Final Report on the Safety Assessment of AloeAndongensis Extract, Aloe Ando...

International Journal of Toxicology, 26(Suppl. 2):150, 2007Copyright c American College of ToxicologyISSN: 1091-5818 print / 1092-874X onlineDOI: 10.1080/10915810701351186

Final Report on the Safety Assessment of Aloe AndongensisExtract, Aloe Andongensis Leaf Juice, Aloe Arborescens LeafExtract, Aloe Arborescens Leaf Juice, Aloe Arborescens LeafProtoplasts, Aloe Barbadensis Flower Extract, AloeBarbadensis Leaf, Aloe Barbadensis Leaf Extract, AloeBarbadensis Leaf Juice, Aloe Barbadensis LeafPolysaccharides, Aloe Barbadensis Leaf Water, Aloe FeroxLeaf Extract, Aloe Ferox Leaf Juice, and Aloe Ferox LeafJuice Extract1

Plant materials derived from the Aloe plant are used as cosmeticingredients, including Aloe Andongensis Extract, Aloe Andongen-sis Leaf Juice, Aloe Arborescens Leaf Extract, Aloe ArborescensLeaf Juice, Aloe Arborescens Leaf Protoplasts, Aloe BarbadensisFlower Extract, Aloe Barbadensis Leaf, Aloe Barbadensis Leaf Ex-tract, Aloe Barbadensis Leaf Juice, Aloe Barbadensis Leaf Polysac-charides, Aloe Barbadensis Leaf Water, Aloe Ferox Leaf Extract,Aloe Ferox Leaf Juice, and Aloe Ferox Leaf Juice Extract. Theseingredients function primarily as skin-conditioning agents and areincluded in cosmetics only at low concentrations. The Aloe leaf con-sists of the pericyclic cells, found just below the plants skin, and theinner central area of the leaf, i.e., the gel, which is used for cosmeticproducts. The pericyclic cells produce a bitter, yellow latex con-taining a number of anthraquinones, phototoxic compounds thatare also gastrointestinal irritants responsible for cathartic effects.The gel contains polysaccharides, which can be acetylated, par-tially acetylated, or not acetylated. An industry established limitfor anthraquinones in aloe-derived material for nonmedicinal useis 50 ppm or lower. Aloe-derived ingredients are used in a wide vari-ety of cosmetic product types at concentrations of raw material thatare 0.1% or less, although can be as high as 20%. The concentra-tion of Aloe in the raw material also may vary from 100% to a lowof 0.0005%. Oral administration of various anthraquinone compo-nents results in a rise in their blood concentrations, wide systemicdistribution, accumulation in the liver and kidneys, and excretionin urine and feces; polysaccharide components are distributed sys-temically and metabolized into smaller molecules. aloe-derived ma-terial has fungicidal, antimicrobial, and antiviral activities, and hasbeen effective in wound healing and infection treatment in animals.Aloe barbadensis (also known as Aloe vera)derived ingredientswere not toxic in acute oral studies using mice and rats. In par-enteral studies, the LD50 using mice was >200 mg/kg, rats was

Received 30 November 2006; accepted 5 March 2007.1Reviewed by the Cosmetic Ingredient Review Expert Panel.

Address correspondence to F. Alan Andersen, Cosmetic Ingredient Re-view, 1101 17th Street, NW, Suite 412, Washington, DC 20036, USA.

>50 mg/kg, and using dogs was >50 mg/kg. In intravenous stud-ies the LD50 using mice was>80 mg/kg, rats was>15 mg/kg, anddogs was>10 mg/kg. The 14-day no observed effect level (NOEL)for the Aloe polysaccharide, acemannan, in the diet of Sprague-Dawley rats, was 50,000 ppm or 4.1 to 4.6 g/kg day1. In a 3-monthstudy using mice, Aloe vera (extracted in ethanol) given orally indrinking water at 100 mg/kg produced reproductive toxicity, in-flammation, and mortality above that seen in control animals. Aloevera extracted in methanol and given to mice at 100 mg/kg in drink-ing water for 3 months caused significant sperm damage comparedto controls. Aloe barbadensis extracted with water and given topregnant Charles Foster albino rats on gestational days (GDs) 0through 9 was an abortifacient and produced skeletal abnormal-ities. Both negative and positive results were found in bacterialand mammalian cell genotoxicity assays using Aloe barbadensisderived material, Aloe Feroxderived material, and various an-thraquinones derived from Aloe. Aloin (an anthraquinone) did notproduce tumors when included in the feed of mice for 20 weeks, nordid aloin increase the incidence of colorectal tumors induced with1,2-dimethylhydrazine. Aloe-emodin (an anthraquinone) given tomice in which tumor cells had been injected inhibited growth ofmalignant tumors. Other animal data also suggest that compo-nents of Aloe inhibit tumor growth and improve survival. Vari-ous in vitro assays also demonstrated anticarcinogenic activity ofaloe-emodin. Diarrhea was the only adverse effect of note with theuse of Aloe-derived ingredients to treat asthma, ischemic heart dis-ease, diabetes, ulcers, skin disease, and cancer. Case reports includeacute eczema, contact urticaria, and dermatitis in individuals whoapplied Aloe-derived ingredients topically. The Cosmetic Ingredi-ent Review Expert Panel concluded that anthraquinone levels inthe several Aloe Barbadensis extracts are well understood and canconform to the industry-established level of 50 ppm. Although thephototoxicity anthraquinone components of Aloe plants have beendemonstrated, several clinical studies of preparations derived fromAloe barbadensis plants demonstrated no phototoxicity, confirm-ing that the concentrations of anthraquinones in such prepara-tions are too low to induce phototoxicity. The characterization ofaloe-derived ingredients from other species is not clear. In the ab-sence of well-characterized derivatives, biological studies of thesematerials are considered necessary. The studies needed are 28-day

1

2 COSMETIC INGREDIENT REVIEW

dermal toxicity studies on Aloe Andongensis Extract, Aloe Andon-gensis Leaf Juice, Aloe Arborescens Leaf Extract, Aloe ArborescensLeaf Juice, Aloe Ferox Leaf Extract, Aloe Ferox Leaf Juice, andAloe Ferox Leaf Juice (ingredients should be tested at current useconcentrations). In Aloe-derived ingredients used in cosmetics, re-gardless of species, anthraquinone levels should not exceed 50 ppm.The Cosmetic Ingredient Review Expert Panel advised the industrythat the total polychlorobiphenyl (PCB)/pesticide contamination ofany plant-derived cosmetic ingredient should be limited to not morethan 40 ppm, with not more than 10 ppm for any specific residueand that limits were appropriate for the following impurities: ar-senic (3 mg/kg maximum), heavy metals (20 mg/kg maximum), andlead (5 mg/kg maximum).

INTRODUCTIONAloe Andongensis Extract, Aloe Andongensis Leaf Juice,

Aloe Arborescens Leaf Extract, Aloe Arborescens Leaf Juice,Aloe Arborescens Leaf Protoplasts, Aloe Barbadensis FlowerExtract, Aloe Barbadensis Leaf, Aloe Barbadensis Leaf Extract,Aloe Barbadensis Leaf Juice, Aloe Barbadensis Leaf Polysac-charides, Aloe Barbadensis Leaf Water,Aloe Ferox Leaf Extract,Aloe Ferox Leaf Juice, and Aloe Ferox Leaf Juice Extract arecosmetic ingredients derived from Aloe. This review presentsinformation relevant to the safety of these Aloe-derived ingre-dients as considered by the Cosmetic Ingredient Review (CIR)Expert Panel.

The general Aloe CAS number is 8001-97-6. Aloe barbaden-sis extract has the CAS number 85507-69-3. As described in theInternational Cosmetic Ingredient Dictionary and Handbook(Gottschalck and McEwen 2004), these ingredients function pri-marily as skin-conditioning agents or have no specific functionlisted. In most cases, the definitions of these ingredients are tau-tologies, e.g., Aloe Barbadensis Flower Extract is an extract ofthe flowers of Aloe barbadensis.

CHEMISTRY

DefinitionPlant material derived from the aloe plant is characterized

by its source (e.g., what part of the plant), the species of plant,the physical description of the material, and by the constituentsfound in the material.

The definitions of each ingredient, according to the In-ternational Cosmetic Ingredient Dictionary and Handbook(Gottschalck and McEwen 2004), are:

Aloe Andongensis Extract is the extract of the leaves of Aloeandongensis.

Aloe Andongensis Leaf Juice is the liquid expressed from theleaves of Aloe andongensis.

Aloe Arborescens Leaf Extract is the extract of the leaves ofAloe arborescens.

Aloe Arborescens Leaf Juice is the juice expressed from theleaves of Aloe arborescens.

Aloe Arborescens Leaf Protoplasts are the protoplasts ob-tained form the leaves of Aloe arborescens.

Aloe Barbadensis Flower Extract is the extract of the flowersof Aloe barbadensis.

Aloe Barbadensis Leaf is a plant material derived from theleaves of Aloe barbadensis.

Aloe Barbadensis Leaf Extract is an extract of the leaves ofAloe barbadensis.

Aloe Barbadensis Leaf Juice is the juice expressed from theleaves of Aloe barbadensis.

Aloe Barbadensis Leaf Polysaccharides is the polysaccharidefraction isolated from the leaf of Aloe barbadensis.

Aloe Barbadensis Leaf Water is an aqueous solution of theodoriferous principles distilled from the leaves of Aloe bar-badensis.

Aloe Ferox Leaf Extract is an extract of the leaves of Aloeferox.

Aloe Ferox Leaf Juice is the juice expressed from the leaves ofAloe ferox.

Aloe Ferox Leaf Juice Extract is an extract of the juice of theleaf of Aloe ferox.

Table 1 gives a list of synonyms for these ingredients as givenin the International Cosmetic Ingredient Dictionary and Hand-book (Gottschalck and McEwen 2004). Aloe vera is a frequentlyused synonym for Aloe barbadensis.

Composition and Physical and Chemical PropertiesGjerstad (1969) described Aloe as the dried juice of the lower

portion of the leaves of any three geographical varieties of theAloe genus. The juice appears blackish-brown, opaque, andsmooth.

