yan2r2021 09:04 ‘AplcagSes de bromelaina de residuos de abacaxi em acne Try out PMC Labs and tell us what you think. Learn More. ELSEVIER Saudi Journal of Guide for Authors | About | Explore tht Journal Ci Saudi. Biol Sch. 2021 Jan; 28(1): 1001-1008, PMCID: PMC7785454 Published online 2020 Nov 11. do: 10,1016) sbs 2020.11 032 PMID: 33424393 Applications of bromelain from pineapple waste towards acne ‘Sukaina Abbas,* Tejashree Shanbhag, and Amruta Kothare, Departamento de Ciéncias da Vida, Kishinchand Chellaram College, Churchgate, Mumbai, Maharasthra 400- 020, India ‘Sukaina Abbas: suk24me@gmail.com * Autor para correspondéncia, suk24me@gmail,com Recebido em 23 de julho de 2020; Revisado em 26 de outubro de 2020; Aceito em 4 de novembro de 2020. Copyright © 2020 Os Autores Este 6 um artigo de acesso aberto sob a licenga CC BY-NC-ND (hitp:licreativecommons.orglicenses/by-nc- ndi4.00) Abstract Bromelain is a proteolytic mixture obtained from pineapple (Ananas comosus (L. Mert)) It has. diversified clinical properties and is used in alleviation of cancer, inflammation and oxidative stress, The current study focuses on extraction of bromelain from different parts of pineapple such as core, crown, fruit, peel and stem. The extracted enzyme was precipitated using ammonium sulphate at 40% saturation followed by dialysis. The fold of purification obtained for peel, crown, core, fruit and stem were found to be 1.948, 1.536, 1,027, 1.989, and 1.232 respectively. Bromelain activity was e: using Azocasein assay, the highest activity was seen in peel at 3.417 U/ug. Antimicrobial activity and MIC of the bromelain purified and crude fractions was studied against the test organisms. Peel crude and purified extract exhibited highest inhibitory effect towards S. aureus followed by P. acne. The antioxidant activity was evaluated using DPPH antioxidant assay. ICS0 values peel, fruit, stem and. crown are found to be 13.158 yigiml, 24.13 jigiml and 23.33 jig/ml and 113.79 igiml respectively. The purified bromelain from peel, stem and crown was used to ereate a facewash formulation towards pathogens frequently associated with skin infections. Common skin pathogens like S. aureus and P. acne were found highly sensitive to its action, The aim of this study was to evaluate the potential of bromelain isolated from waste parts of pineapple in alleviation of acne due to its diverse antimicrobial properties. imated Keywords: Bromelain, Acne, Antioxidant, Antimicrobial, Formulation 1. Introduction Pineapple (Ananas comosus) has been used as a traditional medicine by several cultures throughout time and bromelain has been established since 1876. Bromelain gets its properties mainly due to the presence of its sulfhydryl proteolytic enzymes, Bromelain is classified as stem bromelain or fruit bromelain depending on the origin of the protease. Bromelain is present throughout the pineapple plant hitps:lwww.nebirim.nih.gowipmetarticles!PMC7785454) ane‘13/1212021 08:04 ‘AlicagBes de bromelaina de residuos de abacaxi em acne however the concentration and composition may vary depending on the part of the fruit and its variety (Gautam et al,.2010). Extraction of bromelain makes it feasible as its bioavailability rises during ‘maturity of fruit rather than during fruit development (Maurer, 2001 Extensive studies have been done with bromelain to explore its clinical properties. Thus, finding effective extraction methods is necessary. Bromelain is found in pineapple wastes such as core, peel and leaves in reduced quantities compared to the fruit (Sriwatanapongse et al,, 2000). Various purification strategies have been discussed and developed for extraction of bromelain. It precipitates at 20% to 60% with ammonium sulphate showing highest recovery at 20%. Dialysis to be followed after ammonium precipitation to concentrate and to purify bromelain further by a cost-effective manner (Pardhi et al, 2016), Skin problems like wrinkles, acne and dry skin scemed to be solved using proteases such as papain and bromelain (Ozlen and Chatsworth, 1995). Loon et al., (2018) observed sensitivity of S. aureus against pineapple extract, Partially purified bromelain from the pineapple core also inhibited the growth of S. epidermidis and P. acne (Hidayat et al,, 2018). Bromelain has also been extensively used as a line of treatment for periodontal gingivitis and found to be effective against periodontal pathogens namely, S. mutans, A. actinomycetemcomitans, P. gingivalis (Praveen et al,, 2014). Edema caused by post-surgical trauma also has been effectively treated by bromelain (Mackay and Miller, 2008). ‘Acne is a skin condition affecting the skin’s oil glands is a very prominent condition. It can create a lot of psychological disturbances and stress on the individual that it allects. P acne is an opportunistic pathogen that plays an important role in the growth and cause of acne. S. aureus is a part of the normal skin microbiota and is associated with skin conditions such as folliculitis and also reported to enhance the effect of other microbes in acne lesions (Kumar et al,.2016). Hence this study aims to evaluate the potential of bromelain in alleviation of acne owing to its diverse antimicrobial properties. Active bromelain was isolated from waste parts of pineapple and its eff bacterial pathogens like acne was studied, Bromelain will aid as a potent anti-inflammatory agent that will cial additive to the formulation. Using bromelain from the waste parts of pineapple will help in waste recycling as well as make the whole process cost-effective. ct as ber 2. Materials Biological material: Pineapple, Turmeric, Aloe vera gel, Chemicals: Acetic acid (Chemdyes corp.), Agar Agar Type I (Hi-Media), Ammonium Sulphate (Loba Chemie), Ampicillin antibiotic discs (Hi-Media), Ascorbic Acid (Loba Chemi), Azocasein (Sigma- Aldrich), Beef Extract (Ili-Media), Bovine Serum Albumin (Sigma-Aldrich), Cone. IICI (Chemdyes) Copper Sulphate, DPPH (Hi-Media), Folin- Ciocalteu reagent (Thermofisher scientific), Peptone (Hi- Media), Sodium Acetate Trihydrate, Sodium Carbonate, Sodium Chloride (SDFCL chemicals), Sodium Hydroxide, Sodium Potassium Tartrate, Tris Base (Hi-Media), Tris HCI (Hi-Media), Yeast Extract (Hi- Media) Microbiological Strains: S. aureus (MTCC 96), C. diphtheriae, E. coli (ACC: 25922), Pseudomonas aeruginosa (ATCC: 227853), P. acne (MTCC: 1951). 3._Methodology 3.1. Sample preparation 3.4.4. Crude extract preparation The pineapple parts were categorized as fruit, stem, peel, core and crown. They were individually homogenized with 0.1 M of sodium acetate buffer, (pH ~7.0). The extracts obtained was filtered and centrifuged at 6000 rpm for 20 min at 4 °C. These extracts were labelled as fruit crude extract, stem crude extract, peel crude extract, core crude extract and crown crude extract of bromelain, The parts are observed in Eig. hitps:lwww.nebirim.nih.govipmetarticles!PMC7785454 anoyan2r2021 09:04 ‘AplicacSes de bromelaina de residuos de abacaxi em acne en in a separate window Parts of the pineapple, Crown, stem, pec! (left to right) 3.4.2, Ammonium sulphate precipitation The crude extract was subjected to ammonium sulphate precipitation at 40% saturation (226 gil). The extraction was carried out in an ice box over a magnetic stirrer, Ammonium sulphate was gradually added pinch by pinch in a period of 30 min, It was allowed to settle for 24 h at 4 °C. The solution was then centrifuged at 6000 rpm for 10 min at 4 °C. The pellet was later reconstituted in minimum volume of 10 mM Tris, (pH —7.0) and then subjected to dialysis. ‘The supernatant obtained was further precipitation at 60% saturation (120g/1), The solution was allowed to stand for 24 h at 4 °C and then centrifuged at 6000 rpm for 10 min at 4 °C. The pellet obtained was reconstituted in minimum volume of 10 mM Tris, (pH—7.0). 