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Juliet Rice
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1 Quassia Vinegar

They reproduce quickly by laying eggs, known as nits, that females lay on the hair shaft, close to the scalp.

2 INTRODUCTION WHAT ARE HEADLICE? Headlice, Pediculus humanus capitis, are small parasitic insects living exclusively in the human scalp (they do not infest or live on domestic animals), and feeding on the blood they suck. They reproduce quickly by laying eggs, known as nits, that females lay on the hair shaft, close to the scalp. Nits are yellowish-white and have a chitin envelope, which protects them from external hazards and attaches them to the hair shaft. Their life cycle is approximately days. The eggs, or nits attached to the hair shafts, hatch in 7-10 days and the immature form (nymph) smaller than the adult emerges. It reaches the adult size in about 2 weeks and lives as an adult for about 28 days. The main symptom of the headlice parasitosis or infestation, known as Pediculosis, is an itchy scalp resulting from the irritation produced by the insect s saliva deposited in the bite wounds. Although it can be found everywhere on the head, it is usually located behind the ears, on the nape of the neck and the top of the head. In some cases, it can also affect eyebrows, eyelashes and beard. Two transmission ways exist: a direct way, by a head contacting another head, and an indirect way, by sharing carrier objects such as hairbrushes, combs, hair ribbons, hairpins, hats and scarves. HEADLICE INFESTATION: PEDICULOSIS Infestation or pediculosis is rather frequent among school-aged children, mainly those between 4 and 6 years old, the incidence being smaller between 6 and 10 years old children. At present, adults are not prone to infestation. V04-01/ ,82390,

The only possible strategy is to do an early diagnosis and a quick intervention with an easily applicable, efficient, minimally toxic, and easily removable product.

3 It is an endemic problem, which seems to have no solution. It does not depend on the geography, the hygiene habits, the social class, illnesses, etc. Short hair or daily shampooing does not prevent infestation. The only possible strategy is to do an early diagnosis and a quick intervention with an easily applicable, efficient, minimally toxic, and easily removable product. CURRENT ANTI-HEADLICE TREATMENTS There are several types of treatments in the market. 1. CHEMICAL TREATMENTS These conventional treatments contain different insecticide substances: Pyrethrin and permethrin Malathion Lindane Piperonyl All of these pediculicide agents act efficiently against the adult headlice and less efficiently against nits. However, some of them are neurotoxic. Since they are not selective, they act on nervous system of insects as well as on that of higher life forms. Other elements strengthen their toxic effects: They are only mildly effective in eliminating eggs before they hatch, thus requiring repeated treatment applications. They must be applied on a rather permeable area (the scalp) in a population mainly composed by children, whose less developed immune system and detoxification processes makes them most sensitive to insecticides. V04-01/ ,82390,

4 The continued use of these products induces resistance. Consequently, higher concentrations or longer treatments are required, thus increasing the risk for adverse effects. Lindane, an organochlorine pesticide, is the most toxic of them and it is currently forbidden, or in process of becoming forbidden, in many countries. The main absorption ways are the oral, dermal and the respiratory system. Because of its high liposolubility, it is stored in the adipose tissue and other fat-rich tissues such as liver, kidneys and nervous system. Symptoms resulting from the exposition to lindane resemble DDT intoxication. Malathion is an organophosphorus insecticide, very toxic by ingestion, irritant by skin or eyes contact and probably mutagen. Pyrethrins are natural insecticides produced by the species Chrysanthemum cinaerefolium obtained from the extract of pellitory, used as a domestic insecticide because of their high insectkilling power and low toxicity for mammals. However, they undergo photolysis when exposed to sunlight and air and degrade yielding inactive products. To cope with these stability problems, derived substances such as permethrin were synthesized. Permethrin is recommended as a first-choice treatment, although recent studies report decreased sensitivity and increased resistance of lice to these products. Pyrethrins and permethrins are considered the safest products because of the low absorption occurring at the concentration levels present in most of commercial formulas. 2. Traditional treatments One of the most traditional and homemade methods is to use vinegar because of its efficacy against headlice as well as against nits. It must be directly applied on the dry hair and left to act for a couple of hours. Other home remedies usually employed are olive oil and mayonnaise, which act by immobilizing the parasites. Hair conditioning products or Vaseline can also be used. Hair must be washed with a standard shampoo; a large amount of a hair-conditioning product or of Vaseline must be subsequently applied and only removed after the termination of the treatment. That will immobilize headlice and prevent them from attaching to the hair shafts. V04-01/ ,82390,