Ghannam et al. (1986) described the solid residue of Aloebarbadensis (native to Mediterranean countries and the SaudiArabian peninsula) obtained by evaporating the sap drained fromthe cut leaves. This drained latex solidifies and turns brown onexposure to air. It contains anthraquinone glycosides, barbaloin,and -barbaloin, the hydrolysis of which yields aloe-emodin andd-arabinose.

Natow (1986) stated that Aloe is the name given to the genusthat includes more than 300 plants, grown all over the world.This report considers ingredients from four species: andongen-sis, arborescens, barbadensis, and ferox.

Klein and Penneys (1988), Briggs (1995), and the M.D. An-derson Cancer Center (2003) described the leaf of Aloe plantsas consisting of two main parts. One part, the pericyclic cells, isfound just below the plants skin. The pericyclic cells produce abitter, yellow latex known as Aloe juice, or latex. When this juicedries it forms a dark brown solid material. The latex containsthe anthraquinone, emodin, a gastrointestinal irritant responsi-ble for cathartic effects. The second part, the inner central areaof the leaf, contains the thin walled parenchymal cells that pro-duce the clear slightly viscous (mucilaginous) fluid known asAloe gel or inner gel. This gel contains the polysaccharides.There are generally more than one polysaccharide found in the

ALOE 3

TABLE 1List of synonyms (Gottschalck and McEwen 2004)

Ingredient Synonyms

Aloe Andongensis Extract Aloe ExtractExtract of Aloe andongensis

Aloe Andongensis Leaf Juice Aloe andongensisAloe Arborescens Leaf Extract Aloe arborescens

Kidachi AloeAloe Arborescens Leaf Juice Aloe arborescens

Kidachi Aloe KajyuKidachi Aloe YoujyuKidachi Aloe Youjyu Masto

Aloe Arborescens Leaf Protoplasts Aloe arborescensAloe arborescens CRS

Aloe Barbadensis Flower Extract Aloe barbadensisAloe barbadensis ExtractAloe Flower ExtractExtract of Aloe barbadensis FlowerExtract of Aloe Flowers

Aloe Barbadensis Leaf AloeAloe barbadensisAloe Leaf PowderAloe vera

Aloe Barbadensis Leaf Extract Aloe barbadensisAloe ExtractBarbados Aloe ExtractCuracao Aloe ExtractExtract of AloeExtract of Aloe barbadensisExtract of Aloe leavesAloe vera

Aloe Barbadensis Leaf Juice Aloe barbadensisAloe GelAloe barbadensis GelAloe JuiceAloe vera Gel

Aloe Barbadensis Leaf Polysaccharides Aloe barbadensisAloe Barbadensis Leaf Water Aloe barbadensisAloe Ferox Leaf Extract Aloe ferox

Extract of Aloe feroxCape Aloe

Aloe Ferox Leaf Juice Aloe feroxCape Aloe

Aloe Ferox Leaf Juice Extract Aloe feroxCape Aloe Ekisu

gel; they can be acetylated, partially acetylated, or not acety-lated. The gel portion (Aloe leaf juice) is the part of the plantgenerally used for emollient and moisturizing effects desired incosmetics, and the healing action desired for certain medicinalproducts.

The International Aloe Science Council (IASC) distinguishedbetween the anthraquinone found in the outer cell layer of thealoe leaf and the rest of the plant (IASC 2003). According tothis group, the maximum allowable aloin content in aloe-derivedmaterial for nonmedicinal use is 50 ppm or lower.


TABLE 2Description of Aloe products (Carrington Labs 2001)

Product name Appearance Odor Taste pH

Aloe vera Whole Leaf 1:1 Light amber liquid Light vegetable Slight bitter vegetable 3.04.0Aloe vera gel 1:1 decolorized Clear, slightly translucent Light vegetable Slight characteristic 3.55.0

vegetableAloe vera gel 10:1 decolorized Semiviscous, light to medium amber Light vegetable Slight vegetable 3.55.0Aloe vera gel 40:1 decolorized Semiviscous, light to medium amber Light vegetable Slight vegetable 3.55.0Aloe vera Gel Freeze Dried White to light tan powder Not given Not given Not given

Powder 200:1

Danhof (1998) stated that aloe polysaccharides consist oflinear chains of -1-4-linked glucose and mannose molecules, inwhich mannose predominates. Chain lengths may be as small asfrom a few molecules up to several thousand, but by conventionthe lower end of the molecular weight is usually considered to be1000 daltons, consistent with being a polysaccharide. Dependingon the particular chain length, this material will have differentphysical properties.

The IASC (1998) maintains a certification program in whichwhole leaf aloe vera gel is expected to adhere to the followingspecifications: solids between 0.46 to 1.31%; pH between 3.5and 4.7; calcium between 98.2 and 448 mg/L; magnesiumbetween 23.4 and 118 mg/L; and malic acid between 817.8 and3427.8 mg/L. Consistent with these specifications, Rowe andParks (1941) determined the pH of fresh Aloe vera leaves to be4.7.

TABLE 3Constituents of Aloe Vera (Shelton 1991; Vogler and Ernst 1999)

Nonessential Inorganic EssentialAnthraquinones Saccharides Vitamins amino acids compounds Enzymes amino acids Miscellaneous

Aloin Cellulose B1 Histidine Calcium Cyclooxygenase Lysine CholesterolBarbaloin Glucose B2 Arginine Sodium Oxidase Threonine TriglyceridesIsobarbaloin Mannose B6 Hydroxyproline Chlorine Amylase Valine SteroidsAnthranol L-Rhamnose Choline Aspartic Acid Manganese Catalase Leucine -SitosterolAloetic Acid Aldopentose Folic Acid Glutamic Acid Zinc Lipase Isoleucine LigninsCinnamic Acid C Proline Chromium Alkaline Phenylalanine Uric Acid

Ester phosphataseAloe-emodin -Tocopherol Glycine Copper Carboxypeptidase Methionine GibberellinEmodin -Carotene Alanine Magnesium Lectin-like

substanceChrysophanic Tyrosine Iron Salicylic Acid

AcidResistannol Arachidonic AcidAnthracene Potassium SorbateEthereal Oil

Table 2 presents descriptions and chemical properties of com-mercial Aloe products from one company analyzed by Carring-ton Labs (2001). All products listed in the table are consid-ered acceptable for use in cosmetics, and are derived from Aloebarbadensis.

Active Organics (2002) described a trade name product, Ac-tiphyte, of Aloe Vera 10 Fold, as consisting of Propylene Glycoland Aloe Barbadensis Leaf Extract. The product was a clear topale yellow liquid, with a characteristic odor, and a flash point of228F, boiling point of 225F, specific gravity of 1.02 to 1.05 (at25C), complete solubility in water, refractive index of 1.3620to 1.3700 (at 25C), and a pH of 4.0 to 6.5 (at 25C).

As noted earlier, aloe-derived ingredients also are character-ized by their specific constituents. The constituents reported byShelton (1991) and Vogler and Ernst (1999) found in Aloe veraare listed in Table 3.

ALOE 5

FIGURE 1Structures of the main anthraquinone compounds of Aloe species (Suga and Hirata 1983; van Wyk et al. 1995).

Mabusela et al. (1990) stated that the glucomannans of Aloeferox are mainly arabinogalactan and rhamnogalactan. Accord-ing to van Wyk et al. (1995), aloesin, aloeresin a, and Aloin makeup 70% to 98% of the total dry weight of Aloe ferox leaf exu-date. A description and some physical and chemical propertiesof these and other compounds present in the various Aloe speciesare found in Table 4.

Figure 1 shows the chemical structures of anthraquinonesand anthraquinone derivatives of Aloe barbadensis, Aloe ar-borescens, and Aloe ferox (Suga and Hirata 1983; van Wyk et al.1995). These compounds are generally found in the pericycliccells but can be found within the gel as well (see Methods ofManufacture and Impurities).

Ultraviolet AbsorptionProserpi (1976) examined the ultraviolet (UV) absorption of

various Aloe species and aloin, a component of Aloe species.Both Aloe extracts and aloin have spectrophotometric peaksat about 297 nm. The aloin spectrogram has a second peak at360 nm. The author concluded that cosmetics containing 1% to2% of aloe extract should give effective sunburn protection dueto the substances contained within Aloe performing selectivescreening of UV radiation, by absorbing mainly in the erythe-mogenic rays.

Bader et al. (1981) examined dry extracts of Aloe (contain-ing anthraquinones) for their ability to absorb light in the UVB

range. Extraction was done with a 50% waterethyl alcohol solu-tion. The aloe extracts had maximum absorption around 294 nm.The coefficient of extinction for a glycolic extract of Aloe verawas 73.8 at 308 nm and 85.0 at 295 nm (Bobin et al. 1994).Absorption and fluorescence emission spectra of aloe emodin inethanol yielded a maximum absorption wavelength of 451 nmand a maximum fluorescence emission peak at 537 nm (Vargaset al. 2002).

Absorption and fluorescence emission spectra of aloe emodinin acetonitrile yielded an absorption spectrum with a maximumat 430 nm. The fluorescence emission spectrum for aloe emodinat 430 nm exhibits with emissions peaks at 565, 530, and 495 nm.Increasing the fraction of water increased the intensity of the 565and 530 nm fluorescence peaks relative to the 495 nm peak. Aloeemodin was photostable at UVA radiation irradiances of 2.9 103 W/cm2 (Vath et al. 2002).

Methods of ManufactureAccording to Grindlay and Reynolds (1986), commercial cul-

tivation of Aloe vera for its gel began in the 1920s in Florida.Current cultivation in the United States primarily takes place inFlorida, Texas, and Arizona. Aloe vera can be harvested by hand,with the leaves cut off at the base of the plant. Individual leavesare wrapped, crated, and transported to processing plants wherethey are cleaned. Next the outer layers are removed by filleting;this allows the removal of the central filet of gel. Once the gel is

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antithyroidal agents. The Aloe vera gel was prepared by remov-ing the outer rind and the colorless parenchyma was ground ina blender and centrifuged to remove fibers. Swiss albino malemice were dosed with 125 mg/kg of Aloe vera by gastric intu-bation. Levels of hepatic lipid peroxidation (LPO), superoxidedismutase (SOD) and catalase (CAT) were not affected by ad-ministration of 125 mg/kg of Aloe vera as compared to vehiclecontrols.