3.4.3. Dialysis 3.1.3.1 Activation of dialysis membrane 7 em of dialysis membrane was added into boiling distilled water with 2% sodium carbonate for 45 min, The membrane was boiled again for 45 min in distilled water, the membrane was left overnight in Acetate buffer (pH-7.0). 3.1.3.2 Dialysis The fractions from ammonium precipitation were loaded into the activated dialysis, ‘membranes and tagged. They were equilibrated into a beaker with acetate buffer. The process was carried out for 24 h in an ice box with the replacement of butler every 6 b. The samples from ‘membranes were then unloaded and labelled as purified bromelain samples. The setup is observed in Fig.2. hitps:lwww.nebirim.nih.govipmetarticles!PMC7785454 angyan2r2021 09:04 ‘AplicacSes de bromelaina de residuos de abacaxi em acne Open in a sey indow Dialysis setup, Sample loaded in dialysis membrane (Left to right) 3.2. Estimation of protein content by Folin-Lowry assay The protease mixtures were estimated for the protein content by using Folin-Lowry assay. Bovine serum albumin (BSA) was used as a standard at | mg/ml. Freshly prepared alkaline copper sulphate solution ((50:1) 2% Na2CO3 in 0.1 N NaOH: 0.5% CuSO4 in 1% NaK), Folin-Ciocalteu Phenol reagent (FC) 1 N was used, | ml of protein was added to 5.5 ml of alkaline copper solution and incubated for 10 min, 0.5 ml of FC reagent was added to the mixture and incubated in dark for 30 min and absorbance was recorded (Ay = 660 nm). (Plummer and Plummer, 1988) 3.3. Estimation of proteolytic activity 3.3.1. Calibration curve Azocasein was used as the substrate in the concentration range of 0.1-2% Azocasein was digested using enzyme sample for 300 min and using regression analysis, an equation was derived correlating the absorbance/color intensity to the substrate concentration. 3.3.2, Proteolytic activity of samples Proteolytic activity of samples was tested varying the azocasein concentration from 0.1 to 2%. Equal quantities of substrate and enzymatic samples (i.e. 100 yl) were incubated for 10 min. The reaction was terminated using 5% Trichloroacetic acid (TCA) and the vials are centrifuged at 4 °C for 10 min, The supernatant is taken by 1:1 dilution of 0.5 N NaOH and its absorbance is read (Aq = 440 nm). The blank was obtained by mixing the TCA to the substrate prior to enzyme addition, Calculations were done as specified by Coslho etal. (2016) 3.4, Antimicrobial activity testing 3.4.1. Bacterial maintenance S. aureus (MTCC 96), C. diphtheria, E. coli (ATCC: 25922), P aeruginosa (ATCC: 227853), P. acne (MTCC: 1951) were maintained on Nutrient Agar slants and incubated at 37 °C. These cultures are going to be further mentioned as test cultures. 3.4.2. Ditch plate technique Antimicrobial susceptibility of the organisms towards the crude extract, ‘was observed by using the ditch plate technique. Bromelain extract was mixed with molten agar and poured into the ditch, The test organism is plated across the ditch, The plates incubated at 37 °C for 24h, The Zone of Inhibition was measured (Benson et al,, 2015), 3.4.3. Minimum inhibitory concentration by turbidity test The MIC of the purified extracts of peel, fruit and stem against the test cultures in a 10-100% (v/v) dilution, Saline was used as a negative control in place of test samples. Absence of turbidity signifies no growth of culture whereas presence hitps:lwww.nebirim.nih.gowipmetarticles!PMC7785454) angyan2r2021 09:04 ‘AplcagSes de bromelaina de residuos de abacaxi em acne of turbidity signifies growth of culture (Wiegand et al, 2008). 