects, many are also effective against mosquitoes and other biting Diptera, especially those volatile components released as a consequence of herbivory [1]. The fact that several of these compounds are repellent to haematophagous insects could be an evolutionary relict from a plant-feeding ancestor, as many of these compounds evolved as repellents to phytophagous insects [2], and this repellent response to potentially toxic compounds is well conserved in the lineage of Diptera (True Flies). Insects detect odours when that volatile odour binds to odorant receptor (OR) proteins displayed on ciliated dendrites of specialized odour receptor neurons (ORNs) that are exposed to the external environment, often on the antennae and maxillary palps of the insect, and some ORNs, such as OR83b that is important in olfaction and blocked by the gold-standard synthetic repellent DEET (N, N-diethyl-3-methylbenzamide) [3], are highly conserved across insect species [4, 5]. Plants commonly produce volatile “green leaf volatiles” when leaves are damaged in order to deter herbivores [6], and several authors have shown strong responses of mosquito odour receptors to this class of volatiles including geranyl acetate and citronellal [7], 6-methyl-5- hepten-2-one and geranylacetone [8]. Interestingly, the same odour receptors that respond to DEET also respond to thujone eucalyptol and linalool in Culex quinquefasciatus[9]. In Anopheles gambiae, the DEET receptor OR83b is stimulated by citronellal, but is also modulated by the TRPA1 cation channel [10]. However, it is most likely that many plant volatiles are deterrent or repellent because they have high vapour toxicity to insects [11, 12].

      This repellency of plant material has been exploited for thousands of years by man, most simply by hanging bruised plants in houses, a practice that is still in wide use throughout the developing countries [13]. Plants have also been used for centuries in the form of crude fumigants where plants were burnt to drive away nuisance mosquitoes and later as oil formulations applied to the skin or clothes which was first recorded in writings by ancient Greek [14], Roman [15] and Indian scholars [16] (Figure 1). Plant-based repellents are still extensively used in this traditional way throughout rural communities in the tropics because for many of the poorest communities the only means of protection from mosquito bites that are available [13], and indeed for some of these communities [17], as in the Europe and North America [18] “natural” smelling repellents are preferred because plants are perceived as a safe and trusted means of mosquito bite prevention.
      http://static-content.springer.com/image/art%3A10.1186%2F1475-2875-10-S1-S11/MediaObjects/12936_2011_1642_Fig1_HTML.jpg
      Figure 1

      Moghul painting illustrating a man burning neem leaves near a river where biting insects would be present (© Dr Sarah Moore)

      The discovery of new plant-based repellents is heavily reliant on ethnobotany. This is the targeted search for medicinal plants through in-depth interviews with key informants knowledgeable in folk-lore and traditional medicine. It is common practice to conduct ethnobotanical surveys using structured interviews, combined with the collection of plant voucher Specimens (Figure 2), to evaluate plant use by indigenous ethnic groups [19]. Questions are asked about plant usage, abundance and source. This is a more direct method of identifying plants with a potential use than general screening of all plants in an area. A second means is bio-prospecting, where plants are systematically screened for a particular mode of action, which is a costly and labour intensive means of identifying new repellents. However, mass screening of plants for repellent activity was the way by which PMD (para-methane 3-8, diol), an effective and commercially available repellent was discovered in the 1960s [20].
      http://static-content.springer.com/image/art%3A10.1186%2F1475-2875-10-S1-S11/MediaObjects/12936_2011_1642_Fig2_HTML.jpg
      Figure 2

      A village herbalist in rural Yunnan, Southern China. This lady was a key informant for an ethnobotanical study into plants used to repel mosquitoes (© Dr Sarah Moore)

      PMD from lemon eucalyptus (Corymbia citriodora) extract

      Corymbia citriodora (Myrtaceae), also known as lemon eucalyptus, is a potent natural repellent extracted from the leaves of lemon eucalyptus trees (Table 1). It was discovered in the 1960s during mass screenings of plants used in Chinese traditional medicine. Lemon eucalyptus essential oil, comprising 85% citronellal, is used by cosmetic industries due to its fresh smell [21]. However, it was discovered that the waste distillate remaining after hydro-distillation of the essential oil was far more effective at repelling mosquitoes than the essential oil itself. Many plant extracts and oils repel mosquitoes, with their effect lasting from several minutes to several hours (Table 1). Their active ingredients tend to be highly volatile, so although they are effective repellents for a short period after application, they rapidly evaporate leaving the user unprotected. The exception to this is para-menthane 3, 8 diol, which has a lower vapour pressure than volatile monoterpines found in most plant oils [22] and provides very high protection from a broad range of insect vectors over several hours [23], whereas the essential oil is repellent for around one hour [24]. PMD is the only plant-based repellent that has been advocated for use in disease endemic areas by the CDC (Centres for Disease Control) [25], due to its proven clinical efficacy to prevent malaria [26] and is considered to pose no risk to human health [27]. It should be noted that the essential oil of lemon eucalyptus does not have EPA (Environmental Protection Agency) registration for use as an insect repellent.
      Table 1