Hypotensive EffectSaleem et al. (2001) reported that aloe-emodin and aloin A

extracted from Aloe barbadensis had a hypotensive effect onSprague-Dawley rats. The mean arterial blood pressure was re-duced by 26%, 52%, and 79% at corresponding doses of 0.5, 1,and 3 mg/kg.

ANIMAL TOXICOLOGY

Acute Oral ToxicityShah et al. (1989) administered Aloe vera extracted in ethanol

orally to Swiss albino male mice at doses of 500 mg/kg, 1 g/kg,and 3 g/kg. No signs of toxicity were observed, except in thehigher dose groups, in which there was a decrease in centralnervous system (CNS) activity noted (CNS activity measure-ment not reported).

Lagarto Parra et al. (2001) administered an aloe extractto Swiss albino mice (number not given) for estimation ofLD50. Aloe vera leaves were dried, chopped, and extracted inmethanol. The extracts (concentrations were not given) weregiven orally. The estimated LD50 for Aloe vera 24 h after dosingwas 120.65 mg/kg.

MDS Pharm Services (2000) reported a study in which AloeBarbadensis Leaf Water (2.01 ml volume of 100% Aloe) wasadministered by gavage to 10 Sprague-Dawley rats (5 females,5 males). There were no clinical signs of toxicity, mortality,weight changes, and no gross findings at necropsy (at day 14).

Acute Parenteral ToxicityDhar et al. (1968) conducted a study using the entire Aloe

barbadensis plant, ground up into a powder, extracted with 50%

ethanol, and dried. The dried extract was suspended in either0.1% agar solution or 1.0% gum acacia in distilled water. Thisextract was administered intraperitoneally to adult albino mice(either sex, two to three animals) at an initial dose of 400 to500 mg/kg. The dose was increased or decreased by a factor oftwo. The maximum tolerated dose (MTD) was 100 mg/kg bodyweight and the LD50 was 250 mg/kg.

Fogelman et al. (1992a) administered acemannan (in saline)once intravenously by tail vein to Sprague-Dawley rats andCD-1 mice or by cephalic vein to purebred Beagle dogs orby intraperitoneal injection to rats, mice, and dogs at volumesconsidered to be maximal without undue discomfort and thatdid not interfere with the interpretation of results. Each animalwas observed during the 1st hour following dosing, at 4 h, anddaily thereafter for 14 days. Body weights were recorded. Allanimals surviving to day 14 were euthanized and necropsied.Organ weights were recorded for liver, kidney, pancreas,thymus, thyroid, and spleen. No significant adverse effects,deaths or gross pathology occurred in any of the animals. Studyresults are summarized in Table 10.

Acute Dermal ToxicityAloe Ferox Leaf Extract was applied to shaved skin of male

white New Zealand rabbits. The shaved skin was scarified (exci-sion with a scalpel cutting the epidermis only) or not damaged.The two skin sites were exposed to 0.5 ml of the products andoccluded with hydrophilic patches. One of the six rabbits hada reaction of a very slight erythema on the scarified tissue thatcleared after 72 h and Aloe Ferox Leaf Extract was considereda nonirritant (0.04:8) (Institut Francais de Recherches et EssaisBiologoques 1981).

Short-Term Oral ToxicityAjabnoor (1990) reported that male albino mice (MFI strain)

were orally administered the bitter principal of Aloes in 0.9%saline (up to 20 mg/kg) for 2 weeks. Further experimental detailswere not provided. No mortality was observed.

Fogelman et al. (1992b) mixed acemannan in the basal diet ofSprague-Dawley rats at dose levels of 0, 6000, 12,500, 25,000,

ALOE 27

or 50,000 ppm for 2 weeks. Animals were observed twice daily.Five randomly selected rats/sex/group were evaluated for hema-tologic and serum chemistry parameters at termination (day 14).All survivors were necropsied and brain, heart, liver, kidney,adrenals, and testes weights were recorded. No mortalities oc-curred. Body weights, body weight gains, and food consumptionwere comparable in all groups. Hematology and serum chem-istry values were comparable between treated and control ani-mals. At necropsy, three males in the 12,500 ppm dose grouphad red/brown foci on the stomach; all other males were nor-mal. A female in the 50,000 ppm group had a reddened stomach.These observations were sporadic and not dose-related. Abso-lute and relative organ weights were not significantly differentfrom controls. The 14-day NOEL (no observed effect level) foracemannan in the diet, in rats, was 50,000 ppm, equivalent to4068 mg acemannan/kg/day in male rats and 4570 mg aceman-nan/kg/day in female rats.

Short-Term Parenteral ToxicityMale albino mice (MFI strain) were administered the bitter

principal of Aloes in 0.9% saline (up to 20 mg/kg) intraperi-toneally for 2 weeks. No mortalities were observed (Ajabnoor1990).

Fogelman et al. (1992a) reported that no significant toxicitieswere seen in rats, mice, and dogs after eight doses of acemannan(in saline) administered intravenously or intraperitoneally atfour-day intervals over 30 days. The results are summarized inTable 10.

Dose levels were 4 to 50 mg/kg for Sprague-Dawley rats,20 to 200 mg/kg for CD-1 mice, and 0.3 to 50 mg/kg for Bea-gle dogs. Rats and mice were distributed into groups of 10 persex and Beagle dogs were distributed into groups of 2 per sex.All animals were observed daily, and 1 and 4 hours after dosing.Body weights for dogs were recorded prior to each injection andweekly for the rats and mice. Food consumption was measuredweekly for rats and mice. In addition, all animals had ophthalmo-logic examinations prior to study initiation and at termination.All animals had hematologic and serum chemistry evaluations.Tissues obtained at necropsy were examined. In addition, selectorgan weights were recorded.

Of mice intravenously administered acemannan, 3/20 at the40 mg/kg/day and 6/20 at the 80 mg/kg/day dose level died.Three of these deaths were judged to be unrelated to the treat-ment. Clinical signs seen in the mice were limited to decreasedactivity, abnormal gait and stance, flaccid body tone, piloerec-tion, and tremors. These signs appeared to be transient as re-covery was apparent within 24 h. Mean body weights of the80 mg/kg/day acemannan group were significantly lower thancontrols on days 8, 15, and 22. Hematology and serum chemistryvalues were comparable to controls. Treatment related histolog-ical changes were found in the lungs at all dose levels and werecharacterized as minimal to slight multifocal accumulations ofmacrophages.

Acemannan administered intravenously to rats did not af-fect body weight and food consumption or produce any clinicalsigns of toxicity. Three rats died while on study but the causeof death was not apparent. Histologically, the test rats had thesame observations in the lungs as the mice.

Body weights of dogs treated intravenously with acemannanwere not affected. Clinical signs were limited to emesis seensporadically among the groups. Hematologic and serum chem-istry values were within normal ranges for Beagles (comparedto historical data). Histological findings in the dogs treated in-travenously were similar to those seen in the rats and mice.

In both rats and mice (intraperitoneal administration), bodyweights and food consumption were similar to the controls. Clin-ical signs of toxicity were not seen in the rats; however, themice had abnormal gait and stance, poor grooming, piloerec-tion, and ptosis. Five mice died from treatment related causes;no rats died while on study. In the rats and mice, hematology andserum chemistry values were generally similar to the controls.However, male mice at the 100 mg/kg/day dose and femalesat the 200 mg/kg/day dose levels had a significant increase intotal leukocytes. Significant differences in absolute or relativeorgan weights were seen in the mid- and high-dose male mice,high-dose female mice, and the mid- and high-dose female rats.Histological changes for both the mice and rats were limited tomicrogranulomas seen on the surfaces of the peritoneal organs.

As seen in the intravenous study, dogs treated with aceman-nan intraperitoneally did not have any changes in their bodyweights. The intraperitoneally treated dogs experienced emesis,decreased activity, and abdominal discomfort. Hematology andserum chemistry values were not significantly different fromcontrols; however, there were increases in the monocyte lev-els. No significant differences occurred in organ weights of thetreated dogs. Histological responses were similar to those seenin the rats and mice.

Maximum dose levels with no observed adverse effects were20 mg/kg intravenous or intraperitoneal in mice, and 4 mg/kgintravenous and 50 mg/kg intraperitoneal in rats. The no adverseeffect level for dogs was observed with 1 mg/kg intravenousand the lowest observed adverse effect level for intraperitonealadministration was 5 mg/kg (Fogelman et al. 1992a).

Subchronic Oral ToxicityShah et al. (1989) conducted a study in which 20 Swiss al-

bino male mice were orally administered Aloe vera (extractedin ethanol) in their drinking water at a dose of 100 mg/kg for3 months. Control animals were dosed with distilled water. Allanimals were observed for signs of toxicity and mortality, andbody weights were determined during the study. Blood and or-gans were analyzed from five animals in the control and Aloevera treatment groups at study termination. Body weights andorgan weights in treated animals were not different from con-trols. No abnormalities of the viscera were observed in treatedor control animals. Six of the twenty Aloe veratreated mice


died while on study; this was significant when compared to thecontrols. Alopecia of the genital region and degeneration andputrification of the sex organs occurred in 20% of the animals.Ten percent of the animals had inflammation of the hind limb.Red blood cell counts were considerably decreased in the Aloeveratreated animals.

Fogelman et al. (1992b) mixed acemannan in a predeter-mined quantity of feed to deliver doses of 0, 100, 400, or 1500mg/kg/day for each purebred Beagle dog over a 90-day dos-ing period. The dogs were observed twice daily for clinicalsigns of toxicity. Detailed examinations were done prior to ini-tiation of dosing and at weeks 4, 5, 8, 9, 10, 11, 12, and 13.Body weights were recorded weekly. Ophthalmic examinationswere conducted prior to study initiation and at termination ofthe study. Serum chemistry, hematologic, and urinalysis datawere recorded at day 45 and at termination. The animals wereeuthanized at day 90 and subjected to a complete necropsy.