5 These homemade remedies are effective. However, some of them are not comfortable to use, others are difficult to remove and no program or treatment schedule neither efficacy test exists for them. 3. ALTERNATIVE TREATMENTS: QUASSIA VINEGAR The problems and disadvantages of chemical treatments (especially their toxic effects) and homemade remedies, have promoted the research on new generations of alternative products with pediculicide activity. Alternative treatments act only against the harmful species (insect) without affecting other organisms, including humans. For that purpose, products altering physiological processes exclusive of insects are pursued. Examples of alternative products are: Inhibitors of the chitin synthesis Products interfering with the synthesis of chitin during the larval change. As new chitin cannot be synthesized, the cuticle cannot resist the turgor pressure. The resulting injuries eventually produce death. Adult individuals are not affected, but larvae born from treated females either do not hatch or else, they are affected. The toxicity of these compounds is very low and does not affect higher animals. Repellent products QUASSIA VINEGAR, Anti-headlice Complex It combines both above described actions due to the presence of Quassia Amara, a natural repellent and inhibitor of the chitin synthesis, and the presence of Vinegar, which prevents nits from attaching to the hair shafts by dissolving their chitin envelopes, additionally facilitating the removal of the already attached ones. Its characteristic chemical compounds, the quassinoids, act by inhibiting the insects growth. Quassin, another identified substance, acts as an insecticide because of its larvicide effects, which result from inhibition of the tyrosinase activity in the insect larvae. Since that enzyme is directly involved in the cuticle sclerotization process, this active compound would prevent the insect cuticle formation. V04-01/ ,82390,

The action mechanism described for Quassia Amara and the actions of Vinegar result in a pediculosis treatment with much weaker toxic effects than those of chemical agents.

6 The action mechanism described for Quassia Amara and the actions of Vinegar result in a pediculosis treatment with much weaker toxic effects than those of chemical agents. BOTANICS AND CHEMICAL COMPOSITION Quassia amara L. Common names: amargo, bitter ash, bitterwood, quassia, jamaica bark, jamaica quassia, quassiawood, surinam wood, surinam quassia. Quassia amara, also known as Surinam Quassia, is a bush belonging to the family of Simaroubaceae, native from the tropical America, cultivated in many Amazon countries, whose wood is never attacked by insects. The trunk, roots, branches, leaves and fruits have a bitter flavour, remarkably strong in the log or wood, which contains several substances 50-fold bitter than quinine. Therefore, the log or wood is the main part used in the traditional medicine, although the leaves can also be used. These bitter substances, characteristic of the Quassia log chemical composition, are the quassinoids. They were isolated by the end of the XIX century, although the structure of quassin was not described until 125 years later, in 1961, because of its high complexity and the difficulties to isolate it in the pure state, separated from other active compounds. Quassinoids are modified triterpenes. Two major quassinoids, quassin and neoquassin, and other minor quassinoids such as parain, nigakilactone A, 18- hydroxyquassin, quassialactol, isoquassin, quassol and quassinol have been identified. Quassinoid glycosides are also present. O M e H O O H H H O H R 1 R 2 11-dihydro-12-norneoquassin (R 1 =H, R 2 = OH) Paraine( R 1, R 2 = O) Figure 1.Quassinoids O N 6 5 O H N O M e 4-methoxy-5-hydroxy canthin-6-one 4 3 V04-01/ ,82390,

PEDICULICIDE ACTIVITY: TREATMENT AND PREVENTION Quassia amara is a plant widely employed by the natives from South America and Costa Rica, as a tonic against malaria ulcers and to combat

7 PEDICULICIDE ACTIVITY: TREATMENT AND PREVENTION Quassia amara is a plant widely employed by the natives from South America and Costa Rica, as a tonic against malaria ulcers and to combat colic-associated stomachache. It is often prepared as a cold-water maceration or as an infusion and consumed as a bitter, appetite-stimulating tonic. It is a major medicine for Costa Rica natives. The name of this plant derives from a Surinam native, named Quassia, who became famous by treating the malignant fever by the middle of century XVIII. The properties of this plant were studied in Stockholm by 1756; the first reference to it appeared by 1764 as Lignum quassie; it became then a very popular febrifuge, anti-dysentery and tonic medicine. The medicinal use of Quassia is widespread throughout Latin America, for the treatment of fever, digestive problems, or intestinal parasites. Although the earliest use antecedents, from century XVIII, were as a medicinal herb, Quassia amara extracts were used in Europe and the United States to manufacture natural insecticides. Such a use is well documented between 1884 and 1940, when synthetic pesticides emerged and the use of natural products decreased. Several species belonging to the family of Simaroubaceae, traditionally called marupá in Brazil, are well-known medicinal, insecticide and antihelmintic agents. The earliest scientific research on the pediculicide activity of Quassia dates from 1978, when Jensen et al. studied the effects of Quassia extract on 454 headlice infested patients. After a week, nits were only found on three individuals, and that finding could be explained by re-infestation. That study confirmed the high Quassia effectivity as an alternative to the chemical products (chlorophenothane) that patients had previously used. V04-01/ ,82390,

A subsequent double blind study including a placebo was carried out to evaluate the preventive and therapeutic action of Quassia on human pediculosis.

