An ethnobotanical study of medicinal plants used to treat skin diseases in northern Pakistan

Background Skin diseases are a major health concern especially in association with human immune deficiency syndrome and acquired an immune deficiency. The aim of this study was to document the ethnomedicinal information of plants used to treat skin diseases in Northern Pakistan. This is the first quantitative ethnobotanical study of therapeutic herbs utilized by the indigenous people of Northern Pakistan for skin diseases. Methods Interviews were taken to obtain information from 180 participants. Quantitative methods including fidelity level (FL), Frequency of citation (FC), Use-value (UV), Jaccard indices (JI), Family importance value (FIV), Relative frequency of citation (RFC) and Chi-square test were applied. Medicinal plants uses are also compared with 50 national and international publications. Results In this study, we recorded 106 plant species belonged to 56 floral families for treatment of skin ailments. The dominant life form reported was herb while the preferred method of utilization was powder, along with leaf as the most used plant part. RFC ranges from 0.07 to 0.25% whereas the highest FIV was recorded for family Pteridaceae. FL values range from 36.8 to 100%. The study reported 88% of new plant reports for the treatment of skin diseases. Conclusion The present study revealed the importance of several plants used to treat skin diseases by the local communities of Northern Pakistan. The available literature supported the evidence of plant dermatological properties. Plants having high UV and RFC can be considered for further scientific analysis. There is dire need to create awareness among local, government and scientific communities for the preservation of medicinal species and ethnomedicinal knowledge in Northern Pakistan. Electronic supplementary material The online version of this article (10.1186/s12906-019-2605-6) contains supplementary material, which is available to authorized users.

and psoriasis are caused by a variety of microorganisms [7]. In previous reports, it was found that wound healing, eczema, dermatitis, fungal diseases, pyoderma, scabies, and skin allergies are the largest group of skin diseases that occur in most of the countries. Most of the plants used for treating skin disorders possibly have other additional properties like anti-inflammatory, antimicrobial, anti-viral, cicatrizant, hemostatic, analgesic effects that require pharmacological confirmation [8]. In literature, various plants have been reported to be used against skin infections like wound healing, scabies, swellings, boils, etc. [9][10][11][12][13][14][15][16].
In Pakistan, the number of patients suffering from skin diseases increases every year. The majority suffer from psoriasis, followed by pigment disorder, eczema, urticaria and fungal infection [17]. Climatic conditions like hot and humid weather intensify the prevalence of skin disorders. Although the mortality for skin infection is relatively low, the infection affects the quality of life. Modern skin therapies depend on the cause of the ailment. A skin disease caused by fungal and bacterial infection is medicated using antibiotics such as tolnaftate, clotrimazole, and gentamicin. It is believed that modern therapies have many disadvantages like antibiotic resistance, allergic and adverse reactions in some patients [18]. Modern medicines are very expensive with costly treatments so an alternative approach such as herbal medication in practiced.
Ethno-medicinal studies showed that herbal medicine is an alternative therapy for treatment and control of skin ailments [19]. Herbal anti-skin medicines have many useful properties including low side effects and cost treatment with high significant efficacy [20,21]. Medicinal flora have shown a pivotal part in management of dermatological conditions [11,22], particularly communities in developing countries local communities depend on traditional medicine for their health care [23]. The World Health Organization has a deep interest in the documentation of medicinal plant knowledge from from different areas of globe [24]. Currently, the Ministry of Public Health of Pakistan is promoting the usage of therapeutic herbs in health maintenance system [25].
In Pakistan, few previous reports exist the usage of therapeutic flora in skin care [1]. Therapeutic flora usage for treatment of skin ailments are documented in the literature [26], but, no specific study exists treatment of skin diseases. Various medicinal plants are also reported worldwide usage for the cure of skin disorders [7,[27][28][29][30]. The ethnobotanical literature on medicinal usage of flora for various ailments in Pakistan were mentioned in literature [31][32][33][34][35][36][37], but no systematic ethnomedicinal study has specifically focused on skin problems in the tribal areas of Northern Pakistan.
The objective of this research work is to document and examine the diversity of therapeutic flora used for treating the skin diseases in Northern Pakistan. This research will facilitate future scientific authentication through antimicrobial, pharmacological and phytochemical studies.