      An overview of repellent plant efficacy from literature review

      Plant

      Location

      Other names

      Repellent compound(s)

      Tested mode of use

      Repellency % protection

      Study type

      Ref

      MYRTACEAE

             

      Corymbia citriodora

      Australia

      Brazil

      Bolivia

      China

      India

      Ethiopia

      Tanzania

      Kenya

      lemon eucalyptus

      lemon scented gum

      quwenling

      citronellal

      PMD (by product of

      hidrodistillation)

      (p-menthane-3,8-diol)

      citronellol

      limonene

      geraniol

      isopulegol

      δ-pinene

      30% PMD applied topically

      96.88% protection from mosquitoes for 4 hours

      field study in Bolivia

      [35]

          

      PMD towelette (0.575g) applied topically

      90% protection from An. arabiensis for 6 hours

      laboratory study

      [95]

          

      50% PMD applied topically

      100% protection from An gambiae and An. funestus for 6-7 hours

      field study in Tanzania

      [96]

          

      20% PMD (1.7 mg/cm2) applied topically

      100% protection for 11-12 hours against A. stephensi

      laboratory study

      [52]

          

      20% PMD applied topically

      100% protection against Ae. Aegypti for 120 minutes

      Laboratory study

      [42]

          

      thermal expulsion (leaves)

      78.7 % protection from An. arabiensis

      76.8% protection from An. pharaoensis

      field study in Ethiopia

      [97]

          

      direct burning (leaves)

      70.1 % protection from An. arabiensis

      72.9% protection from An. pharaoensis

      field study in Ethiopia

      [97]

          

      periodic thermal expulsion (leaves)

      74.5% protection from An. gambiae s.s.

      semi-field study in Kenya

      [50]

          

      periodic direct burning (leaves)

      51.3% protection from An. gambiae s.s.

      semi-field study in Kenya

      [50]

          

      thermal expulsion (leaves)

      48.71% protection from An. gambiae s.l.

      field study in Kenya

      [98]

      Eucalyptus spp.

      Guinea-Bissau

      Ethiopia

      Tanzania

      Portugal

      eucalyptus

      1,8-cineole

      citronellal

      Z- and α- citral

      α-pinene

      thermal expulsion (leaves)

      72.2% protection from mosquitoes for 2 hours

      field study in Guinea Bissau

      [99]

      E. camaldulensis

      Ethiopia

        

      thermal expulsion (leaves)

      71.9 % protection from An. arabiensis

      72.2% protection from An. pharaoensis

      field study in Ethiopia

      [97]

          

      direct burning (leaves)

      65.3 % protection from An. arabiensis

      66.6% protection from An. pharaoensis

      field study in Ethiopia

      [97]

      Eugenia caryophyllus or Syzygium aromaticum or Eugenia aromaticu

      India

      clove

      lavang

      cravinho-da-india

      Eugenol

      carvacrol

      thymol

      cinnamaldehyde

      100% essential oil applied topically

      100% protection against Ae. aegypti for 225 minutes

      100% protection against An. albimanus for 213 minutes

      laboratory study

      [53]

          

      100% essential oil applied topically

      100% protection against Ae. aegypti for 120 min.

      100% protection against C. quinquefasciatus for 240 min.

      100% protection against An. dirus for 210 min.

      laboratory study

      [23]

      VERBENACEAE

             

      Lippia spp.

      Kenya

      Tanzania

      Ghana

      Zimbabwe

      lemon bush

      myrcene

      linalool

      α-pinene

      eucalyptol

          

      L. javanica

        

      alloparinol

      camphor

      limonene

      α –terpeneol

      verbenone

      5mg/cm2 plant extract applied topically

      100% protection against Ae. aegypti for 8 hours

      laboratory study

      [100]

          

      alcohol plant extract applied topically

      76.7% protection against An arabiensis for 4 hours

      laboratory study

      [101]

      L. uckambensis

       

      fever tea

       

      potted plant

      33.3% protection against An. gambiae s.s

      semi-field study in Kenya

      [102]

          

      periodic thermal expulsion (leaves)

      45.9% protection against An. gambiae s.s.

      semi-field system in Kenya

      [50]

          

      periodic direct burning (leaves)

      33.4% protection against An. gambiae s.s

      semi-field system in Kenya

      [50]

          

      potted plant

      25.01% protection against An.gambiae s.l

      field study in Kenya

      [98]

      L. cheraliera

        

      eucalyptol

      caryophyllene

      ipsdienone

      p-cymene

          

      Lantana camara

      Kenya

      Tanzania

      lantana

      spanish flag

      West Indian

      lantana

      Wild sage

      caryophylene

      potted plant

      32.4% protection against An. gambiae s.s

      semi-field study in Kenya

      [102]

          

      potted plant

      27.22% protection against An. gambiae s.l.