8 A subsequent double blind study including a placebo was carried out to evaluate the preventive and therapeutic action of Quassia on human pediculosis. The results confirmed the effectivity of Quassia for treatment, as well as for prevention of the headlice infestation (Ninci, 1991). Several studies have evaluated the insecticide activity of Quassia. Evans et al (1991) observed that the water extract of Quassia had anti-larval activity on Culex quinquefasciatus. The results showed that Quassia could be used as an insecticide, without the toxic effects of the synthetic insecticides. More recently, the above cited research group studied the effects of Quassia extract on the metabolism of the aromatic amino acids phenylalanine and tyrosine and their derivates L-dopa and adrenaline in the mosquito larvae (Culex quinquefasciatus). Tyrosine concentration was higher than that of phenylalanine throughout these mosquitoes life cycle. Phenylalanine, L-dopa and adrenaline levels did not vary significantly during the larval stage. However, their concentrations in the adult stage were higher than those in the development phase. After the addition of Quassia extract (5ppm) and a 6 hours incubation period, double levels of phenylalanine, tyrosine and L-dopa were observed, without any modification in the levels of adrenaline. The fact that this extract does not affect the enzyme monoamine oxidase can explain the finding that, adrenaline concentration remained the same. However, the enzyme tyrosinase is known to be very active during the development stages and to play an important role in the cuticle sclerotization. This enzyme was found to be highly sensitive to Quassia, which completely inhibited it even at low concentrations. These findings can explain the action mechanism of the Quassia extract as an anti-larval agent (Evans et al., 1992). In order to evaluate the anti-malaria activity of Quassia, a preliminary in vivo study was carried out in mice, with Plasmodium berghei berghei and extracts of Quassia amara and Quassia undulata. The results showed that the Q.amara and Q.undulata extracts had a strong anti-malaria activity, that of Q.amara being the strongest (Ajaiyeoba et al., 1999). Quassia is a bitter tonic, which stimulates the digestion related organs, thus augmenting the secretions of saliva and bile. It is also a mild activator of the blood circulation. Studies using white rats and mice support the traditional use of this plant as a digestive agent. Additionally, these V04-01/ ,82390,

studies show that the oral administration of Quassia-wood water extract has no direct effect on the nervous system or on other systems in the organism (García M, et al. 1997).

9 studies show that the oral administration of Quassia-wood water extract has no direct effect on the nervous system or on other systems in the organism (García M, et al. 1997). Vinegar, liquid obtained from the acetic fermentation of fruit juices and therefore acid in nature, is a very used traditional treatment for prevention, as well as for elimination, of the headlice infestation. A combination of 1 part of vinegar plus 2 parts of water is often used to rinse the hair. The pediculicide efficacy of this product results from its ability to soften chitin, thus favouring the elimination of nits from the hair shaft. The abundant information available on headlice infestation still suggests applying vinegar when rinsing the hair for prevention, as well as for treatment of the infestation. According to Giardelli et al. 2001, a 10 minutes exposition to Vinegar 100% produces a pediculostatic effect, namely an inhibition of the headlice movements. However, strong local irritation effects were also observed. Vinegar was demonstrated to have a repellent effect for approximately 60 minutes after the application of the studied concentrations (25% and 50%). SAFETY AND TOXICITY Bibliographic data about the toxicity of this plant and its active compounds are available. One of its main active compounds is quassin, registered in Europe and the USA as an alimentary additive, specifically as an aroma additive. In July 2002 the Scientific Committee on Food of the European Commission issued a report on the safety of quassin. That report compiled all of the data on its toxicity available to that date. In Europe, it is allowed to include up to a maximum quassin concentration of 5 mg/kg in food and beverage, 10 mg/kg in pastries and 50 mg/kg in alcoholic drinks (88/388/EEC). A re-evaluation of these concentration limits was requested because some studies had shown toxic effects of quassin and Q.amara extract on the reproductive capacity of rats. No sign of toxicity was observed when doses up to 1000 mg/kg Quassia water extract were orally administered to rats and mice. In the USA, Quassia amara has been included into the list of safe ingredients for food (21CFR ). V04-01/ ,82390,