Description of study area
Northern Pakistan is home to the world's largest peaks and high mountain ranges i.e., Karakorum, Alai Ranges, Kunlun, Hindukush and Tien Shan [38]. Its topography differs from rock parts in North to green plains and forest in South. These areas are rich in floral variation of therapeutic plant species [39]. This area includes Hazara division, Swat valley, Mansehra, Kaghan and some tribal areas of Northern parts (Fig.1). The area is located at 72°35'to -73°31′E and 33°50′-to 34°23′ N. The province borders Afghanistan to North Western side, Kashmir to East Punjab Islamabad capital territory to East and FATA to South. The average temperature recorded in the past was minimum in January as 1.7°C while the mean maximum was 32.41°C in June [40]. The average annual rainfall is about 1125 mm. The major tribes residing in the area include Khattak, Yusufzai, Marwat, Shinwari, Afridi, Orakzai, Mahsud, Mohmand, Abbassies, Wazir, Tareen, Mashwani, Jadoon, Tanolis, Awans, Sardars, Sheikhs and Qureshi [1]. Northern Pakistan is a hilly area and the cultivated land is not enough for sustenance [41]. Medicinal plant collection and other nontimber forest products provide an additional source of income (12%), while daily salaries and wages constitute 20%, transmittals from other areas of Pakistan and overseas (17%), and other occupations (10%) [41]. About 80% population in Pakistan is rural households and has easy access to medicinal plants.

Ethical compliance
The present study was carefully designed with strict compliance of bio-ethics and approved by the Institutional Bioethics Committee (IBC) of Quaid-i-Azam University, Islamabad, Pakistan under the approval No PT-5695. The rules for plant collection and identification were followed according to National Biodiversity Action Plan as per the guidelines of Herbarium of Pakistan (ISL), Quaid-i-Azam University, Islamabad, Pakistan. Prior to data collection, a brief group discussion was held with the participants for agreement, to tell the objectives of research and to guarantee the safety of indigenous knowledge. These practices clear the aim of research and develop confidence in participants so they give reliable knowledge without any hesitation. Initially, 200 participants were selected of them were but among them, 20 were hesitant in providing knowledge leaving a total of 180 participants for data collection. While data documentation, all participants were contacted 3 times for the authentication of the knowledge given by informants. Any deviance of the informants idea from authentic knowledge given, the information was excluded and regarded inapplicable. The data quality was ensured through proper training of data collectors, pointing out missing information, duplication of the material, and careful analysis. The data quality was ensured through proper training of data collectors, pointing out missing information, duplication of the material, and careful analysis. The few plants in the MS are listed on the IUCN red list such as Taxus wallichiana (plant #104) is endangered, Colubrina oppositifolia (#84) is critical, Aconitum chasmanthum (#79) is critical and Plantago lanceolata (#69) is vulnerable. All plants listed in this study are authorized by the biodiversity action plan and duly authenticate by ethical committee of Quaid-i-Azam University, Islamabad and then included in the MS. The native communities of the area have knowledge about sustainable use of these plants and use of these plants with care (criteria of IUCN) so that they don't get vanished and are save for next generation.

Field study and data collection
This research work focused on the use of traditional plant resources with specific reference to the treatment of skin ailments. Fieldwork was performed between April 2015 to August 2015. Collectively, 180 participants were interviewed after receiving their prior informed consent. Data was collected from native indigenous health practitioners (THPs) and local participants (female and males of altered groups of age, experiences and education levels). During field surveys, face to face interviews and semi-structured interviews were also conducted. Guided field walks were also conducted [42]. The questionnaire used for data collection includes two parts, (i) part dealing with the demographic data of participants, and (ii) part focusing on information about plants´local name, mode of administration, preparation and part of the plant used against skin diseases. Documentation of data while field survey was evaluated and organized by usage of quantitative analysis. In addition, data was compared with previously published research articles on ethnomedicinal uses of plants to validate the plants with higher medicinal values for skin diseases.

Collection identification and preservation
In the current study, therapeutic flora documented by participants was identified by their common names [43]. The plant specimens were further authenticated by a Plant Taxonomist, Professor Mir Ajab Khan (Ph.D. in Plant Systematics) at the Herbarium of Pakistan (ISL), QAU Islamabad, Pakistan. All the plants species were further authenticated through available literature [42] and compared with herbarium specimens. In addition, some plants used by the local healers were photographed. Voucher plant specimens were collected in duplicate. Herbarium specimens were deposited in Herbarium of Pakistan (ISL, Registered at Index Herbarium http://sweetgum.nybg.org/science/ih/) and voucher specimens are presented in.