      field study in Kenya

      [98]

          

      flower extract in coconut oil

      94.5% protection against Ae. aegypti and Ae. albopictus for one hour

      laboratory study

      [103]

          

      periodic thermal expulsion (leaves)

      42.4% protection against An. gambiae s.s

      semi-field study in Kenya

      [50]

      LAMIACEAE

             

      Ocimum spp. O.americanum

      Kenya

      Tanzania

      Zimbabwe

      Nigeria

      Ghana

      Cameroon

      Eritrea

      Ethiopia (…)

      Tree basil

      nchu avum

      lime basil

      kivumbasi

      Myeni madongo

      African blue basil

      hairy basil

      p-cymene

      estragosl

      linalool

      linoleic acid

      eucalyptol

      eugenol

      camphor

      citral

      thujone

      limonene

      ocimene

      and others

      potted plant

      39.70% protection against An. gambiae s.s

      semi-field study in Kenya

      [102]

          

      potted plant

      37.91% protection against An. gambiae s.l.

      field study in Kenya

      [98]

          

      fresh plants combined with O. suave bruised and applied topically

      50% protection against An. gambiae s.l.

      field study in Tanzania

      [104]

          

      periodic thermal expulsion (leaves and seeds)

      43.1.% protection against An gambiae s.s

      semi-field study in Kenya

      [50]

          

      periodic direct burning (leaves and seeds)

      20.9% protection against An. gambiae s.s

      semi-field study in Kenya

      [50]

          

      100% essential oil combined with vanillin 5% applied topically

      100% protection against Ae. aegypti for 6.5 hours1

      100% protection against C. quinquefasciatus for 8 hours

      100% protection against An. dirus for 8 hours

      laboratory study

      [26]

      O. suave

         

      thermal expulsion (leaves)

      73.6 % protection from An. arabiensis

      75.1% protection from An. pharaoensis

      field study in Ethiopia

      [97]

          

      direct burning (leaves)

      71.5 % protection from An. arabiensis

      79.7% protection from An. pharaoensis

      field study in Ethiopia

      [97]

          

      periodic thermal expulsion (leaves and seeds)

      53.1% protection from An. gambiae s.s.

      semi-field study in Kenya

      [50]

          

      periodic direct burning (leaves and seeds)

      28.0% protection from An. gambiae s.s.

      semi-field study in Kenya

      [50]

      O. basilicum

         

      thermal expulsion (leaves)

      78.7 % protection from An. arabiensis

      79.2% protection from An. pharaoensis

      field study in Ethiopia

      [97]

          

      direct burning (leaves)

      73.1 % protection from An. arabiensis

      70.0% protection from An. pharaoensis

      field study in Ethiopia

      [97]

          

      100% essential oil applied topically

      100% protection for 70 minutes

      laboratory study

      [23]

      O. kilimandscharikum

         

      thermal expulsion (leaves and seeds)

      44.54% protection against An. gambiae s.l.

      field study in Kenya

      [98]

          

      thermal expulsion (leaves and seeds)

      37.63% protection against An. funestus

      field study in Kenya

      [98]

          

      periodic thermal expulsion (leaves and seeds)

      52.0% protection against An. gambiae s.s.

      semi-field study in Kenya

      [50]

          

      periodic direct burning (leaves and seeds)

      26.4% protection against An. gambiae s.s

      semi-field study in Kenya

      [50]

      O. forskolei

         

      fresh plants hung indoors

      53% protection against mosquitoes entering human dwelling

      field study in Eritrea

      [105]

      Hyptis spp. Hyptis suaveolens

      Kenya

      Tanzania

      Ghana

      The Gambiae

      bushmint

      wild hops

      wild spikenard

      hangazimu

      hortelã-do-campo

      myrcene

      smouldering on charcoal

      85.4% repellency against mosquitoes for 2 hours

      field study in Guinea Bissau

      [99]

          

      fresh leaves

      73.2% repellency against mosquitoes for 2 hours

      field study in Guinea Bissau

      [99]

          

      periodic direct burning (leaves and flowers)

      20.8% repellency against An. gambiae s.s

      semi-field study in Kenya

      [50]

      Mentha spp. M. piperata

      Brazil

      Bolivia

      hortelã-do-campo

      peppermint

       

      100% essential oil applied topically

      100% protection against Ae. aegypti for 45 minutes

      laboratory study

      [53]

      M. arvensis

       

      menta

      Japanese mint

       

      100% essential oil volatilized in a kerosene lamp

      41% protection indoors against Mansonia spp

      field study in Bolivia

      [9]

      Thymus spp. Th. vulgaris

      China

      Former Soviet

      Union

      Korea

      Middle-East

      Mediterranean

      thyme

      α-terpinene

      carvacrol

      thymol

      p-cymene

      linalool

      geraniol

      α-terpinene topically

      97.3% protection against Culex pipiens sallens for 82 min

      laboratory study

      [106]

          

      carvacrol topically

      94.7% protection against C. pipiens sallens for 80 min

        
          

      thymol topically

      91.8% protection against C. pipiens sallens for 70 min

      laboratory study

      [106]

          

      linalool topically

      91.7% protection agains C. pipiens sallens for 65 min

        
          

      p-cymene

      89.0% protection agains C. pipiens sallens for 45.2 min

        
          

      100% essential oil applied topically

      100% protection against An. albimanus for 105 minutes and Ae. aegypti for 135 minutes

      laboratory study

      [53]

          

      direct burning (leaves)

      85-09% protection for 60-90 min

      field study

      [12]

      Pogostemon spp.