10 IN-VIVO EFFICACY TEST The effects of QUASSIA VINEGAR and Andiroba oil were evaluated for the treatment and prevention of human pediculosis. An in-vivo study on the scalp of volunteers infested with Pediculus humanus was carried out. A dermatologist visually evaluated the action of the tested products. TESTED PRODUCTS QUASSIA VINEGAR combines the action of Quassia amara as a natural repellent of Pediculus capitis and of Vinegar, which prevents nits from attaching to the hair shaft by dissolving their chitin envelopes. Andiroba oil was included in this study because of its general insect repellent properties. To perform the present study 3% QUASSIA VINEGAR was added to three different formulations for hair: hair lotion, shampoo and hair conditioner. Andiroba oil 1.3% was added only to the lotion. METHOD Efficacy was evaluated on 21 male and female volunteers undergoing the following treatment for 14 days: every evening, the lotion was applied on the whole scalp; on the following day, their hair was shampooed and the hair conditioner was subsequently applied. After the treatment period, the volunteers went back to their usual shampoo for an additional 14 days period. This study allowed for a follow-up evaluation of the prevention effects since these volunteers were constantly in contact with untreated pediculosis-infested subjects. Controls were carried out every 7 days, during the treatment period and the following 14 additional days until a total of 28 observation days. Pediculosis cases were classified according to the infestation degree, as shown in the following table. V04-01/ ,82390,

11 Category No infestation Mild infestation Moderate infestation Severe infestation Very severe infestation Quantification No pediculosis Less than 10 nits Between 11 and 50 nits More than 51 nits Uncountable nits Table1 RESULTS Dermatological control (nº and % volunteers per group) Category D1 D7 D14 D21 D28 No pediculosis 0 (0%) 5 (24%) 12 (57%) 20 (95%) 21 (100%) < 10 nits 5 (24%) 8 (38%) 9 (43%) 1 (5%) 0 (0%) nits 8 (38%) 4 (19%) 0 (0%) 0 (0%) 0 (0%) > 51 nits 3 (14%) 3 (14%) 0 (0%) 0 (0%) 0 (0%) Uncountable 5 (24%) 1 (5%) 0 (0%) 0 (0%) 0 (0%) Table 2 The initial pediculosis infestation, observed in the first control of the study, progressively diminished in all of the treated volunteers, eventually disappearing by the end of the experimental period (D28). The results also showed that no re-infestation took place, even when the volunteers were in contact with infested subjects throughout the study period. The following graphic shows the efficacy of the tested products on pediculosis as a whole, namely when all of the categories were included in a group, as a function of time. V04-01/ ,82390,

12 % volunteers with pediculosis Efficacy against pediculosis D1 D7 D14 D21 D28 Dermatologic control Figure 2. Efficacy against pediculosis CONCLUSIONS The present study demonstrated that hair treatment using QUASSIA VINEGAR and Andiroba oil was effective already from the first treatment week and completely eliminated nits in all of the volunteers. Additionally, it was demonstrated that the tested products act as preventive agents since no reinfestation was recorded under the test conditions. COSMETIC PROPERTIES AND USAGE DOSE 100% botanical Natural origin, from renewable source Safe / Non-toxic 100 % Proven anti-headlice efficacy Less resistance Hair conditioner V04-01/ ,82390,

13 Usage dose: Pediculicide treatment: 3.0% Preventive treatment: 3.0% BIBLIOGRAPHY A.O.A.C Official Methods of Analysis. 11ª ed., pag. 520, Ajaiyeoba EO, et al. In vivo antimalarial activities of Quassia amara and Quassia undulata plant extracts in mice. Journal of Ethnopharmacology 1999; 67: (ref. 5748). Andiroba Oil: Repellence Test in Flies. Unpublished report of efficacy test. Badilla B, et al. Actividad gastrointestinal del extracto acuoso bruto de Quassia amara (Simarubaceae). Journal of Tropical Biology 1998; 46 (3): 1-8 (ref. 5747). Barbetti P, et al. Indole alkaloids from Quassia amara. Planta Medica 1987; 53 (3): (ref. 5741). Barbetti P, et al. New canthin-6-one alkaloids from Quassia amara. Planta Medica 1990; 56: (ref. 5742). Barbetti P, et al. Quassinoids from Quassia amara. Phytochemistry 1993; 32 (4): (ref. 5740). Belitz HD, Grosch W. Química de los alimentos. Zaragoza: Acribia, 1997; CADIME. Pediculosis: Tratamiento. Bol Ter Andal 2002; 18 (3): 9-13 (ref. 5757). Chapter Three. Phytochemistry study on Quassia amara, pp 1-27 (ref. 5753) Dincin D. Hierbas para la belleza natural. Palma de Mallorca: José J. De Olañeta, 1986; Dou J, et al. A new quassinoid from crude quassin-extract of Quassia amara. International Journal of Pharmacognosy 1996; 34 (5): (ref. 5739). V04-01/ ,82390,