3.5. Antioxidant activity testing by DPPH method Antioxidant activity (DPPH free radical scavenging activity): The antioxidant activity of the extracts and standards was based on the radical scavenging effects of 2, 2 diphenyl-I-pieryIhydrazyl (DPPH) The diluted working solutions of the samples were made in water. Ascorbic was used 10-100ug/ml. 0.002% DPPH was prepared in ethanol. The test sample/standard was mixed with DPPH in 1:1 ratio and incubated in the dark for 30 min. The solution mixtures were read at 517 nm, The OD was recorded and percentage inhibition was calculated by the formula below. Ethanol with DPPH (1:1) is used as a blank (Khalaf et al., 2008, Yuris and Siow, 2014. A-B Percent inhibition = x100)where A is OD of blank and B is OD o} sample. 3.6. Formulation ‘Three formulations were synthesized using the stem bromelain, peel bromelain and crown bromelain as the primary base See Table | and Fig. 3. Table 1 Formulation of Bromelain Facewash, Function Bromelain Antimicrabia/Antioxidant Turmeric Antioxidant Aloe Vera Gel Moistrizer Xantham gum ‘Thickener Tea re il Essential oilPreservative NaOH, SLES, Sodium stearate, Propylene glycol Soap Base hitps:lwww.nebirim.nih.govipmetarticles!PMC7785454 519yan2r2021 09:04 ‘AplicacSes de bromelaina de residuos de abacaxi em acne Bromelain Facewash Formulations al charac 3.6.1. Physical characteristics of the facewash Phys color, pH, washability, foamability and consistency (Ingle and Meshram, 2018). 3.6.2, Antibacterial testing of formulation The antimicrobial activity of formulation the crude extract was determined using the disc diffusion method. Nutrient agar plates were swabbed with the cultures. Sterile Whatmann discs impregnated with the test samples were placed, The samples were tested in four concentrations of (v/v) percentage at 25%, 50%, 75% and 100%. They were incubated at 37 °C for 24h, The Zone of Inhibition was measured. ( 2015) 3.7. Comparison of commercial products Four commercially available herbal facewashes were selected at random and their antimicrobial action against bacterial cultures were tested at four (v/v) % concentration levels (i.e. 25%, 50%, 75%, 100% These were named as Brand 1, Brand 2, Brand 3 and Brand 4) 4. Results and conclusion 4.1. Estimation of protein content The protein content was estimated by folin-lowry method. The values obtained in crude extracts were fruit (170 pg/ml), crown (167 jigiml), peel (128 g/ml), stem (127 pg/ml) and core (126 jig/ml). The crude extracts were purified by ammonium sulphate precipitation and dialysis. The highest protein content after purification was observered in peel (103 ig/ml), followed by stem (100 ig/ml), crown (93 ugiml) and core (92 jig/ml). The results are shown in Graph 1 hitps:iwwv.ncb.nim.nih.govipmelarticles/PMC7786454) aneyan2r2021 09:04 ‘AplicacSes de bromelaina de residuos de abacaxi em acne Protein Estimation of Bromelain Extracts 10 Core Crow Frat Pest stem coe #40 860 mines) a DiDl-0 {n_a separate window Graph 1 Estimativa do teor de proteina em extratos brutos ¢ purificadas de bromelaina 4.2, Estimativa da atividade proteolitica A atividade proteolitica foi estimada pelo método Azocasein. O contetido de azocaseina foi calibrado para digestZo maxima usando bromelaina, conforme visto emGrifico 2. Isso foi usado para o célculo das atividades proteoliticas dos extratos brutos ¢ purificados. Um aumento foi observado nas atividades proteoliticas apés a purificagdo. A casca ¢ a fruta demonstraram maior atividade proteolitica a 3,417 U / jig ¢ 2,556 U / pg. Atividades proteoliticas foram descritas emGrifico 3 ¢ mesa 2. 08 y = 0.0555x + 0.1512 R?