Quantitative data analysis Use value (UV)
Use value is calculated to assess all probable usage of plant species. UV of plants gives a quantitative analysis for plant citation. UV tells the relative importance of plant flora recognized locally. UV was analysed according to [44].

UV ¼ u=N
Where u is the total participants stating various uses of a plant and N is whole number of participants. UV is usually (1) if the number of usages is greater, and (0) if the usage report for plants species is less. UV not deliver data on multiple or single usage of plant flora is considerably low. UV does not deliver any data on the single or multiple uses of plant species.

Frequency of citation (FC) and relative frequency of citation (RFC)
FC is used for evaluating the most prefered plants or more used plant species. RFC was analysed to intricate the knowledge of traditional flora about usage of therapeutic flora in the study site.
Where RFC is denoted by relative frequency citation, FC (Frequency of Citation) is the number of participants who stated the plant flora and N is whole number of informants [34].

Fidelity level (FL)
To analyse most prefered plant usage for the cure of a specific disease, we used (FL) index adopted by [37]. FFL indicates the importance of one species over other, to cure specific diseases. Fidelity level shows the percentage of participants who reported the use of specific plant species for a particular disease (Skin disease).
Where, Np is the number of participants that declare the usage of species for definite disease, and N is total participants that use plants as a medicines for the treatment of any given ailment [45].

Jaccard index (JI)
Jaccard index (JI) is evaluated by comparison of formarly published studies from local, regional and global level by analysing the percentage of cited plant species and medicinal usage, by using the following formula: where "a" is the number of species of area A, "b" is number of species of area B, and "c" is number of species common in A and B [46].

Chi-square test
The knowledge of medicinal species distributed between male and female participants between two age categories (36-46 and > 60 years of age) was comparatively analyzed by using Chi-square.

Diversity of medicinal plants used
Therapeutic flora, used to cure skin diseases in Northern Pakistan are documented in Table 2. The study reported 106 medicinal plant species. The main growth habit of the plant flora was herbs 62%, followed by shrubs (20%) and trees (18%). The plants belonged to 56 families. Asteraceae (10 species) and Lamiaceae (7 species) represent the most dominant family in this study site (Fig. 2). The other important families in the study included Polygonaceae (6 species), then Ranunculaceae and Rosaceae (5 species each). The least species (1%) were observed in 37 families (Fig. 2).

Plant parts used in herbal medicines
Leaves (62%) were reported to be the most frequently used plant part to prepare herbal medicine either by singly or mixes by other plant parts. Leaves were followed by roots (19 species), flowers (18 species), seeds (15 species), fruit (11 species), whole plant (8 species) and stem, bulb, latex, aerial parts contributed (1 species each) (Fig.3). A schematic representation of part used of medicinal plants is shown in (Additional file 1).

Mode of preparation
Mode of administration for herbal remedies used for treating skin diseases include decoction, infusion, powder, poultice, raw, extract, juice, cooked, paste and oil. Among various preparation methods, the powder was the most frequently used (23 species), followed by paste (19 species), decoction (16 species), extract (14 species), raw and poultice (each has 8 species) (Fig. 4). A schematic representation of the mode of utilization of medicinal plants is shown in (Additional file 1).

Used categories in skin diseases
In this study, the skin diseases were assembled into 13 groups. The skin category includes pimples, mumps, measles, wound healing, boils, skin burns, abscesses, inflammation, skin irritation, allergy, burning sensation, skin cleanser and sensation ( Table 2). In this study, the maximum figure of plant was used in handling for wound healing (34 species) followed by skin burn (11 species). Other important skin ailments treated by plant flora in the area were boils and pimples (9 species). The lowest citation reports (1%) were recorded for mumps, measles and skin irritations (Fig. 5).