There were no significant signs of systemic toxicity. Bodyweights and food consumption were comparable between treatedand nontreated dogs. Serum chemistry, hematology, and urinal-ysis data from the treated dogs were all within the normal limitsat all times evaluated. No significant gross or microscopic le-sions were attributed to the ingestion of acemannan. The NOELfor acemannan, orally administered to Beagle dogs, was at least1500 mg/kg/day (estimated), which is equivalent to 1,170 mgacemannan/kg/day.

In a six month study by these authors, acemannan was mixedin the basal diet of Sprague-Dawley (20 rats/sex/group) rats toprovide acemannan doses of 0, 200, 650, or 2000 mg/kg/day.Individual body weights were determined initially, weekly for15 weeks, biweekly thereafter, and at study termination. Foodconsumption was measured during the week prior to initiationand concurrently with body weights throughout the study. Twicedaily observations were made for mortality and clinical signs oftoxicity. Concurrent with the body weight data collection, de-tailed physical examinations were performed. Ophthalmic ex-aminations were performed initially and at study termination.At 1, 3, and 6 months, 10 rats/sex/group underwent hematology,serum chemistry, and urinalysis determinations. Complete grossnecropsy examinations were conducted on all animals that diedduring the study, 10 rats/sex/group at day 90, and all survivorsat 6 months. Organ weight data were obtained and select tissuesfrom the control and high-dose groups were examined.

Body weights for males and females receiving acemannanwere comparable to controls throughout the study. Male ace-mannan consumption ranged from 193 to1986.3 mg/kg/day andfemale acemannan consumption ranged from 200.9 to 1994.8mg/kg/day. Adverse clinical signs were randomly distributedamong the dosage groups and sexes, and were not consideredto be due to acemannan. Ophthalmological examinations re-vealed no significant changes among the rats. All serum chem-istry, urinalysis, and hematological values were normal. Organweights and gross and microscopic pathology for the treated ratswere normal and similar to the corresponding controls. No gross

lesions attributable to the ingestion of acemannan were noted inany of the rats.

Six female rats died during the study; four deaths were at-tributed to the bleeding procedure. Of the two unexplaineddeaths, one female at the 2000 mg/kg/day dose died on day30 and the gross necropsy revealed enlarged and mottled kid-neys. The other unexplained death occurred on day 139 in afemale receiving the 650 mg/kg/day dose. The cause of deathwas confirmed histologically as chronic pyelonephritis. The 6-month NOEL in rats was 2000 mg/kg/day, which was equivalentto 1549 mg acemannan/kg/day for males and 1555 mg aceman-nan/kg/day for females (Fogelman et al. 1992b).

Herlihy et al. (1998a) administered Aloe barbadensis to fourgroups of SPF Fischer 344 male rats (20 rats per group) for 1.5or 5.5 months. Aloe barbadensis was processed into four for-mulations. Formulation A1 was a 1% preparation of Aloe filetsthat had been homogenized, lyophilized, and frozen. Formula-tion A10 was the same preparation, at a 10% concentration. B1and B10 formulations were prepared the same way as A1 andA10; however, the homogenate was charcoal filtered prior tolyophilization (removal of barbaloin) to more closely resemblecommercial products. The four groups of rats received one of theAloe preparations in their diet; a fifth, the control group, was fednormal rat chow. Data were collected for food consumption, wa-ter consumption, body weights, clinical signs, blood chemistry,histopathological analysis, and gastrointestinal transit time.

Due to diarrhea and restricted growth rates seen in therats fed A10, this group was eliminated from the study.Formulations A1, B1, and B10 did not have any effect on bodyweights, food consumption, organ weights, or gastrointestinaltransit time when compared to the controls. B10 increasedwater consumption; however, analysis of the rat chow/Aloemixture revealed increased salt levels that arose from theconcentration of the salts during lyophilization. In the groupsfed the Aloe preparations, pathology and blood chemistrywere not significantly different from the controls, except fordecreased serum cholesterol, HDL, aspartate transferase, andalanine transferase (Herlihy et al. 1998a).

Ocular ToxicityAloe Ferox Leaf Extract was instilled in the conjunctival sac

of the eye of male New Zealand white rabbits. Six healthy malerabbits received 0.1 ml (no concentration specified) of AloeFerox Leaf Extract in the conjunctival sac of the eye and wereobserved 1 h, 24 h, 2 days, 3 days, and 7 days after administra-tion. No significant changes were seen except for 5/110 com-bined Draize score after 1 h (all six animals exhibited minimalchemosis and 5/6 had slight corneal erythema), a 2/110 com-bined Draize score after 1 day, and 0/110 after day 2 (InstitutFrancais de Recherches et Essais Biologoques 1981).

The Institut DExpertise Clinique (2000) administered AloeBarbadensis Leaf Water (300 l; no concentration specified) tothe vascular chorionic-allantoic membrane of White Leghorn

ALOE 29

PA12 eggs in an alternative ocular irritation assay. After contactfor 20 s, the membrane was washed with 5 ml of physiologicalserum. There were four eggs per treatment; a negative controlof 0.9% physiological serum; and positive control of sodiumdodecyl sulfate (SDS) 0.5% w/v. The mean irritation index forSDS was 12.0 (irritant) and the Aloe irritation index was 1.3(slight irritant).

REPRODUCTIVE AND DEVELOPMENTAL TOXICITYShah et al. (1989) reported that Swiss albino male mice were

orally administered Aloe vera (extracted in methanol) in theirdrinking water for three months at a dose of 100 mg/kg. The ex-tracted Aloe vera caused significant sperm damage (megacephaliand flat head > swollen achrosome > rotated head) when com-pared to the controls (dosed with distilled water).

Parry and Matambo (1992) studied the reproductive toxicityof three Aloe species (Aloe chabaudii, Aloe globuligemma, Aloecryptopoda) in Wistar rats and mice (strain not given). No in-formation was available on the similarity of extracts from thesespecies and those addressed in this report. The three Aloe plantswere prepared the same way: pounded with a mortar and pestle,collection of the juices, filtered through muslin cloth, and freezedried. The dried powder was reconstituted with distilled water.Ten rats received 500 mg/kg of A. chabaudii (eight were dosedintraperitoneally and two were dosed orally) on days 14, 15,and 16 of their pregnancy. Eight rats received 500 mg/kg of A.cryptopoda intraperitoneally and eight rats received 500 mg/kgintraperitoneally of A. globuligemma. Control rats were admin-istered saline intraperitoneally. The mice were only adminis-tered A. chabaudii either orally (four) or intraperitoneally (five)(500 mg/kg) on days 14, 15, and 16 of their pregnancy. Controlgroups received normal saline either intraperitoneally or orally.

Even though a number of rats died due to the toxicity ofthe Aloe, no resorptions of the fetuses occurred. Additionally,no abortions were noted. Rats and mice that survived deliverednormal sized, healthy litters (Parry and Matambo 1992).

Nath et al. (1992) extracted ground up leaves of Aloe bar-badensis with water and orally administered (vehicle was 1%acacia) the material to five pregnant Charles Foster albino rats.Dose administration took place from gestation day 0 though ges-tation day nine. Five control rats were given the vehicle only.Cesarean delivery was performed on gestation day 20. A total of51 fetuses was examined for macroscopic effects, 25 for visceralabnormalities, and 26 for skeletal abnormalities.

In the treatment group, per animal, the implantations were13.0 1.0 (8.6 1.1 in controls); resorptions were 2.8 1.1(none in controls); live births were 10.2 1.3 (8.6 1.1 incontrols); fetal body weight was 2.60 0.85 (4.83 0.44in controls); and fetal body length was 2.87 0.42 (3.87 0.22 in controls). Overall 21.5% abortifacient activity (resorp-tions/implantations) was calculated (0% in controls). No macro-scopic, visceral, or skeletal deformities were reported in controlfetuses. Macroscopic effects in treatment fetuses included kink-

ing of tail (5.9%), clubbing of right hind limb (11.8%), andleft wrist drop (19.6%). No visceral abnormalities were seenin treatment fetuses. Skeletal deformities in the treatment fe-tuses included nonossification of skull bones (15.4%), nonossi-fied ribs (15.4%), fused tarsal (15.4%), and intercostal space inribs (11.5%). The authors noted that traditional medicinal plantsused as abortifacient agents can have an effect on the fetus if not100% effective (Nath et al. 1992).

GENOTOXICITYBoth negative and positive results were found in bacterial

and mammalian cell genotoxicity assays using Aloe barbaden-sis, Aloe ferox and various components of Aloe. The data aresummarized in Table 11.

CARCINOGENICITYSiegers et al. (1993) reported that male NMRI mice were

administered different diets to test if aloin was carcinogenic. Thesubjects were randomized into five groups (20 per group), andeach was fed a different diet for 20 weeks. Group 1 was injectedsubcutaneously with 20 mg/kg of dimethylhydrazine (DMH) toinduce tumors weekly for 10 weeks plus 0.03% (equivalent to100 mg/kg/day) sennosides in diet for 20 weeks. The secondgroup was injected subcutaneously with 10 mg/kg of EDTA(solvent for carcinogen) plus 0.03% sennosides in diet for 20weeks. Group 3 was injected subcutaneously with 20 mg/kgDMH weekly for 10 weeks plus normal diet for 20 weeks. Group4 was injected subcutaneously with 20 mg/kg DMH weekly for10 weeks plus 0.03% aloin in diet for 20 weeks. The final groupwas injected subcutaneously 10 mg/kg EDTA weekly for 10weeks plus 0.03% aloin in diet for 20 weeks. The mice weresacrificed by decapitation at the end of the experimental period.Tumors were detected only in the distal segment of the colonand rectum.

The authors concluded that aloin did not significantly alterthe incidence of colorectal tumors. Group 3 (normal diet plusDMH) produced 10/19 tumor-bearing animals, group 4 (aloinplus DMH) had 7/20 tumor-bearing animals, and group 5 (con-trol for aloin) had 1/ 20 tumor-bearing animals. The mean tumorrate in tumor-bearing animals was 1.50 0.22, 1.0, and 2.0 0.58 for groups 1, 2, and 3, respectively (Siegers et al. 1993).