= 0.9975 sss aaa 0.3 ABSORBANCE (OD) ° ° ey B ° 3 o 1 2 3 4 5 6 7 8 9 0 U CONCENTRATION OF AZOCASEIN (MG/ML) Abra em uma janela separada Grifico2 Gritico de calibraglo da Azocasein. hitps:lwww.nebirim.nih.govipmetarticles!PMC7785454 m9yan2r2021 09:04 ‘AplicacSes de bromelaina de residuos de abacaxi em acne ‘ane ey Cr Crown © care Fat 5. Som 40-40% sm. Sp saturation / © - 6m sam, Sup Seuiaion 40D = 40%. saturation — subject to days oD — 60% saturation — ‘subjctado dass Grifico 3 Atividade especifica exibida pelas frag de Bromelaina pelo ensaio Azocasein. mesa 2 ‘Soma do ensaio da azocaseina. “Atividade especifica do Atividade especifica da fragio Purificagio Rendimento petréleo (U /ug) purificada (U /ug) dobrada _percentual (%) Casa 1,758 3417 1988 12.19 coma 1.258 1.933 1.336 105,483 ‘Testemunho 1.615 Lass 1.027 102.435 Fra 1.285 2556 1989 1149032 ‘Tronco 1.798 2216 1232 108.627 4.3. Estimativa de teste de susceptibilidade antimicrobiana de extratos de bromelaina teste de susceptibilidade antimicrobiana dos extratos brutos e putificados de bromelaina foi realizado pelo método da placa vala, O extrato bruto e purificado da casca exibiu maior efeito inibitério para S. aureus seguido por P. acne, E. cali, C. diphtheria e P, aeruginosa, P, aeruginosa foi mais suscetivel & ago da bromelaina, De acordo com a agdo inibitéria, os extratos potentes foram casca, fruta, caule, coroa e carogo. Os resultados sto mostrados emGirificos 4, 5 e Fig 4. hitps:lwww.nebirim.nih.govipmetarticles!PMC7785454 aneyan2r2021 09:04 ‘AplicacSes de bromelaina de residuos de abacaxi em acne Grifico 4 Atividade antimicrobiana exibida por extrates brutos de bromelaina, Grifico S Atividade antimicrobiana exibida pela Bromelaina puri hitps:lwww.nebirim.nih.govipmetarticles!PMC7785454 sngyan2r2021 09:04 ‘AplicacSes de bromelaina de residuos de abacaxi em acne ‘Uma petriplaca mostrando sensibilidade antimierobiana por Bromelaina por téenica de placa Ditch, 4,4. Estimativa da concentragao inibitéria minima (MIC) por teste de turbidez A concentragio inibitéria minima dos extratos purificados foi estudada contra as culturas de teste. P. acne foi considerada suscetivel contra os fracionados purificados em concentragées téo baixas quanto 19 1g / ml do extrato da fruta, Enquanto P. aeruginosa e C. diphtheria s3o suscetiveis, eles requerem maior concentragdo da enzima para inibir seu crescimento. As observagdes do teste de turbide mostradas emTabela 4. Os valores MIC so mostrados emGrifico abela 34 hitps:lwww.nebirim.nih.govipmetarticles!PMC7785454 1019yan2r2021 09:04 ‘AplicacSes de bromelaina de residuos de abacaxi em acne MIC (ug/ml) 9a 0.48 we | 2 3856 3618 #9 # fog 2g & a 5 ef 8G 2 = goa ze Feit stem ‘Abra em uma janela separada Resultados de MIC de extratos contra culturas. hitps:lwww.nebirim.nih.govipmetarticles!PMC7785454 wmneyan2r2021 09:04 ‘AplicacSes de bromelaina de residuos de abacaxi em acne Tabela 3 Resultados de MIC de extratos v/s cultura, Papel Organismo MIC (ug/ mi) Casca Caiferia 45 Eeeoli 22,5 Pacne 30 Pacrurginosa 52,5 Scaweus 15 Fruita C. diphtheriae 67,998 E.coli 19.428 Pacne 19.428 P.acrurginasa 87.426 S.aweus 38856 ‘Tronco diphtheriae 96,48 E.col 4824 Pacne 36,18 Pacrurginasa 9648 Saureus 48,24 hitps:lwww.nebirim.nih.govipmetarticles!PMC7785454 raneyan2r2021 09:04 ‘AplicacSes de bromelaina de residuos de abacaxi em acne Tabela 4 Observagdes do teste de turbidez MIC. Extrato Purificado Peet Organismo 100% 90% 80% 70% 60% 50% 40% 30% 20% 10% C diphtheria - coe RR Eco = eee ee ee Pane = coe P.acroginosa ~ coe RR RR Scaweus = Boe Extrato Purificedo de Frutas Organismo 100% 90% 80% 70% 60% SO% 40% 30% 202% 10% diphtheria - coe eR RR Foot nes Pane es P.aeroginosa ~ oe RR eR Re RS Scaweus —- coe ee ee Extrato Purificedo de Caule Organism 100% 90% 80% 70% 60% 50% 40% 30% 20% 10% C diphiheria ~ coe ot Re eR et Eco = coe Pane es Pacruginosa es Sauweus = coe ‘Abra em uma janela separada 4.5. At idade antioxidante 0s antioxidantes naturais estio presentes na bromelaina que auxiliam no combate ao estresse oxidative. As atividades antioxidantes de varias amostras de bromelaina sio significativamente diferentes, (ANOVA de uma via; 4-0,05; valor P - 0,703). Os valores de IC50 das amostras de bromelaina de casca, fruta, caule ¢ coroa sdo 13,158 ug / ml, 24,13 jig/ml e 23,33 pg/ml € 113,79 yg / ml. A atividade antioxidante da bromelaina é comparada com 0 dcido ascérbico (usado como padrio). 