Quantitative ethnobotany Value of medicinal plant
In addition to the use of questionnaires, various analytical tools were required so it could be possible to do quantification of data by cross verification of indigenous information to treat skin diseases in the study site. Species with the highest use value was Pisum sativum (Fabaceae) (UV 0.143) (

Fidelity level (FL)
FL value is calculated for handling specific ailment in this study site. We examined the disease categories to focus the most significant medicinal plant species in each category of skin ailment in terms of FL. It is analyzed for the plant species which were used to cure the most commonly reported category for

Family importance value (FIV)
The analysis of family importance value reported to Pteridaceae has the maximum FIV (26.6%), followed by Fabaceae (22.2%), Scrophulariaceae, Thymelaeaceae and Caryophyllaceae (21.6). Lowest values were observed for Cyperaceae 7.7 (Fig.6). These medicinal plants are explored equally by all the communities on a regular basis and the folk knowledge is constant.

Jaccard index (JI)
A comparison of medicinal uses of plants was made by analyzing 50 research papers from aligned countries ( Table 2). The review of the literature showed that 106 reported medicinal plant species share similar uses fluctuated from 0% [29] to 13.2% while nonsimilar usage from 3.77 [64] to 0% [70]. The lowest degree of similarity was found in the studies reported in India and South Africa on skin diseases by [5,[49][50][51] ( Table 3). The comparison was based on skin disease reports in several studies, presenting the usage of therapeutic plants for the cure of skin infections in local communities.

Chi-square test
The male participants reported more medicinal plants than women, and it could be stated that males possess more knowledge about the use of medicinal plants than women (Additional file 1). The chi-square on the number of species of plants reported by the two age categories showed important differences.  (Table 4). Preliminary in vitro screening of some of the most mentioned plants have been mentioned to validate the findings of the present study (Table 5). In spite of the wide application of active metabolic compounds for humans; they also have a health hazardous effect because of much toxins. These substances not only hamper with the growth of parasite also have lethal effects on mammalian cells (Additional file 1: Table S1). It is, therefore, important to validate the toxic effects of medicinal plant products in relation to their anti-nutritional and other side effects.

Comparison with other studies in neighbouring regions
In the present study, some plants were used alone to treat the particular diseases, while in some cases plant parts were mixed to treat diseases. This present study reported 63 novel plants for skin diseases from Northern Pakistan, including Ajuga integrifolia, Anaphalis chitralensis, Capparis himalayensis, Gnaphalium affine, Isodon rugosus, Tamarix aphylla, Nepeta clarkei, Launaea nudicaulis, Valeriana jatamansi (Table 2).