Strickland et al. (2000) reported a study in which semi-synthetic and extracted aloe-emodin (from Aloe barbadensis),in 25% ethanol, were applied to specific pathogen free femaleC3H/HeNCr (MTC) mice. Mice were exposed to 15 kJ/m2UVB radiation three times weekly. Treatment was with eitheraloe-emodin or 25% ethanol (1 ml per mouse) and took placebefore UVB exposure. Control mice were not irradiated butwere treated with either aloe-emodins or ethanol. Two addi-tional groups were treated with either a total of 90 kJ/m2 UVradiation for 2 weeks then aloe-emodin three times weekly for31 weeks or a total of 60 g aloe-emodin and then UV radi-ation for 31 weeks. The mice administered either aloe-emodin


TABLE 11Genotoxicity of aloe compounds

Material tested Assay type Treatment Results Reference

Bacterial Cell AssaysAloe-emodin Salmonella

TA1537With and without

activation,100 g/plate

61 His revertants per plate,0.22 per nmole of aloeemodin; activation hadno effect

Brown et al. 1977

Aloe-emodin With and withoutactivation,50 g/plate

330 His revertants perplate, 0.42 per nmole ofemodin; significantactivation

Aloe-emodin SalmonellaTA1535,TA100,TA1537,TA153,TA1538, TA98

With and without S9activation

Negative in all strainsexcept for TA1537without S9

Brown and Dietrich1979

Aloin Salmonella/microsomeTA1535,TA100,TA1537TA1538, TA98

With and without S9,50250 g/plate

Negative all strains Brown and Dietrich1979

Aloe ferox waterextracted

Bacillus subtilisrec-assay

With and withoutmetabolic activationconcentrations of 5,10, 20, 50, 100 mg/ml

Positive Morimoto et al. 1982

SalmonellaTA98, TA100

Negative

Aloe feroxethanolextracted

Bacillus subtilisrec-assay

Negative


Negative

Aloe-emodin SalmonellaTA1537,TA1538, TA98

With and withoutactivation11000 g/ml

Mutagenic in all strainswith and withoutactivation

Westendorf et al. 1990

Aloe-emodin Salmonella TA1537, TA1538,TA98

105000 g/platewith/without S9

TA 1537 and TA 98negative with S9,positive without S9;TA1538 positivewith/without S9

Heidemaan et al. 1993

Aloe-emodin Micronucleus test NMRI mice,1500 mg/kg

Negative Heidemaan et al. 1993

Aloe-emodin ChromosomeAberration test

Wistar Rats, 2000mg/kg


Crushed leaves ofAloebarbadensis


TA98 with and withoutmetabolic activation,6 mg/ml

All strains negative Badria 1994

ALOE 31

TABLE 11Genotoxicity of aloe compounds (Continued)

Material tested Assay type Treatment Results Reference

Aloe-emodin SalmonellaTA1535,TA1537,TA1538, TA98

105000 g/plate withor without S9

TA1535 negativewith/without S9;TA1537 and TA 98negative with S9,positive without S9;TA1538 positivewith/without S9


Mammalian Cell AssaysAloe-emodin C3H/M2

fibroblasts130 g/ml Positive reaction in

transformation of cellsWestendorf et al. 1990

Aloe-emodin V79-HGPRTassay

530 g/ml Weakly mutagenic Westendorf et al. 1990


CHO cells with andwithout S9, at18.7575.00 g/ml

With S9, positive at3775 g/ml


Without S9, positive at18.7575 g/ml

Aloe-emodin MouseTFT-resistanceL5178Y tk+/mutation assay

37111 M tk mutations were inducedin mouse lymphomacells L5178Y

Mueller et al. 1996

Aloe-emodin V79-HGPRT 1.6 106 cells exposedwith and without S9,5350 g/ml

Negative with/without S9 Heidemaan et al. 1996

Animal AssaysAloe-emodin Mouse Spot test NMRI females crossed

with DBA malesgiven 200 and 2000mg/kg on gestationday 9


Aloe-emodin Micronucleus test NMRI mice,1500 mg/kg



Wistar rats, 2000 mg/kg Negative Heidemaan et al. 1996

In Vitro AssaysAloe-emodin UDS test In vitro and in vivo in

Wistar rats, 100 and1000 mg/kg


alone or vehicle alone did not develop tumors by study termi-nation. Sixteen out of 20 mice dosed with either 2 g or 5 gof either aloe-emodin and then UV radiation had tumors bystudy termination; however, the number was not significantlydifferent from the ethanol-treated/UV-radiated mice. However,aloe-emodin significantly increased the incidence of melanomatumors when compared to the unirradiated controls. Mice thatwere exposed to UV radiation first and then aloe-emodin didnot develop tumors. Mice that were exposed to aloe-emodin and

then the UV radiation did develop tumors, 30% of which weremelanoma.

AnticarcinogenicityAloe barbadensis

Harris et al. (1991) administered acemannan to 32 dogs and11 cats that had spontaneous malignant tumors that had failedconventional treatment. Each animal received 1 mg/kg of ace-mannan intraperitoneally and 2 mg of acemannan injected into


TABLE 12Effect of acemannan on fibrosarcomas (King et al. 1995)

Species Age Sex Results Tumor-free intervala Survival (days)b

Canine 3 M Free of disease 408+ 442+Canine 7 F Free of disease 559+ 603+Canine 7 M Died post surgery 0 52Canine 8 F Free of disease 409+ 440+Canine 10 F No viable tumor present at time of

surgery303 303

Canine 12 M Died week 11, recurrent disease 77 144Canine 12 F Free of disease 475+ 498+Canine 13 F Died; recurrent tumor outside

radiation field162 225

Feline 6 M Euthanized; recurrent tumor 204 246Feline 7 M Free of disease 555+ 594+Feline 8.5 F Free of disease 428+ 467Feline 12 M Free of disease 450+ 472+Feline 13 F Died post surgery, no viable tumor

present at time of surgery0 97

aTime from surgical intervention.bTime from study entry.

the tumor, if possible. Treatment occurred every 3 weeks for atotal of five treatments. Thirteen of the 43 animals had no sig-nificant clinical or histopathological response. Among the 13animals, 5 were lost to follow-up, 7 died after receiving oneinjection, and 1 had no response after a full course of therapy.Twelve animals had obvious signs of clinical improvement. Im-provement was based on tumor shrinkage, tumor necrosis, oran unexpected long survival period. No toxicity was associatedwith intraperitoneal administration of acemannan.

Peng et al. (1991) reported that acemannan, from Aloe bar-badensis, initiated the production of monokines that supportedantibody dependent cellular cytotoxicity and stimulated blas-togenesis in thymocytes. Additionally, acemannan, in both en-riched and highly purified forms, was administered intraperi-toneally to female CFW mice (number not given) into whichmurine sarcoma cells had been previously subcutaneously im-planted. The highly malignant sarcoma cells grew in 100% ofthe implanted control animals resulting in mortality within 20 to46 days. In the animals treated with acemannan at implantation,approximately 40% survived. The tumors from the acemannan-treated animals had vascular congestion, edema, polymorphonu-clear leukocyte infiltration, and central necrosing foci with hem-orrhage and peripheral fibrosis.

Sheets et al. (1991) administered acemannan intraperi-toneally to cats diagnosed with feline leukemia. Forty-four catswere dosed for 6 weeks at 2 mg/kg and observed for an additional6 weeks after the end of dose administration. At the end of the12-week period, 29/41 cats were still alive. Two of the originalcats were lost in follow-up and 1 died of other causes; the other

15 died of feline leukemia. Two months after the completion ofthe study the owners of 22 cats were interviewed. Twenty-oneof those cats were still alive, and according to the owners, happyand healthy pets.

King et al. (1995) treated eight dogs and five cats with new orrecurring fibrosarcomas with acemannan. All animals received 1mg/kg body weight of acemannan weekly for 6 weeks, followedby monthly injections for 1 year. Prior to surgery, 2 mg of ace-mannan was injected into the tumor weekly for up to 6 weeks.Between the 4th and 7th weeks following initiation of intraperi-toneal injections, surgical incision was attempted on the tumors.All animals that survived surgery immediately underwent radi-ation therapy. Of the 13 animals in the study, 7 survived andremained tumor free. Mean survival time was 240 days. Of thesix animals that died, two died from postsurgical complicationsand one died from other causes. Study results are summarizedin Table 12.

Kim and Lee (1997) tested aloe gel extracts from Aloe bar-badensis for their chemopreventive capabilities in vitro and invivo. Rat hepatocytes treated with [3H]benzo[a]pyrene in theabsence and presence of the Aloe gel extracts (0 to 250 g/ml)had a dose-related inhibition of [3H]benzo[a]pyrene adduct for-mation (9.1% to 47.7%) in the Aloe gel extracts group. Nocytotoxic effects on rat hepatocytes were observed. Aloe gelextracts (250 g/ml) and [3H]benzo[a]pyrene incubated withrat hepatocytes for 3 to 48 h had a slight inhibitory effect ofAloe gel extracts on DNA adduct formation at 3 h with max-imum inhibition occurring at 6 h (36%). Male ICR mice (18mice/group) were assigned to the following groups: group 1,

ALOE 33

control; group 2, daily oral treatment with 50 mg/mouse/day ofAloe gel extracts in distilled water; group 3, single oral dose ofbenzo[a]pyrene (10 mg/mouse in corn oil); group 4, single oraldose of benzo[a]pyrene (10 mg/mouse in corn oil) then dailyoral treatment with Aloe gel extracts (10 mg/mouse/day) for16 days; and group 5, oral pretreatment with Aloe gel extracts(10 mg/mouse/day) for 16 days followed by a single oral dose ofbenzo[a]pyrene (10 mg/mouse in corn oil) and then continuoustreatment with 50 mg/mouse/day of Aloe gel extract for 16 days.

DNA adduct formation was significantly decreased in groups4 and 5 when compared to group 3. Glutathione S-transferaseactivity was slightly increased in the liver but cytochrome P450content was not affected by Aloe gel extract. The authors con-cluded that the inhibitory effect of the Aloe gel extract onbenzo[a]pyrene diol epoxideDNA adduct formation might havea chemopreventive effect by inhibition of benzo[a]pyrene ab-sorption (Kim and Lee 1997).