1C50 de dcido ascérbico sendo 9,92 ug / ml. A porcentagem de inibigao das amostras de bromelaina & mostrada emGrifico 7 ¢ Tabela 5. hitps:lwww.nebirim.nih.govipmetarticles!PMC7785454 1319yan2r2021 09:04 ‘AplicacSes de bromelaina de residuos de abacaxi em acne 100 90 80 z, z 7% z 60 = & 50 £40 Zz a Z 30 a = 20 10 0 0 20 40 60 80 CONCENTRATION (V/V)(%) —eAscorbic Acid —m+Peel —t-Fruit Stem Crown Abra em uma janela separada Grifico 7 Atividade antioxidante dos extratos de deido asedrbico e bromelaina, Tabela 5 Valores 1C30 exibides pelas amostras testadas. ‘Amostra Valor 1C50 ‘Acido ascérbico 9,92 ye /ml Casea 13,158 g/ml Fra 2413 pg/ml ‘Tronco 23,33 ug / ml ‘coro 113,79 g/m hitps:lwww.nebirim.nih.gowipmetarticles!PMC7785454) sanyan2r2021 09:04 ‘AplcagSes de bromelaina de residuos de abacaxi em acne 4,6. Teste de parametros fisicos de formulagées de teste Os pardmetros fisicos das formulagées de teste foram descritos em Tabela 6, Tabela 6 ObservagGes fisicas das formulagdes. Fator Formulasio Peel Formulagio de Haste Formulagio da Coroa pH 55 59 53 Cor ‘Amarelo dourado -Amarelo Acastanhado Verde pilido Consisténcia Semi-sélido Semi-sélido Semi-sélido Lavabilidade Boa Boa Boa Foambilidade Aparece pouca espuma Aparece pouce espuma Aparece pouca espuma 4,7. Resultados de suscepti bromelaina lidade antimicrobiana da formulagao do teste de ‘A susceptibilidade antimicrobiana da formulacao foi testada por difusdio em disco contra os organismos de teste. 5, aureus foi o organismo mais suscetivel pelas tr6s formulagSes de teste seguidas por P. acne, ‘A zona de inibigdo é representada graficamente, Os resultados so descritos emGrifico 8 e Fig. 5. Antineroblal testing of Formulation 2% 575% NDING 2M SOT LO STO von Eoraica Seu Feaaaion ee! Fomnaion| Cie Eco Peace Abra em uma janela separada Grifico 8 antimicrobiano de formulagSes de bromelaina para lavagem facial hitps:lwww.nebirim.nih.govipmetarticles!PMC7785454 1519yan2r2021 09:04 ‘AplicacSes de bromelaina de residuos de abacaxi em acne Abra em uma. Eigs Formulago de AST de Bromelaina: eaule, casca e coroa (de cima para baixo) contra B. coli, S. aureus, P acne e C. diphtheria (de esquerda para a dreita). 4,8. Resultados de susceptibilidade antimicrobiana de produtos comerciais ‘Todos os facewashes foram considerados eficazes contra P. acne eS. aureus. No entanto, a marca 2.€ a ‘marea 3 ndo mostraram inibigdo contra E. coli, Os resultados so mostrados emCirifico 9 e Fig. 6. bid dun ‘Abra em uma jancla separada ‘Testes antimicrabianos de anti-sépticos comercia hitps:lwww.nebirim.nih.govipmetarticles!PMC7785454 1619yan2r2021 09:04 ‘AplicacSes de bromelaina de residuos de abacaxi em acne Abra em uma janela separada Eig6 AST de formulagées comerciais contra P. aene 5. Discus: © abacaxi é uma fruta milenar, consumida e utilizada tradicionalmente por muitas de suas propriedades ‘medicinais. Tem sido usado para desbridamento da pele ( Houck etal. 1983 ). A bromelaina também & usada como substituto durante a deficiéneia de pepsina e tripsina devido & sua atividade e estabilidade ‘em uma ampla faixa de pH ( Balakrishnan et al., 1981) Geralmente, apenas a fruta é considerada como © tinico membro comestivel e 0 resto & tratado como residuo, ou seja, a casca, 0 carogo, a coroa © 0 caule, Os residuos do abacaxi sio geralmente descartados e depois biodegradaveis, o que leva & degradayo ambiental principalmente devido ao conteiido rico em carboidratos. De acordo com nosso estudo, a bromelaina ativa também foi obtida das pegas residuais. A easca demonstrou maior atividade proteolitica a 3.417 U / ng seguida pela fruta a 2.556 U / yg. Precipitagdo com sulfato de aménio didlise foram aplicadas para purificagio pela facilidade de uso, viabilidade e facil recuperagdo da cenzima com maior atividade, Esta etapa esté de acordo com ( Pardhi ct al. 2016) que validam 0 uso desses métodos para a purificagdo da bromelaina. A atividade da bromelaina