      China

      Patchouli

       

      100% essential oil applied

      100% protection against Ae. aegypti for 120 min

      laboratory study

      [23]

      Pogostemon cablin

      India

      Malaysia

      Thailand

      Oriza

       

      topically

      100% protection against C. quinquefasciatus for 150 min

      100% protection against An. dirus for 710 minutes

        

      POACEAE

             

      Cymbopogon spp.

      China

      India

      Indonesia

            

      C. nardus

      Brazil

       

      citronellal

      40% essential oil applied topically

      100% protection for 7-8 hours against An. stephensi

      laboratory study

      [52]

          

      100% essential oil applied topically

      100% protection against Ae. aegypti for 120 min

      100% protection against C. quinquefasciatus for 100 min

      100% protection against An. dirus for 70 minutes

      laboratory study

      [23]

          

      10% applied topically

      100% protection against Ae. aegypti for 20 minutes

      laboratory study

      [42]

      C. martini

      Tanzania

      Kenya

      palmarosa

      geraniol

      topically

      (100% essential oil)

      100% protection against An. culicifacies for 12 hours

      96.3% protection against C. quinquefasciatus for 12 hours

      field study in India

      [107]

          

      topically

      (100% essential oil)

      98.8% protection against C. quinquefasciatus for 10 hours

      laboratory study

      [107]

      C. citratus

      USA

      South África Bolívia

      lemongrass oil grass

      citral α-pinene

      topically

      74% protection against An. darlingi for 2.5h

      95% protection against Mansonia spp. for 2.5 hours

      field study in Bolivia

      [9]

          

      Methanol leaf extract applied topically (2.5mg/m2)

      78.8 % protection against An. arabiensis for 12 hours

      laboratory study

      [108]

          

      100% essential oil applied topically

      100% protection for 30 minutes

      laboratory study

      [23]

      C. winterianius

         

      100% essential oil combined with vanillin 5% applied topically

      100% protection against Ae. aegypti for 6.5 hours

      100% protection against C. quinquefasciatus for 8 hours

      100% protection against An. dirus for 8 hours

      laboratory study

      [26]

      C. excavatus

         

      alcohol plant extract applied topically

      66.7% protection against An. arabiensis for 3 hours

      laboratory study

      [101]

      Pelargonium reniforme

       

      rose geranium

       

      alcohol plant extract applied topically

      63.3 protection against An. arabiensis for 3 hours

      laboratory study

      [101]

      MELIACEAE

             

      Azadirachta indica

      India

      Sri Lanka

      China

      Brazil Bolívia

      Pakistan

      Ethiopia

      Guinea Bissau

      Kenya

      Tanzania (…)

      Neem

      azadirachtin

      saponins

      direct burning (leaves)

      76.0% protection from mosquitoes for 2 hours

      field study in Guinea Bissau

      [99]

          

      periodic thermal expulsion (leaves)

      24.5% protection from An. gambiae s.s

      semi-field study in Kenya

      [50]

          

      1% neem oil volatilized in a kerosene lamp

      94.2% protection from Anopheles spp.

      80% protection from Culex spp.

      field study in India

      [109]

          

      2% neem oil applied topically

      56.75% protection from mosquitoes for 4 hours

      field study in Bolivia

      [35]

      ASTERACEAE

             

      Tagetes minuta

      Uganda

      Zimbabwe

      India

      Khaki weed

       

      topically

      86.4% protection againt An. stepehensi for 6 hours

      laboratory study

      [110]

          

      topically

      84.2% protection against C. quinquefasciatus for 6 hours

      laboratory study

      [110]

          

      topically

      75% protection against Ae. aegypti for 6 hours

      laboratory study

      [110]

          

      fresh leaves (4Kg)

      reduced human landings indoors

      field study in Uganda

      [111]

      Artemisia spp.