Discussion
This study was carried out in the native groups of Northern Pakistan. People use medications for the cure of several diseases. Generally the medicinal plants are used in village parts of the area. The majority of professional healers in this study were males, this finding is similar to the literature [290]. According to an estimate, 84% of the rural population relies on herbal traditional medicinal plants [291]. Different origins of the medicinal plant knowledge were recorded. The inherited knowledge of medicinal plants is transferred through orally a cultural practice common in the rural areas in addition to the divine revelation. Most people inherit traditional knowledge from their elders that passed generation to generation [292].
The most dominant life form uses in the study was herbs. Herbs are easily available and collected from roadsides and farmlands [293][294][295]. Asteraceae was the most preferred family used. Previous work [3] also reported Asteraceae (6 species), Lamiaceae (6 species) and Fabaceae (5 species) with large figure of medicinal flora. There seems to be a tendency for a few families of plants to stand out in any pharmacopeia [296]. These plant families have been reported with high pharmacological,     organoleptic and pharmaceutical properties [297]. The fewer species were observed in 37 families that are similar to previous studies [298,299].
Among the reported plant part leaves were the most used plant part. In various studies, leaves were reported to be used as powder and paste on the affected skin
Acute toxicity [273] 98. Saxifragaceae Bergenia ligulata Engl. LI 17 Bergenin, catechin, gallicin and gallic acid [274] Radical toxicity in renal epithelial cell [275] 99 Flavonoids, alkaloids and tannins [283] Less toxic [284] 104. Taxaceae  Taxus wallichiana Zucc.  LI 100 Diterpenoids, lignans, flavonoids, steroids and sugar derivatives [285] Hepatotoxicity [286] 105. Thymelaeaceae Daphne mucronata S Royle LI 39 Coumarins, flavonoids, triterpenoids, lignin, glucosides, daphnine and umbelliferone [287] Leaf extract is highly toxic [287] 106. Urticaceae Urtica dioica L. LI 104 Phytosterols, saponins, flavanoids, tannins, hydrolysable tannins, phenolic compounds, proteins and amino acids [288] Nontoxic [289] areas [300]. The powder was found to be the most preferred method of utilization. The use of powder and decoction is the major mode of utilization in the herbal preparations in the ethnomedicinal studies by [35,301]. The preparations were applied 2-3 times daily until healing occurred. A large number of herbal preparation involved soaking the plant material in water for a few days and taking the infusion, while few involved boiling the parts of plants and take the decoction. The drugs were usually prepared from the paste of the plant part either with water, lime water, rose water, coconut water, milk, ghee, and butter. Sometimes juice extract from fresh parts of plants was used. Treatments were done with single plant parts or a combination of different parts of the same plant. The amount of powder used to make a concoction was defined as a half, full or a quarter of a teaspoon.
In the morning, the mixtures were regularly used before breakfast or afterward dinner, for 3-7 successive days, or till the patient was completely cured. The medicinal plants described in this study for the cure of skin infections might also be utilized additionally for their phytochemical and pharmacological activities. Following reports carried out in various areas also described the common practices of medicinal species usage against the diseases of skin [22,29,48].
The overall effectiveness of the mentioned plant species in the context of curing skin ailments was calculated on the basis of the computed index called used value [40]. This species was mentioned by 21 participants. Wounds and skin burns treated by Pisum sativum showed an increase in oxygen supply as a result of increased blood pressure flow [302]. In other studies glycoprotein extracted from Pisum helped the formation of epidermis tissues [303]. The highest UV for important medicinal plants like Pisum sativum and Cynodon dactylon might be ascribed to the trends of using herbal drugs for skin diseases in the area. It is also observed that plant species that are using repeatedly are more possibly to be active biologically and have good healing properties [53]. Less available in the study site parallel to small UV e-g in case of Adiantum venustum [304].
Relative frequency of citation is applied to choose high potential medicinal plant species for future research anti-skin diseases drug development. The medicinal species that have high RFC should be further analyzed for phytochemical compounds, to recognize their active chemical components for drug discovery [305]. These findings might be considered as of greatest importance for relating and assessing study in associated hypothetical fields for upcoming drug inventory and sustainable utilization of plant species for medicinal purposes [306].
The plant species that were cited only once by a single participant were not considered for the fidelity level study. The high value of FL indicates the choice of participants to treat the specific disease [84]. These plants can be verified as significant medicinal flora on additional estimation by the help of pharmaceutical, phytochemical and biological actions. We have found the species as more significant having 80 FL% or greater.
In [292] the maximum value of FIV was documented for Juglandaceae (45%) followed by Punicaceae (44%) whereas the lowest value was noted for Vitaceae and Rubiaceae (3%) The results of present study vary from previous literature reports due to differences in climate and vegetation of area [307]. The highest percentage of FIV demonstrates that the plants of a particular family are commonly used in curing many diseases as reported by participants.