foi estudada usando 0 ensaio de azocaseina. A maior atividade foi observada na casca ¢ na fruta em 3.417 U/ jig € 2.556 U/ ug respectivamente. Este método esté de acordo com ( Cod 6 ) para estimativa de atividades proteoliticas usando azocaseina. Vuris ¢ Si associou o abacaxi por suas atividades hitpsslww.ncb.nim.nih.govlpmelarticles/PMC7786454 a9‘13/1212021 08:04 ‘AlicagBes de bromelaina de residuos de abacaxi em acne antioxidantes & presenga de fendlicos, o que nos motivou a estudar se a bromelana também exibia atividades antioxidantes. Os valores IC50 de amostras de bromelaina de casca, fruta, caule ¢ coroa so 13,158 pg / ml, 24,13 yg / mle 23,33 wg / ml € 113,79 pg / ml, Acne é uma condigdo da pele que afeta as glindulas sebdceas da pele e é uma condigao muito proeminente, Pode criar muitos distirbios psicologicos ¢ estresse no individuo que afeta, A flora microbiana frequentemente associada a infec¢a0 por acne inclui P. acnes, S. epidermidis e S. aureus. A atividade antimicrobiana da bromelaina foi testada contra P. acne, S. aureus, C. diphtheria, E. coli e P. aeruginosa. S. aureusfoi o organismo mais suscetivel a ago dos extratos brutos e purificados de bromelaina, seguido por P. acne . O extrato brato ¢ purificado da casca exibiu o maior efeito inibitério para S. aureus. A maior resisténcia a atividade da bromel ina foi demonstrada por P. aeruginosa, Um facewash foi preparado usando extratos de casca, caule e bromelaina em coroa, Até onde sabemos, esta é a primeira formulago preparada com bromel ina extraida de foi duos de abacaxi. A susceptibilidade antimicrobiana da formulaga testada por difusdo em disco contra os organismos de teste. S. aureus foi o organismo mais suscetivel pelas trés formulagdes de teste seguidas por P. acne.Escolhemos quatro coletores faciais & base de ervas populares, aclamados por serem usados como anti-acne, e comparamos a ago antimicrobiana das formulagdes. As marcas 2 € 3 ndo mostraram atividade contra E. coli. . As formulagdes de bromelaina apresentaram atividade contra P. acne, S. aureus, C. diphtheria ¢ E.coli . A comparagao da bromelaina com os anti-sépticos faciais comerciais encontrou uma diferenga na atividade contra S. aureus ¢ P. acne 6. conclusées ‘A bromelaina tem se mostrado uma potencial protease que pode ser usada clinicamente para o tratamento da acne devido a sua atividade antimicrobiana, facilidade de purificagdo e extragio dos residuos do abacaxi. f um bom candidato uma vez que a prot se purificada é potente e a extragdo & vidvel. A bromelaina também possui uma ampla faixa de estabilidade em termos de temperatura ¢ pH. A propriedade antioxidante da bromelaina também contribui para o beneficio da pele no combate 20 estresse oxidative. A acne sempre foi causa de grande softimento fisico e social devido & sua natureza sensivel, & preciso ter muito cuidado com a acne para evitar que outro surto cause mais sofrimento a pele. A formulagdo da bromelaina foi a primeira a ser feita a partir de residuos de abacaxi. A bromelaina também pode ser explorada para estudé-la como um gel bactericida com tratamento direto para acne, porém sua interagao com outra flora cutinea também pode precisar ser estudada para ver se € uma protease seletiva ou ndo. As interagdes da bromelaina com outras proteases da pele e peptideos antimictobianos, como a psoriasina, também podem ser exploradas. Agradecimentos Este projeto foi financiado pelo ‘Mumbai, projeto URG - 388. Go: Biofharmaceuticals do Guru Nanak Khalsa College por sua instrumentagio sofisticada e suporte de anilise yuema de subvengdes para pesquisas menores da Universidade de iamos de estender nossa gratiddo ao National Facility for Notas de rodapé Revisdo por pares sob responsabilidade da King Saud University Referéncias 1. Balakrishnan V., Hareendran A., Nair C, Double-blind cross-over trial de uma enzima preparagao em pancreatic steatorrhoea, J. Assoc. Phys. India, 