      A. vulgaris

      India

      Egypt

      Italy

      Canada

      USA

      mugwort

      wormwood

      St. Johns plant

      Old uncle henry

      Sailors tobacco

      camphor

      linalool terpenen-4-ol

      α-and β-thujone

      β-pinene

          

      A. monosperma

      Siberia Brazil

      Felon herb

      Naughty man

      myrcene

      limonene

      cineol

      5% leave extract applied topically

      100 % protection for 4 hours

      field study in Egypt

      112

      CAESALPINIACEAE

             

      Daniellia oliveri

      Guinea-Bissau

      The Gambiae

      churai

      santão

      santang

      santango

       

      direct burning (bark)

      77.9% protection against mosquitoes for 2 hours

      field study in Guinea Bissau

      [99]

          

      direct burning (bark)

      77% protection against mosquitoes

      field study in The Gambiae

      113

      FABACEAE

      Glycine max

      Worldwide

      Soya

       

      2% soya bean oil

      100% protection against Ae. aegypti for 95 minutes

      laboratoty study

      [42]

      RUTACEAE

      Zanthoxylum limonella

      Thailand

      makaen

       

      100% essential oil applied topically

      100% protection against Ae. aegypti for 120 min

      100% protection against C. quinquefasciatus for 170 min

      laboratory study

      [23]

          

      10% essential oil combined with 10% clove oil

      100% protection against An. dirus for 190 minutes

      laboratory study

      [52]

      Citrus hystrix

      Indonesia

      Malaysia

      Thailand

      Laos

      Kaffir lime

      Limau purut

       

      100% essential oil combined with vanillin 5% applied topically

      100% protection against An. stephensi for 8 hours

      100% protection against Ae. aegypti for 3 hours

      100% protection against C. quinquefasciatus for 1.5 hours

      100% protection against An. dirus for 2.5 hours

      laboratory study

      [26]

      ZINGIBERACEAE

      Curcuma longa

       

      Turmeric

      Curcuma

      Indian saffron

       

      100% essential oil combined with vanillin 5% applied topically

      100% protection against Ae. aegypti for 4.5 hours

      100% protection against C. quinquefasciatus for 8 hours

      100% protection against An. dirus for 8 hours

      laboratory study

      [26]

      Citronella

      Essential oils and extracts belonging to plants in the citronella genus (Poaceae) are commonly used as ingredients of plant-based mosquito repellents (Table 1), mainly Cymbopogon nardus that is sold in Europe and North America in commercial preparations. Citronella has found its way into many commercial preparations through its familiarity, rather than its efficacy. Citronella was originally extracted for use in perfumery, and its name derives from the French citronelle around 1858 [28]. It was used by the Indian Army to repel mosquitoes at the beginning of the 20th century [29] and was then registered for commercial use in the USA in 1948 [30]. Today, citronella is one of the most widely used natural repellents on the market, used at concentrations of 5-10%. This is lower than most other commercial repellents but higher concentrations can cause skin sensitivity. However, there are relatively few studies that have been carried out to determine the efficacy of essential oils from citronella as arthropod repellents. Citronella-based repellents only protect from host-seeking mosquitoes for about two hours although formulation of the repellent is very important [31, 32]. Initially, citronella, which contains citronellal, citronellol, geraniol, citral, α pinene, and limonene, is as effective dose for dose as DEET [33], but the oils rapidly evaporate causing loss of efficacy and leaving the user unprotected. However, by mixing the essential oil of Cymbopogon winterianus with a large molecule like vanillin (5%) protection time can be considerable prolonged by reducing the release rate of the volatile oil [34]. Recently, the use of nanotechnology has allowed slower release rates of oils to be achieved, thus prolonging protection time [35]. Encapsulated citronella oil nanoemulsion is prepared by high-pressure homogenization of 2.5% surfactant and 100% glycerol, to create stable droplets that increase the retention of the oil and slow down release. The release rate relates well to the protection time so that a decrease in release rate can prolong mosquito protection time [35]. Another means of prolonging the effect of natural repellents is microencapsulation using gelatin-arabic gum microcapsules, which maintained the repellency of citronella up to 30 days on treated fabric stored at room temperature (22°C) [36]. The use of these technologies to enhance the performance of natural repellents may revolutionize the repellent market and make plant oils a more viable option for use in long-lasting repellents. However, for the time-being travellers to disease endemic areas should not be recommended citronella-based repellents [32]. In contrast, for those communities where more efficacious alternatives are not available, or are prohibitively expensive, the use of citronella to prevent mosquito bites may provide important protection from disease vectors [17].

      The second way to use volatile plant repellents is to continuously evaporate them. Citronella and geraniol candles are widely sold as outdoor repellents, however field studies against mixed populations of nuisance mosquitoes show reductions in biting around 50%, although they do not provide significant protection against mosquito bites [3739].

      Neem

      Neem is widely advertised as a natural alternative to DEET [40], and it has been tested for repellency against range of arthropods of medical importance, with variable results (Table 1). Several field studies from India have shown very high efficacy of Neem-based preparations [4143], contrasting with findings of intermediate repellency by other researchers [44, 45]. However, these contrasting results may be due to differing methodologies, and the solvents used to carry the repellents. The EPA has not approved Neem for use as a topical insect repellent. It has a low dermal toxicity, but can cause skin irritation, such as dermatitis when used undiluted [46]. Due to the paucity of reliable studies, Neem oil is not recommended as an effective repellent for use by travellers to disease endemic areas [32], although it may confer some protection against nuisance biting mosquitoes.