Corsi et al. (1998) conducted a study in which male Fisherrats (number not given) previously injected with Yoshida AH-130 ascite hepatoma cells (2 105 in a saline suspension) intothe pleural space received Aloe vera (20 g suspended in 0.5ml sterile saline) injections daily for 2 weeks. Injections of Aloevera were also administered into the pleural space. At speci-fied times (days 7 and 14) the growth of the tumor cells in thepleural space was evaluated. The use of Aloe vera significantlydecreased tumor cell growth with respect to nontreated controlanimals. The survival of rats was progressively prolonged whenAloe vera was used as a therapy in the tumor-bearing animals.

Shamaan et al. (1998) studied the effects of an aloe derivativein rats. Male Sprague-Dawley rats were divided into six groupsof five to eight rats per group. Groups 1 to 3 received a basaldiet: group 1 was the control group and received no additionaltreatment; group 2 received vitamin C in their drinking water;group 3 received 0.1 g Aloe vera gel extract/100 ml in theirdrinking water for the 9-month study. A single dose of diethyl-nitrosamine (DEN) 200 mg/kg body weight and a diet contain-ing 0.02% (w/w) 2-acetylaminofluorene (AAF)-induced chem-ical hepatocarcinogenesis in groups 4 to 6. Group 5 receivedvitamin C and group 6 received Aloe vera gel extract as pre-viously described. DEN/AAF treatment significantly increasedthe GGT ( -glutamyltransferase) concentration in carcinogen-induced rats; however, the addition of Aloe vera gel extract sig-nificantly decreased the activity of GGT. Both GGT- and GSTP(placental glutathione S-transferase)-positive foci were detectedin the carcinogen treated rats; however, the addition of Aloe veragel extract to the drinking water of group 6 caused a significantdecrease in number of foci on their livers. The livers of the ratsin group 6 were devoid of neoplastic nodules whereas cellulardamage was seen in group 4. Aloe vera gel extract supplemen-tation was found to be able to reduce the severity of chemicalhepatocarcinogenesis.

Kim et al. (1999) screened a polysaccharide of Aloe gel ex-tract of Aloe barbadensis for its chemopreventive effects us-ing biomarkers involved in chemical carcinogenesis. In the

benzo[a]pyrene (B[a]P)-DNA binding assay, a concentration de-pendent inhibition of DNA adduct formation occurred with theAloe polysaccharide (180 g/ml). Oxidative DNA damage by 8-hydroxyoxyguanosine was significantly decreased by the Aloepolysaccharide (180 g/ml). Additionally, the Aloe polysac-charide had a positive effect in a dose-dependent manner oninhibition of phorbol myristic acetate (PMA)-induced tyrosinekinase activity in HL-60 cells and PMA-induced ornithine decar-boxylase activity in BALB/3T3 cells. The Aloe polysaccharidesignificantly inhibited superoxide anion formation.

Pecere et al. (2000) screened aloe-emodin in vitro and invivo for its anticarcinogenicity activity against highly malig-nant tumors: neuroectodermal tumors, Ewings sarcoma, andneuroblastoma. The in vitro study was conducted first cultur-ing neuroblastoma cells, pPNET cells, Ewings sarcoma cells,T-cell leukemia cells, vinblastine-resistant cells, colon adeno-carcinoma cells, and doxorubicin-resistant cells. Cytotoxicityactivity was determined by growing cells in the presence ofaloe-emodin for 72 h, using many different concentrations ofaloe-emodin. After 24 and 48 h, the cells were scraped, washed,fixed overnight in a buffer solution, and processed for viewing.The results showed that aloe-emodin decreased the survival oftumor cells and normal fibroblasts. There was a large difference,however, between the two. The reduction in the neuroectodermaltumor cell lines was much more significant than in the differenttumor cell lines and normal fibroblasts.

The results of the in vitro study led to another antitumor studythat used 6-week-old female severely compromised immunode-ficient (SCID) mice. The mice were organized into three differ-ent groups. The first two groups both received IP injections in thedorsal region of 10 106 human neuroblastoma cells; however,the difference between the two groups was when they startedtreatment. Drug treatment started immediately in the first groupthat continued for 5 days for a total of 5 doses. Five animalsreceived 0.4 ml/day of DMSO (saline solutioncontrol), andthe other mice were treated with 50 mg/kg/dose of aloe-emodin.The second group received the same doses, except treatmentdid not start until 15 days after they were injected with tumorcells. All of the mice were sacrificed in groups 1 and 2 when themean tumor volume in the control was 1.5 cm3. In group 3, thetreatment was exactly the same as group 1; however, these micereceived the same amount of IP injections of colorectal adeno-carcinoma a cells instead of neuroblastoma cells. All of the micewere sacrificed at day 30. In all three groups, the tumors weremeasured with a micrometer caliper twice a week throughoutthe study.

The tumors in group 1 (immediate treatment of aloe-emodin)were very sensitive to the drug, showing a significant reduc-tion of growth in the animal hosts. When aloe-emodin treatmentwas delayed (group 2) until a palpable tumor mass had devel-oped (day 15), tumor growth was halted throughout the periodof drug administration. The group that was injected with col-orectal adenocarcinoma cells was refractory to the treatment.No appreciable signs of acute or chronic toxicity were observed


in any of the treated animals; weight, neurological and intestinalfunctions, and hematological parameters were normal (Pecereet al. 2000).

Singh et al. (2000) investigated the cancer chemopreventivecapabilities of Aloe vera leaf pulp extract by analyzing Aloeseffects on the enzymes that play a role in carcinogen metabolismand those involved in cellular antioxidant activity. Random-bredSwiss albino male mice were divided into four groups: group 1was fed a normal diet and sham treated with distilled water byoral gavage for 14 days; group 2 was also fed a normal diet; how-ever, they were treated with 30 l of Aloe vera leaf pulp extractper animal by oral gavage for 14 days; group 3 was also fed a nor-mal diet; however, their treatment amount was 60 l of Aloe veraleaf pulp extract per animal, also delivered by oral gavage; group4 was fed a diet containing 0.75% butylated hydroxyanisole(BHA) for 14 days. The last group served as a positive controlbecause BHA is known to be an anticarcinogenic compound.The animals were killed and their livers homogenized. The re-sulting supernatant from the livers was used to assay for totalcytosolic glutathione S-transferase, DT-diaphorase, lactate de-hydrogenase, and antioxidant enzymes. The pellet was used forassaying cytochrome P450, cytochrome b5, cytochrome P450reductase, cytochrome b5 reductase, and lipid peroxidation.

During the treatment phase of the experiment there were noanimal deaths or decreases in body weight. Additionally, therewas no indication of cellular damage. Aloe vera leaf pulp de-creased the levels of cytochromes P450 and b5, but increased theactivities of cytochrome P450 and b5 reductases. There were sig-nificant increases in the activity of glutathione S-transferase andDT-diaphorase in the high-dose group as compared to the con-trol. In the low-dose group, the enzyme activity was comparablewith that of the control group. The activity of lactate dehydroge-nase was decreased in a dose-dependent manner. The activitiesof the antioxidant enzymes (glutathione, glutathione peroxidase,glutathione reductase, superoxide dimutase, and catalase) wereall increased in a dose-dependent manner. The extent of lipidperoxidation was decreased significantly in the high-dose group;the low-dose group was also inhibited but it was not significant(Singh et al. 2000).

Lee (2001) reported aloe-emodininduced apoptosis in lungcell carcinoma cell lines CH27 (human lung squamous carci-noma cells) and H460 (human lung nonsmall cell carcinomacells). During apoptosis increases in cytochrome c and activationof caspase-3 were observed and expression of protein kinase Cappeared to occur downstream of caspase-3.

Kuo et al. (2002) reported that aloe-emodin inhibited cell pro-liferation and induced apoptosis in two liver cancer cell lines,Hep G2 and Hep 3B. Hep G2 cells induced p53 (tumor suppres-sor gene) expression and was accompanied by induction of p21(cell cycle regulator) expression that was associated with a cellcycle arrest in G1 phase. Fas/APO 1 (type one I membrane pro-tein) receptors and Bax (proapoptotic protein) expression wasincreased in Hep G2 with aloe-emodin. In Hep 3B cells, theinhibition of cell proliferation by aloe-emodin was mediated

through p21, and neither cell cycle arrest nor an increase in thelevel of Fas/APO1 receptor was observed. Bax expression wasincreased by aloe emodin in Hep 3B cells.

Wasserman et al. (2002) described Merkel cell carcinoma(MCC) as a rare and aggressive tumor of the skin, also known asa primary neuroendocrine carcinoma of the skin. A free floatingcell line was established from a metastasis from a MCC patient.Aloe-emodin decreased the viability of carcinoma cells after 72 hof treatment. The results are statistically significant (p < .02)for 10 mol and higher concentrations (100 mol) (p < .001).Aloin was also investigated at the same concentration range andno effect was detected.

Aloe arborescensYagi et al. (1977) injected aloemannan, a partially acetylated

-d-mannan isolated from the leaves of Aloe arborescens, in-traperitoneally into ICR mice (10 animals/group) that were im-planted with sarcoma-180 cells. Doses of 5 and 100 mg/kg wereadministered daily for 10 days and produced inhibition ratios of38.1% and 48.1%, respectively. No toxicity was observed.

Imanishi et al. (1981) administered aloctin A, a glycoproteinof Aloe arborescens, intraperitoneally to BALB/c mice (5 to 6weeks of age) once daily for 5 days after tumor implantation.Tumors were induced by methylcholanthrene. At 10 mg/kg/dayfor 5 days, aloctin A significantly inhibited the growth of themethylcholanthrene-induced fibrosarcomas. To determine if thealoctin A had direct cytotoxicity on tumor cells or was due tohost-mediated effects, the authors tested the effect of aloctin Aon various tumor cell lines. Aloctin A had almost no inhibitoryeffect on the growth of tumor cell lines up to a concentration of200 g/ml.