1981; 29 : 207-209. [ PubMed ] [ Google Scholar ] 2. Benson, Brown A., Smith H. 13* ed, McGraw-Hill Higher Education; Nova York: 2015. Aplicagao Mictobiolégica: Laboratory Manual in General Microbiology. [ Google Scholar ] hitps:lwww.nebirim.nih.gowipmetarticles!PMC7785454) 1819yan2r2021 09:04 ‘AplcagSes de bromelaina de residuos de abacaxi em acne 3. Coélho DE, Saturnino TP, Fernandes FF, Mazzola PG, Silveira E., Tambourgi EB Azocasein Substrate for Determination of Proteolytic Activity: Reexamining a Traditional Method using Bromelain Samples. Biomédico. Res. Int. 2016; 2016 : 8409183. doi: 10.1155 / 2016/8409183. [ Antigo gratuito PMC ] [ PubMed ] [ CrossRef] [ Google Scholar ] 4. Gautam, SS, Mishra, SK, Dash, V., Goyal, AK, Rath, G., 2010. Estudo comparativo da extragdo, Purificagao e estimativa da bromelaina do caule e fruto da planta do abacaxi, Thai J. Pharm. Sci. 5. Hidayat, Y., Hermawati, E., Setiasih, S., Hudiyono, S., Saepudin, F., 2018. Teste de atividade antibacteriana da bromelaina parcialmente purificada do extrato de niicleo de abacaxi (Ananas ‘comosus [L..] Mert) por fracionamento usando acetona de sulfato de aménio, AIP Conf. Proc. 2023. https://doi.org/10.1063/1 5064064 6, Houck J., Chang C., Klein G, Isolation of an effective debriding agent from the stems of pineapple plants, Int. J. Tissue React. 1983; 5 : 125-134. [ PubMed } | Google Scholar ] 7. Ingle, A., Meshram, MB, 2018. Formulacdo e avaliagdo de sabonete liquido ayurvé Phytopharm. 8. Khalaf, NA, Shakya, AK, Al-Othman, A. some common plants. Turco J. Biol. 9. Kumar B., Pathak R., Mary PB, Jha D., Sardana K., Gautam HK Novos insights sobre a patogénese da acne: Explorando o papel das populagdes microbianas associadas & acne. Dermatol. Pecado. 2016 doi: 10.1016 / jdsi.2015.12.004. [ CrossRef ][ Google Scholar } 10, Loon, YK, Satari, MH, Dewi, W., 2018, Antibacterial effect of abacaxi (Ananas comosus) extract para Staphylococcus aureus. Padjadjaran J. Dent. https:/doi.org/10.24198/pjd.vol30n01. 16099. 11, Mackay D., Miller A. Nutritional Support for Wound Healing, Altern. Med. Rev. 2003; 8 : 359— 377. [ PubMed ] [ Google Scholar ] 12, Maurer HR Bromelain: bioquimica, farmacologia ¢ uso médico. Célula. Mol. Life Sci. C. 2001; 58 : 1234-1245. doi: 10.1007 / P1.00000936. | PubMed | [ CrossRef ] [ Google Scholar ] 13, Orlen S., Chatsworth C. Patente dos Estados Unidos; 1995. Composigo cosmética contendo alfa Int. J :l-Agbar, Z., Farah, H., 2008. Antioxidant activity of hidroxiacidos, dcido salicilico e mistura de enzimas de bromelaina e papaina.e. [ Google Scholar ] 14, Pardhi V., Chivte P., Kothare A., Shanbhag T. An Overview of Purification and Activity of Bromelain. Int. J. Recent Sci. Res. 2016 { Google Scholar } 15, Plummer M., Plummer DY 3* ed. Teta Me-Graw Hill Publishing Company Ltd .; 1988. Introdugao & Bioquimica Prética. { Google Scholar ] 16, Praveen NC, Rajesh A., Madan M., Chaurasia VR, Hitemath NV, Sharma AM Avaliagdo In vitro da Eficécia Antibacteriana do Extrato de Abacaxi (Bromelaina) em Patégenos Periodontais. J Int, Cura oral. JIOH. 2014; 6 : 96-98. [ Artizo gratis PMC ] [ PubMed ] [ Google Scholar] 17, Sriwatanapongse, A., Balaban, M., Teixeira, A., 2000. Thermal inactivation kinetics of Bromelain in pineapple juice. Trans. Sou, Soe. Agric. Eng. https://doi.org/10.13031/2013.3071. 18, Wiegand, I, Hilpert, K., Hancock, REW, 2008. Métodos de diluigo em agar e caldo para determinar a concentragao inibitéria minima (MIC) de substancias antimicrobianas. Nat, Protoc. Ittps://doi.org/10,1038/nprot.2007.521. [ PubMed } 19, Yuris A., Siow L.+F. Um estudo comparativo das propriedades antioxidantes de trés variedades is comosus L.). J. Food Stud. 2014 doi: 10,5296 /jfs.v3i1.4995. [ CrossRef ] Os artigos do Saudi Journal of Biological Sciences so fornecidos aqui como cortesia da Elsevier hitps:lwww.nebirim.nih.gowipmetarticles!PMC7785454) 1919