      Natural oils and emulsions

      Several oils have shown repellency against mosquitoes. It is likely that they work in several ways 1) by reducing short range attractive cues i.e. kairomones, water vapour and temperature [4749]; 2) by reducing the evaporation and absorption of repellent actives due to the presence of long-chained fatty molecules [50]; 3) by containing fatty acids are known to be repellent to mosquitoes at high concentrations [51]. Bite Blocker, a commercial preparation containing glycerin, lecithin, vanillin, oils of coconut, geranium, and 2% soybean oil can achieve similar repellency to DEET, providing 7.2 hours mean protection time against a dengue vector and nuisance biting mosquitoes in one study [44], and protection for 1.5 hours, equivalent to that of low concentration DEET in a second study [52]. It would appear that the soybean oil in Bite Blocker helps only contributes to repellency as it is not repellent when evaluated on its own [53]. Soybean oil is not EPA registered, but it has low dermal toxicity, although no recommended maximum exposure or chronic exposure limits have been established [54]. Other plant-based oils that have shown some repellent efficacy are coconut oil, palm nut oils [55] and andiroba oil [56], although all of these three oils are far less effective than DEET, they may be useful as carriers for other repellent actives as they are cheap and contain unsaturated fatty acids and emulsifiers that improve repellent coverage and slow evaporation of volatile repellent molecules [50, 53, 57].

      Essential oils

      Essential oils distilled from members of the Lamiaceae (mint family that includes most culinary herbs), Poaceae (aromatic grasses) and Pinaceae (pine and cedar family) are commonly used as insect repellents throughout the globe (Table 1). Many members of these families are used in rural communities through burning or hanging them within homes [5862]. In Europe and North America there is a strong history of use of the oils dating back to Ancient times. Almost all of the plants used as repellents are also used for food flavouring or in the perfume industry, which may explain the association with these oils as safer natural alternatives to DEET despite many oils causing contact dermatitis (Table 2[63]). Many commercial repellents contain a number of plant essential oils either for fragrance or as repellents including peppermint, lemongrass, geraniol, pine oil, pennyroyal, cedar oil, thyme oil and patchouli. The most effective of these include thyme oil, geraniol, peppermint oil, cedar oil, patchouli and clove that have been found to repel malaria, filarial and yellow fever vectors for a period of 60-180 mins [6466]. Most of these essential oils are highly volatile and this contributes to their poor longevity as mosquito repellents. However, this problem can be addressed by using fixatives or careful formulation to improve their longevity. For example, oils from turmeric and hairy basil with addition of 5% vanillin repelled 3 species of mosquitoes under cage conditions for a period of 6-8 hours depending on the mosquito species [34]. Although essential oils are exempt from registration through the EPA, they can be irritating to the skin and their repellent effect is variable, dependent on formulation and concentration. Repellents containing only essential oils in the absence of an active ingredient such as DEET should not be recommended as repellents for use in disease endemic areas, and those containing high levels of essential oils could cause skin irritation, especially in the presence of sunlight.
      Table 2

      Some common ingredients in natural repellents that may be hazardous. Reproduced with permission from [63]

      Common Name

      Scientific Name

      Safe Concentration

      Hazard

      Anise

      Pimpinella anisum

      3.6%

      Based on 0.11% methyl eugenol; carcinogen

      Basil

      Ocimum sp

      0.07%

      Based on 6% methyl eugenol; carcinogen

      Bergamot

      Citrus aurantium bergamia

      0.4%

      Sensitising and phototoxic; skin irritant

      Cajeput

      Melaleuca alternifolia

      0.004%

      Based on 97% methyl eugenol; carcinogen

      Cedar

      Chamaecyparis nootkatensis

      1%

      Likely allergenic contaminants if nootkatone not 98% pure

      Cassia

      Cinnamonium cassia

      0.2% or 9%

      Sensitising skin irritant

      Citronella

      Cymbopogon nardus

      2%

      Safety is controversial; based on 0.2% methyl eugenol or 1.3% citral; sensitising skin irritant

      Citronella (Java)