Jaccard index is used to find out the similarity of medicinal uses with previous studies carried out on skin ailments. The maximum level of resemblance was present in findings carried out in North-West Frontier Province, Pakistan and Gilgit Baltistan Pakistan on skin diseases [1,73]) with Jaccard index value 11.81 and 5.30, respectively. About 12% average similarity is reported among different areas and the study regions. The recent study represents a high level of novelty index with respect to the use of medicinal species in skin diseases and its significance in old traditional recipes [308] specified in his study work that the medicinal plants repeatedly cited must be utilized as herbal drug development. The comparison of similarities shows the significant authenticity of documented data. Similarly, the medicinal plants which are not cited in previous work should be assessed for pharmacological and phytochemical analysis for drug discovery development.
In this research, the use of medicinal plants against skin diseases were studied for the occurrence of various toxicity and phtochemicals stated in former literature (see Table 5, Additional file 1). Mostly all the species had been described previously for their one or more phytochemical important compound representing their importance in medicinal cures. In the study, phytochemical analysis on genus, Aconitum has directed to the identification of alkaloids, benzoyl mecasonine and mesaconitine [249]. Some species of Aconitum are slightly poisonous when used in the excess amount [250]. In other studies, Bergenia ciliate was reported to contain active compounds such as alkaloids, carbohydrates, cardiac glycosides, saponins, phenols, flavonoids and diterpenes [272]. Allium sativum is rich with saponins, steroids, tannins, carbohydrates, allicin and cardiac glycosides which possess essential skin diseases curing activity [132]. Alkaloids, flavonoids, phenols, saponins and diterpenes compounds of Juniperus excels also have reported skin properties [185]. High consumption of flavonoids and phenolics may inhibit enzyme activity and cause oxidative damage [309]. Some alkaloids can inhibit enzyme activity, block ion channels loss of coordination, convulsions, hallucination and even death [310]. Myrsine Africana reported to have an acute toxic effect and Malus pumila cause hepatotoxicity [148], Rubus fruticosus damage cell activity that was stated by [261]. Discovery of drugs from medicinal plants links a multidisciplinary approach to joining pharmacological, botanical, ethnomedicinal and natural methods. Some natural products of plant derivatives are in the phase of the trial and are in experimental use [311]. Therefore further pharmacological, ethnomedicinal and phytochemical studies should be carried out to authenticate the use of plant species in skin diseases and to discover new drugs.
The root of Butea monosperma was reported for skin diseases in the present study while it is reported as a blood purifier and skin diseases in the work of [312]. Coriandrum sativum was used to control hypertension, joint pain, stomach complaints, and Gastrointestinal tracts problems [313], but in the present study, it is reported to treat pimples and skin problems. Fruits of Lagenaria siceraria were reported to treat severe body pain [314], while our study revealed that fruits and seeds can be used for skin problems. The leaves of Justicia adhatoda have been used for muscular pains in a study of [315], but this study documented that the leaves can be used for wound healing. Leaves of Myrsine africana were reported for stomach problems in the previous studies of [313], these results are in accord with the present study. The flowers and leaves of Verbascum thapsus were used for wounds [314], while the current study found that aerial parts of plant's may be utilized for the cure of blemishes and several skin related problems.
Launaea nudicaulis and Gnaphalium affine were used often for skin ailments. Asteraceae are generally rich in flavonoids, sesquiterpenes, diterpenes, triterpenes, phytosterols [146]. Nepeta clarkei, Ajuga integrifolia, and Isodon rugosus were used for curing of boils, wound healing and skin problems, respectively. Capparis himalayensis was used for wound healing in areas of Northern Pakistan. The medicinal use of species related to wound healing was not reported earlier. Euphorbia helioscopia was reported for the treatment of cholera, jaundice, respiratory diseases, cancer [46], but the present study reported it for wound healing. Brassica juncea was found to treat some skin problems while the literature suggested it for the treatment of ulcers [316]. In this study, Cucumis melo was used to treat skin burn while in a previous study it was used to treat liver diseases [314]. This study showed that Rheum emodii can be used for skin ailments, while in literature it is mostly reported for the treatment of cancer [317]. Our research also found that Swertia alata, as used for skin diseases, while the previous study reported it only as used for rheumatic disorders [314]. Onosma hispida was documented to treat skin burns, compared to use as skin tonic [318]. Verbascum thapsus also served for curing skin ailments, while traditionally it was reported for stomach diseases [319]. Melia azedarach was found as a treatment for pimples and wound healing, but literature reported this species for sexual problems and as skin tonics [320]. The present work therefore suggest that public sector administrator in study area should make policies in order to protect people from health problems and use of medicinal plants by local people for treatment of diseases.

Conclusions
This is the first quantitative ethnomedicinal study that provides information about the use of 106 species that belonging to 90 genera and 56 families for the treatment of skin diseases in Northern Pakistan. Key findings of the study revealed leaves to be the most used plant parts (58%), herb to be dominant life form (63%) and powder to be the most frequent method of administration (22%). The highest skin disease category was recorded for wound healing (40%). RFC ranged from 0.07 to 0.25%, highest use-value reported for Pisum sativum (0.143 UV), highest FIV was observed for Pteridaceae (26.6 FIV) while FL values ranged from 100% to 36.8. The medicinal information documented in this study could be explored in the future for phytochemical and pharmacological investigations which may lead to plant-based nano-medicine drug discovery and development.

Additional file
Additional file 1: Table S1. Chi-square test χ 2 test for gender wise distribution. Figure S1. Schematic representation of medicinal plant parts used prepared by NVivo software for skin diseases in Northern Pakistan.