Uehara et al. (1996) measured the formation of 2-amino-3-methylimidazo[4,5-f]quinoline (IQ)-DNA adducts by 32P-postlabeling analysis. Cytochrome P450 (CYP) 1A1 and 1A2protein levels were analyzed by ELISA (enzyme-linked im-munosorbent assay) to assess the mechanism of chemopreven-tion of hepatocarcinogenesis by freeze-dried whole leaves ofAloe arborescens. Ten male F344 rats were fed a diet contain-ing 30% Aloe arborescens over an 8-day period. The 11 controlanimals were fed a basal diet only. On day 7, all animals weresubjected to a two-third partial hepatectomy (PH). Twelve hoursafter the PH, the rats received a single intragastric dose of thecarcinogenic food pyrolysate (IQ) (100 mg/kg) to initiate hepa-tocarcinogenesis. Rats were killed 6, 12, 24, and 48 h after IQadministration.

The levels of adducts, expressed as relative adduct labelingvalues, in rats treated with Aloe arborescens were decreased ascompared with the control group at hour 24 (36 h after PH) andhad further decreases at hour 48. The levels of CYP1A2, knownto be responsible for the activation of IQ, also were decreased athour 48. The authors concluded that Aloe arborescens has thepotential to decrease IQ-DNA adduct formation, presumably asa result of decreased formation of active metabolites (Ueharaet al. 1996).

ALOE 35

Shimpo et al. (2001) examined freeze-dried powder of whole-leaf Aloe arborescens Miller var. natalensis Berger (Japanesename Kidachi aloe) for its modifying effect on azoxymethane(AOM)-induced aberrant crypt foci (ACF), putative neoplasticlesions, in male F344 (4-week-old) rats. Rats were fed a basaldiet (groups 1 and 5), experimental diet with 1% (groups 2, 3,and 6), or 5% (groups 4 and 7) aloe for 5 weeks. On days 7, 14,and 21, groups 1 to 4 were given a subcutaneous injection of15 mg/kg of AOM and groups 5 to 7 received a subcutaneousinjection of normal saline (4 ml/kg).

No rats died. The 5% aloe diet rats had a mud-like or softfeces. Body weight was reduced significantly in all the AOM-dosed rats compared to the AOM-untreated rats. ACF developedonly in the rats treated with AOM (with or without aloe) and noACF were present in the untreated rats. ACF were decreased by35% and 14% with the 1% and 5% aloe treatments, respectively.AOM had no effect in the quinone reductase (QR) levels in theliver, although QR activity was significantly greater in the liverof rats fed 5% aloe with or without AOM treatment as comparedto the rats fed a basal diet (Shimpo et al. 2001).

Furukawa et al. (2002) reported that freeze-dried whole leafpowder of Aloe arborescens modified the initiation phase ofcancer in female Syrian hamsters (total = 90; group = 30).N -nitrosobis(2-oxopropyl)amine (BOP) was administered sub-cutaneously for 4 weeks at a dose of 10 mg/kg. Aloe was givenin the diet at 0%, 1%, or 5% for 5 weeks. At 54 weeks all surviv-ing animals were killed and the development of neoplastic andpreneoplastic lesions was assessed histopathologically.

Aloe at 5% decreased significantly the number of animalswith pancreatic adenocarcinomas (58.6%) compared to BOPalone (86.6%) and decreased significantly the number of ani-mals with atypical hyperplasia (27.5%) compared to BOP alone(56.6%). Adenocarcinomas per animal were decreased signif-icantly by Aloe at 5% (0.69 0.66) compared to BOP alone(1.20 0.76) and atypical hyperplasias per animal were de-creased significantly (0.34 0.61) compared to BOP alone(0.76 0.77). The decreases seen in the group treated withAloe at 1% were significant only for the number of animalswith adenocarcinomas and the number of adenocarcinomas peranimal, compared to BOP alone. The authors concluded thattreatment with the Aloe arborescens derivative has an inhibitoryeffect during the induction phase for pancreatic neoplastic andpreneoplastic lesions (Furukawa et al. 2002).

Shimpo et al. (2002) reported that the ethyl acetate extractof an acetone soluble Aloe arborescens Miller var. natalensisBerger (Japanese name Kidachi aloe) fraction inhibited 12-O-tetra-decanoylphorbol-13-acetate (TPA)-induced ear edema (anearly marker of tumor promotion) in ICR mice (n: control =4; TPA = 6; TPA + Aloe extract = 6). The effect of topicaltreatment of 1 mg of aloe with 1.6 nmol topically applied TPAto the ear was 40% reduction of edema (p < .0005). Aloe ex-tract itself did not induce edema. The 20 mg of aloe extracttogether with 5 nmol TPA also inhibited the TPA-induced in-crease in epidermal putrescine level by 30% (p < .005). Aloe

extract affected the promotion of skin papillomas by TPA in7,12-dimethylbenz[a]anthracene (DMBA)-initiated mice (n =20/group). The extract lowered the percentage of tumor-bearingmice by 45% after 10 weeks (p < .005) and average number oftumors decreased (no tumors in solvent control, 25.3 tumor inTPA-treated, and 7.7 tumors in aloe-treated mice) (p < .001).

PHOTOTOXICITYVath et al. (2002) exposed human skin fibroblast cell lines

(ATCC CRL 1634) to 1 M aloe-emodin and UVA radiation(320 to 400 nm) from two GE F40BL bulbs filtered through 3mm of soft glass. The irradiance of this source was 2.9 103W/cm2. UVA exposure alone did not affect cell survival up to ex-posures of 5 J/cm2. UVA-plus aloe-emodin reduced survival to50% at around 3 J/cm2. UVA plus aloe-emodin in D2O (extendsthe lifetime of singlet oxygen) reduced survival to 50% at around1 J/cm2. The authors suggested that this finding supports a pho-totoxic mechanism that involves formation of singlet oxygenby aloe-emodin in the presence of UVA. Cells exposed to aloe-emodin in the dark had no effect on survivability. Aloe-emodinwas shown to bind to DNA, although single strand breaks inplasmid (E. coli) DNA were not observed.

Vargas et al. (2002) exposed human red blood cells to aloe-emodin in phosphate-buffered saline (PBS) solution (20 to80 g/ml) and UVA-irradiated the cells under a Rayonet pho-tochemical reactor equipped with 16 phosphor lamps (340to 500 nm) or alternatively with an Osram HQL 250 Wattmedium-pressure Hg lamp in a pyrex immersion-well photore-actor (radiation dose of 4.5 J/cm2). Aloe-emodin caused cell ly-sis (>50%) within 70 min after irradiation of erythrocyte suspen-sion. Hemolysis did not occur in the dark controls. Free radicalscavengers reduced glutathione (GSH), and superoxide dismu-tase (SOD) had little protective effect on the photohemolytic pro-cess. Photohemolysis was decreased 20% by addition of serumproteins. Preirradiated aloe-emodin was responsible for 60%hemolysis during 80 min. When the photohemolysis tests weredone under argon there was a 54% decrease in the effect. Pho-toperoxidation was observed in irradiated aloe-emodin/linoleicacid in PBS solution. Significant amounts of hydroperoxideswere formed. When this test was performed under argon a 34%reduction in lipid peroxidation was seen.

The effect of 5 106 M aloe-emodin on acetyl-cholinesterase (ACE) activity on human erythrocyte membranesirradiated with UV light (340 to 500 nm) was determined. ACEactivity was inhibited by 50% with respect to controls; other frac-tions were incubated with reactive oxygen scavengers butylatedhydroxyanisole (BHA) (1 mM), sodium azide (1 mM), diazabi-cyclooctane (DABCO) (1 mM), and SOD (0.01 mg/ml). TheACE activity recovered by 34.1 5.4%, 21.0 4.8%, 21.9 3.8%, and 3.8 3.0% (not significant), respectively. The au-thors thought the radical species and singlet oxygen productionand subsequent peroxyl radical formation are involved in thephotoinhibitory effects of aloe-emodin on ACE activity (Vargaset al. 2002).


Shiseido Safety Research Labs (2002) reported photoaller-genicity of Aloe Arborescens CRS was evaluated by Adjuvantand Strip test method using female Hartley strain albino guineapigs (five subjects per group). During the induction phase of theexperiment, 0.1 ml of emulsified Freunds complete adjuvant(FCA) was prepared and injected intradermally at 4 corners ofa clipped and shaved nuchal area of the subject. Then, 0.1 ml of3% Aloe Arborescens CRS in distilled water was applied to thearea and irradiated with 10.2 J/cm2 of UVA. Similar applicationand irradiation continued for 4 days, and control groups weretreated in the same manner, only using distilled water. During thechallenge phase of the experiment (followed a 2-week restingperiod), 0.02 ml of Aloe Arborescens CRS6%, 3%, 0.6%, and0.3% in distilled waterwere applied to a clipped and shavedflank symmetrically in duplicate. One side was irradiated with10.2 J/cm2 of UVA, and the other side was covered with alu-minum foil to prevent exposure to the light. The reactions withrespect to erythema, eschar formation, and edema were scoredat 24, 48, and 72 h and followed a scoring system to evaluate.

It was concluded that Aloe Arborescens CRS does not pos-sess photosensitizing potential under the test conditions sinceno reactions were observed in the animals of both the treatedand control groups. One animal in the control group, however,was excluded from the estimation due to persistent light reaction(Shiseido Safety Research Labs 2002).

Shiseido Safety Research Labs (2002) reported on anotherstudy analyzing skin phototoxicity of Aloe Arborescens CRS inHartley strain male albino guinea pigs. There were five subjectsin each of the treatment and control groups. The test materialfor the treatment group was 100% and 3% Aloe ArborescensCRS in distilled water and for the control group it was 0.02%8-methoxypsoralen (8-MOP) in ethanol. The dorsum of the sub-jects was shaved and depilated, and 24 h later, 0.02 ml of the testmaterials was applied to the hairless area symmetrically in du-plicate. One side was irradiated with 14 J/cm2 of UVA, and theother side was covered in aluminum foil to prevent exposure tothe light. The skin reactions with respect to erythema and edemawere evaluated at 24, 48, and 72 h after irradiation with a scoringsystem. There were no skin reactions for Aloe Arborescens CRSat the irradiated and unirradiated sites. However, the assessmentscore of 8-MOP (positive control) at concentration 0.02% wasseverely phototoxic. The Shiseido Safety Research Lab con-cluded that Aloe Arborescens CRS does not possess phototoxicpotential under the test condition.