      Cymbopogon winterianius

      2%

      Based on 0.2% methyl eugenol; carcinogen

      Citrus oils

      Citrus sp

      16-25%

      Based on 0.005%-0.0025% bergapten; phototoxic skin irritant

      Clove

      Syzyguim aromaticum

      0.5%

      Based on 92% eugenol; sensitising skin irritant

      Fever tea, lemon bush

      Lippia javanica

      2%

      Based on 5% citral in related species; sensitising skin irritant

      Geranium

      Pelargonium graveolens

      6%

      Based on 1.5% citral; sensitising skin irritant

      Ginger

      Zingiber sp

      12%

      Based on 0.8% citral; sensitising skin irritant

      Huon oil, Macquarie pine

      Langarostrobus franklini

      0.004%

      Based on 98% methyl eugenol; carcinogen

      Lemongrass

      Cymbopogon citratus

      0.1%

      Based on 90% citral; sensitising skin irritant

      Lime

      Citrus aurantifolia

      0.7%

      Phototoxic skin irritant

      Litsea

      Litsea cubeba

      0.1%

      Based on 78% citral; sensitising skin irritant

      Marigold

      Tagates minuta

      0.01%

      Phototoxic skin irritant

      Mexican tea, American wormseed

      Chenopodium ambrosioides

      Prohibited

      Toxic

      Mint

      Mentha piperata and spicata

      2%

      Based on 0.1% trans-2-hexenal; sensitising skin irritant

      Nutmeg

      Myristica fragrans

      0.4%

      Based on 1% methyl eugenol; carcinogen

      Palmarosa

      Cymbopogon martini

      16%

      Based on 1.2% farnesol; sensitizing skin irritant

      Pennyroyal

      Mentha pulegium or Hedeoma pulegioides

      Prohibited

      Toxic

      Pine

      Pinus sylvestris

      Prepare with antioxidants

      Oxidation creates phototoxic skin irritants

      Rosemary

      Rosemarinus officinalis

      36%

      Based on 0.011% methyl eugenol; carcinogen

      Rue

      Ruta chalepensis

      0.15%

      Based on presence of psoralenes; phototoxic skin irritant

      Thyme

      Thymus vulgaris

      2%

      Based on 0.1% trans-2-hexenal; sensitising skin irritant

      Violet

      Viola odorata

      2%

      Based on 0.1% trans-2-hexenal; sensitising skin irritant

      Ylang-ylang

      Canagium odoratum

      2%

      Based on 4% farnesol; sensitizing skin irritant

      Considerations for repellent testing methodology

      In a Pubmed search using the terms “plant” and “repellent” and “mosquito” in the past 5 years, 87 results were shown. These studies can be broken down into a series of categories: 1) standard ethnobotanical studies and evaluations of plants that are traditionally used to repel mosquitoes [17, 6770]; 2) standard dose response [33] laboratory evaluations of solvent extractions of plants without DEET positive controls [71]; 3) standard dose response [33] laboratory evaluations of solvent or extractions or essential oils of plants with DEET positive controls [72] coupled with GC-MS (coupled gas chromatography-mass spectrometry) [73]; 4) laboratory evaluations using time to first bite method [74] comparing the plant repellents to DEET [75] and in addition several of those studies also analysed the constituents of the oil through GC-MS [76, 77]. In addition there were a large number of studies that did not use the accepted standard methodology [78] (Table 3), and should be interpreted with caution. Only two studies considered safety [79] or adverse effects [80] and only one study considered randomization and blinding [52], and almost all repellent studies did not consider the number of human participants needed to minimize sampling error [81]. It is important for the future development of plant based repellents that the standard WHO methodology is followed [78], including a DEET control to allow simple comparison of multiple studies, and reporting of standard errors to understand the reliability of that repellent compound to provide the observed protection.
      Table 3

      Guidelines on repellent testing adapted from [78]

      WHOPES approved repellent testing methodology

      Laboratory Testing

      Use 20% deet in ethanol as a positive comparison

      Human subjects preferable to reflect the end user

      Before the test the test area of skin should be washed with unscented soap then rinsed with 70% ethanol / isopropyl alcohol

      Mosquitoes should be reared under standard 27 ± 2 C temperature, ≥80 ± 10% relative humidity, and a 12:12 (light:dark) photoperiod.

      Mosquitoes should be 3 to 5 days old, nulliparous females, starved for 12 hours preceding the test

      Tests should be conducted with three or more species

      40 x 40 x 40 cm cages with 50 – 100 mosquitoes for effective dose testing

      40 x 40 x 40 cm cages with 200 - 250 mosquitoes for complete protection time testing

      Control arms should be used to estimate mosquito readiness to feed

      Treatment arms should be offered to mosquitoes after avidity has been measured

      Field Testing

      Use 20% deet in ethanol as a positive comparison

      Human subjects preferable to reflect the end user

      Before the test the test area of skin should be washed with unscented soap then rinsed with 70% ethanol / isopropyl alcohol

      Volunteers should sit >20 metres apart

      Design should be completely randomised

      Trials should be conducted with medium biting pressures of representative vector species

      All participants should be recruited on informed consent from the local area and be provided with malaria prophylaxis

      In all testing monitoring of adverse effects should be carried out

      Some fallacies about plant based or natural repellents

      It is commonly assumed that plant-based repellents are safer than DEET because they are natural. However, some natural repellents are safer than others, and it cannot be assumed that natural equates to safe [18]. DEET has undergone stringent testing and has a good safety profile. An estimated 15 million people in the U.K., 78 million people in the U.S.A. [82], and 200 million people globally use DEET each year [83]. Provided that DEET is used safely, i.e. it is applied to the skin at the correct dose (such as that in a commercial preparation) and it is not swallowed or rubbed into the mucous membranes then it does not cause adverse effects [84]. DEET has been used since 1946 with a tiny number of reported adverse effects, many of which had a history of excessive or inappropriate use of repellent [85, 86]. Its toxicology has been more closely scrutinized than any other repellent, and it has been deemed safe for human use [82, 87], including use on children [88], pregnant women [89], and lactating women [84]. In contrast, plant-based repellents do not have this rigorously tested safety record, with most being deemed safe because they have simply been used for a long time [90]. However, many plant-based repellents contain compounds that should be used with caution (Table 1).