14 Evans DA, et al. Effecto of quassin on the metabolism of catecholamines in different life cycle stages of Culex quinquefasciatus. Indian Journal of Biochemistry Biophysics 1992; 29: (ref. 5750). Evans DA, et al. Larvicidal efficacy of quassin against Culex quinquefasciatus. Indian J Med Res 1991; 93: (ref. 5749). García M, et al. Actividad farmacológica del extracto acuoso de la madera de Quassia amara (Simarubaceae) en ratas y ratones albinos. Rev Biol Trop 1997; 44-45:47-50 (ref. 5744). Garcia M, Quintero R, López-Munguía A. Biotecnología alimentaria. México D.F.: Limusa, 1999; Giardelli M, et al. Pediculosis y escabiosis. Arch. Argen. Pediatr 2001; 99(1): (ref. 5754). Grandolini G, et al. A new neoquassin derivative from Quassia amara. Phytochemistry 1987; 26 (11): (ref. 5738). Instituto de Racionalización del Trabajo. U.N.E Jensen O, et al. Pediculosis capitis treated with Quassia tincture. Acta Dermatovener (Stockholm) 1978; 58: (ref. 5751). Kligman AM. Topical treatments for photoaged skin. Postgraduate Medicine 1997; 102 (2): Manrique T, et al. Estudio comparativo entre una fórmula de aceite mineral vs. un champú pediculicida para el tratamiento de la infestación por piojos. Archivos Venezolanos de Puericultura y Pediatría 2000; 63 (1): (ref. 5745). Nakatani, N. Phenolic compounds in food and their effects on health II. American Chemical Society, 1992;72-86 (ref. 709). Ninci M. Profilaxis y tratamiento de pediculosis con cuasia amarga. Revista de la Facultad de Ciencias Médicas de la Universidad de Cordoba 1991; 49 (2): (ref. 5752). Njar VCO, et al. 2-Methoxycanthin-6-one: a new alkaloid from the stem wood of Quassia amara. Planta Medica 1993; 59: (ref. 5743). V04-01/ ,82390,

15 Pérez-Cotapos ML, et al. Pediculosis y escabiosis. Boletín Escuela de Medicina. Pontificia Universidad Católica de Chile 1996; 25: (ref. 5755). Robins RJ, et al. High-performance liquid chromatographic methods for the analysis and purification of quassinoids from Quassia amara L. Journal of Chromatography 1984; 283: (ref. 5746). Salunkhe DK, Kadam SS. Handbook of fruit science and technology: production, composition, storage and processing. New York: Marcel Dekker, 1995; , , Scholz D, Brooks GJ, Parish DF, Burmeister F. Fruit acid extracts, a fresh approach to skin renewal. International Journal of Cosmetic Sience 1994; 16: (ref. 697). Villalobos R, et al. Desarrollo de criterios silviculturales para el manejo de Quassia amara, un producto no maderable del bosque tropical. Simposio Internacional Posibilidades de Manejo Forestal Sostenible en America Tropical.Santa Cruz de la Sierra, Bolivia, de julio 1997 (ref. 5737). Vollmer G, Josst G, Schenker D, Sturn W, Vreden N. Elementos de bromatología descriptiva. Zaragoza: Acribia, 1995; Wang X. A theory for the mechanism of actionof the -hydroxy acids applied to the skin. Medica Hypotheses, 1999; 53 (5): (ref. 4261). Witkowski JA, et al. Pediculosis and resistance: the perennial problem. Clinics in Dermatology 2002; 20: (ref. 5756). V04-01/ ,82390,

Fruit Vinegar Exclusive N.A. Distributor CENTERCHEM, INC. NORWALK, CT Ph: Fax:

Fruit Vinegar Exclusive N.A. Distributor CENTERCHEM, INC. NORWALK, CT Ph: Fax: Fruit Vinegar DESCRIPTION FRUIT VINEGAR is a product obtained from the maceration of Apple Vinegar with the following fruit, APRICOT, APPLE and PEACH BOTANY AND CHEMISTRY APRICOT Prunus armeniaca L. This