CLINICAL ASSESSMENT OF SAFETY

Dermal Irritation/SensitizationThe Institut DExpertise Clinique (2000) applied Aloe Bar-

badensis Leaf Water to the back of 10 healthy female adults. Thetest substance (0.02 ml) was applied to the back on a 50-mm2area of skin and occluded for 48 h. No reactions or pathologicalirritation were observed other than a very slight erythema in oneperson.

Photoallergy/PhotosentizationIvy Laboratories (1987a) applied a 0.1% Aloe extract (suntan

product) to the lower back of 25 healthy Caucasian subjectswith no history of sun sensitivity. Each subject underwent threephases: pretesting, induction, and challenge. In the pretestingphase the minimal erythema dose (MED) was established onthe midback (1-cm diameter) in 25% increments from thexenon arc solar simulator (150-watt compact xenon arc sourceequipped with UV-reflecting dichroic mirror and 1-mm-thickSchott WG-320 filter to produce simulation of solar UVBspectrum 290 to 320 nm). The MED was the area with distincterythema after 20 to 24 h.

The induction phase involved application of (10 l/cm2) ofaloe extract to a 2 2-cm section of the lower back and coveredfor 24 h. The site was then wiped and exposed to the MED andleft open for 24 h and occluded for another 24 h. The patcheswere removed and exposed to 3 MEDs of solar simulator radi-ation. This sequence was repeated for the same test sites twiceweekly for 3 weeks. The challenge phase occurred 10 to 14 daysafter the last induction exposure. The aloe was reapplied to thesame area and in the same manner but in duplicate to a newlydesignated skin site of the same dimensions on the opposite sideof the lower back and occluded for 24 h. Each site was irra-diated with 4 J/cm2 of UVA (UVB was filtered from the solarsimulator with a 1-mm-thick UG5 filter). The duplicate site wasunirradiated; another skin site untreated with aloe was exposedto 4 J/cm2 of UVA. No adverse reaction at any phase of theexperiment was seen (Ivy Laboratories 1987a).

Ivy Laboratories (1987b) applied a suntan preparation con-taining 0.1% aloe extract to the lower back of 10 healthy,Caucasian adults with fair skin (skin type I: always burns eas-ily, never tans; II: always burns easily, tans minimally; andIII: burns moderately, tans gradually). All subjects were overthe age of 18, with no medical or dermatological illness. Aloe(50 l) was applied to a 2 2-cm area in duplicate, allowedto dry, and covered. Six hours later one site was exposed to30 J/cm2 of UVA radiation. An adjacent skin site was treatedwith vehicle (hydrophilic ointment USP) and exposed to theUVA. Reactions were graded immediately, and at 24 and 48hours.

No abnormal reactions were observed. One subject had min-imal erythema reaction immediately. One individual had a min-imal erythema to the vehicle control and the aloe immediatelythat persisted at 24 h and cleared by 48 h (Ivy Laboratories1987b).

Ivy Laboratories (1988) applied a 0.01% Aloe BarbadensisFlower Extract (suntan product) to the lower back of 25 healthyCaucasian subjects with no history of sun sensitivity. Each sub-ject underwent three phases: pretesting, induction, and chal-lenge. In the pretesting phase the minimal erythema dose (MED)was established on the midback (1-cm diameter) in 25% incre-ments from the xenon arc solar simulator (150-watt compactxenon arc source equipped with UV-reflecting dichroic mirrorand 1-mm-thick Schott WG-320 filter to produce simulation of

ALOE 37

solar UVB spectrum 290 to 320 nm). The MED was the areawith distinct erythema after 20 to 24 h.

The induction phase involved application of aloe extract,10 l/cm2, to a 2 2-cm section of the lower back and coveredfor 24 h. The site was then wiped and exposed to the MED andleft open for 24 h and occluded for another 24 h. The patcheswere removed and exposed to 3 MEDs of solar simulatorradiation. This sequence was repeated for the same test sitestwice weekly for 3 weeks. The challenge phase occurred 10 to14 days after the last induction exposure. The aloe was reappliedto the same area and in the same manner but in duplicate toa newly designated skin site of the same dimensions on theopposite side of the lower back and occluded for 24 h. Each sitewas irradiated with 4 J/cm2 of UVA (UVB was filtered from thesolar simulator with a 1-mm-thick UG5 filter). The duplicatesite was unirradiated; another skin site untreated with aloe wasexposed to 4 J/cm2 of UVA.

No adverse reactions were noted, except for mild to moderateerythema, scaling, and tanning (Ivy Laboratories 1988).

The Consumer Product Testing Co. (1996) applied a 0.1%Aloe extract (suntan product) to the lower back of 26 healthyCaucasian subjects with no history of sun sensitivity. Each sub-ject underwent three phases: pretesting, induction, and chal-lenge. One subject dropped out for personal reasons. In thepretesting phase the minimal erythema dose (MED) was es-tablished on the midback (1-cm diameter) in 25% incrementsfrom the xenon arc solar simulator (150-watt compact xenonarc source equipped with UV-reflecting dichroic mirror and 1-mm-thick Schott WG-320 filter to produce simulation of solarUVB spectrum 290 to 320 nm).

The induction phase involved application of aloe extract(0.2 g) to a 3/4 3/4-inch area of the lower back and cov-ered for 24 h. The site was then wiped and exposed to the MEDand left open for 24 h and occluded for another 24 h. The patcheswere removed and exposed to two times the MED as determinedfor each subject. This sequence was repeated for the same testsites twice weekly for 3 weeks. The challenge phase occurred10 to 14 days after the last induction exposure. The product wasreapplied to the same area and in the same manner but in du-plicate to a new designated skin site of the same dimensions onthe opposite side of the lower back and occluded for 24 h. Eachsite was irradiated for 3 min with UVA (UVB was filtered fromthe solar simulator with a 1-mm-thick UG5 filter). The dupli-cate site was unirradiated; another skin site untreated with theproduct was exposed to 4 J/cm2 of UVA.

No adverse reactions at any phase of the experiment wereobserved (Consumer Product Testing Co. 1996).

Ivy Laboratories (1996) applied a 0.5% Aloe extract (othersuntan product) to the lower back of 31 healthy Caucasian sub-jects with no history of sun sensitivity. Each subject underwentthree phases: pretesting, induction, and challenge. In the pretest-ing phase the minimal erythema dose (MED) was establishedon the midback (1-cm diameter) in 25% increments from thexenon arc solar simulator (150-watt compact xenon arc source

equipped with UV-reflecting dichroic mirror and 1-mm-thickSchott WG-320 filter to produce simulation of solar UVB spec-trum 290 to 320 nm). The MED was the area with distinct ery-thema after 20 to 24 h.

The induction phase involved application of 80 mg of theproduct to a 2 2-cm area of the lower back and covered for24 h. The site was then wiped and exposed to the MED and leftopen for 24 h and occluded for another 24 h. The patches wereremoved and exposed to 3 MEDs of solar simulator radiation.This sequence was repeated for the same test sites twice weeklyfor 3 weeks. The challenge phase occurred 10 to 14 days after thelast induction exposure. The product was reapplied to the samearea and in the same manner but in duplicate to a new designatedskin site of the same dimensions on the opposite side of the lowerback and occluded for 24 h. Each site was irradiated with 4 J/cm2of UVA (UVB was filtered from the solar simulator with a 1-mm-thick UG345 filter). The duplicate site was unirradiated;another skin site untreated with the product was exposed to 4J/cm2 of UVA.

A total of 25 subjects exhibited, at most, mild erythema,desquamation, and tanning at any phase of the experiment. Sixsubjects dropped out of the study for personal reasons (Ivy Lab-oratories 1996).

KGL Inc. (2000) applied a product containing 0.1% aloe ex-tract to the lower back of 26 Caucasian adults with fair skin (skintype I: always burns easily, never tans; II: always burns easily,tans minimally; and III: burns moderately, tans gradually) overthe age of 18 with no medical or dermatological illness and nohistory of sun sensitivity. The subjects underwent three phases:pretesting, induction, and challenge. In the pretesting phase theminimal erythema dose (MED) was established on the midback(1-cm diameter) in 25% increments from the xenon arc solar sim-ulator (150-watt compact xenon arc source equipped with UV-reflecting dichroic mirror and 1-mm-thick Schott WG-320 filterto produce simulation of solar UVB spectrum 290 to 400 nm).The MED was the area with distinct erythema after 20 to24 h.

The induction phase involved application of 40 mg of theproduct to a 2 2-cm area of the lower back and covered for24 h. The site was then wiped and exposed to the MED andleft open for 48 h and occluded for another 24 h. The patcheswere removed and exposed to 3 times the MED as determinedfor each subject. This sequence was repeated for the same testsites twice weekly for 3 weeks. The challenge phase occurred 12days after the last induction exposure. The product was reappliedto the same area and in the same manner but in duplicate to anew designated skin site of the same dimensions on the oppositeside of the lower back and occluded for 24 h. Each site wasirradiated with 4 J/cm2 of UVA (UVB was filtered from the solarsimulator with a 1-mm-thick UG5 filter). The duplicate site wasunirradiated; another skin site untreated with the product wasexposed to 4 J/cm2 of UVA.

No adverse reactions at any phase of the experiment wereseen (KGL Inc. 2000).


KGL Inc. (2001) applied a product containing 0.5% aloe ex-tract to the lower back of 26 healthy Caucasian subjects with nohistory of sun sensitivity. Subjects were of skin type I: alwaysburns easily, never tans; II: always burns easily, tans minimally;and III: burns moderately, tans gradually and over the age of18 with no medical or dermatological illness. The subjects un-derwent three phases: pretesting, induction, and challenge. Onesubject dropped out of the study for personal reasons. In thepretesting phase the minimal erythema dose (MED) was es-tablished on the midback (1-cm diameter) in 25% incrementsfrom the xenon arc solar simulator (150-watt compact xenonarc source equipped with UV-reflecting dichroic mirror and 1-mm-thick Sch

Final Report on the Safety Assessment of AloeAndongensis Extract, Aloe Ando...

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