      It is also commonly stated that plant based repellents are better for the environment than synthetic molecules. While plant volatiles are naturally derived, distillation requires biomass energy, extraction commonly uses organic solvents that must be disposed of carefully, growing the plants uses agrichemicals, such as fertilizers and pesticides (unless sourced from a sustainable and organic source). However, if carefully practiced, cash cropping of plants used for repellents provides a vital source of income for small scale farmers in developing countries [91] and can have beneficial environmental impact when planted in intercropping systems to prevent soil erosions [92]. Therefore, it is important to carefully source of repellent plants to avoid pitfalls associated with unsustainable cropping practices. Another common misconception is that garlic is an effective repellent. It does have a moderate repellent effect when rubbed on the skin [93], although there are far more effective repellents available that also have a more pleasing odour. The consumption of garlic however, has not been shown to be effective at repelling mosquitoes.

      Promising developments in plant based repellents

      The field of plant-based repellents is moving forward as consumers demand means of protection from arthropod bites that are safe, pleasant to use and environmentally sustainable. Perhaps the most important consideration is improving the longevity of those repellents that are effective but volatile such as citronella. Several studies looked at improving formulations of plant oils to increase their longevity through development of nanoemulsions [35, 94], improved formulations and fixatives [9597]; while alternate uses such as spatial activity [98102] and excitorepellency [103, 104] have also been investigated. There has been a single clinical study of PMD to lower malaria incidence [26]. This is an exciting discovery since PMD may be recovered from distillation of leaves of E. citroidora or chemical modification of citronellal [105]– available from plants of the genus Cymbopogon. These plants are already commercially cropped in malaria endemic countries including South America, especially Brazil (6 million trees), southern China, India, Sri Lanka, Congo (Zaire), Kenya and most countries in southern Africa, where it is grown for essential oil production and timber [106]. Local production of insect repellent would remove the high cost of importation in developing countries.

      New developments have also been seen in understanding the function of plant-based repellents in insects. Several studies have investigated the behavioural mode of action of repellents through structure-activity studies of contact versus spatial repellency [107], or olfactometry that demonstrated that DEET inhibited mosquito response to human odour whereas Ocimum forskolei repels but does not inhibit response to human odour [108]. A further study demonstrates that citronellal directly activates cation channels [10], which is similar to the excitorepellent effect of pyrethrin – another plant based terpine [109], but contrasts with the inhibition effect of DEET [3].

      The field of repellent development from plants is extremely fertile due to wealth of insecticidal compounds found in plants as defences against insects [2]. The modern pyrethroids that are the mainstay of the current malaria elimination program that is making excellent progress [110], are synthetic analogues based on the chemical structure of pyrethrins, discovered in the pyrethrum daisy, Tanacetum cinerariifolium from the Dalmation region and Tanacetum coccineum of Persian origin. The insecticidal component comprising six esters (pyrethrins) is found in tiny oil-containing glands on the surface of the seed case in the flower head to protect the seed from insect attack. Pyrethrins are highly effective insecticides, that are relatively harmless to mammals [111], although it must be emphasised that many other plant produce compounds that are highly toxic to mammals and / or irritating to the skin, and natural does not equate to safe. In the past few years, a plant derived repellent, PMD has been proven to be suitably efficacious and safe to compete with DEET in the field of disease prevention, and repellents have been recognised by WHO as a useful disease prevention tool to complement insecticide-based means of vector control. The field of plant-based repellent evaluation and development had become far more rigorous in recent years and developments in methods of dispensing plant-based volatiles means that extension in the duration of repellency and consequent efficacy of plant-based repellents will be possible in future.

      Declarations

      Acknowledgements and funding

      Authors receive salary support from Bill and Melinda Gates Foundation 51431. We would like to thank Coronel Mustapha Debboun for permission to reproduce Table 2 and the two anonymous reviewers who greatly improved the manuscript through their comments and suggestions.

      This article has been published as part of Malaria Journal Volume 10 Supplement 1, 2011: Natural products for the control of malaria. The full contents of the supplement are available online at http://​www.​malariajournal.​com/​supplements/​10/​S1.

      Authors’ Affiliations

      (1)
      Disease Control Department, London School of Hygiene and Tropical Medicine
      (2)
      Biomedical and Environmental Thematic Group, Ifakara Health Institute

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