Plant cell culture as emerging technology for production of active cosmetic ingredients

Abstract

Plants have always been the main source for active cosmetic ingredients, having proven health beneficial effects on human, such as anti‐aging, antioxidant, anti‐inflammatory, UV‐protective, anti‐cancer, anti‐wrinkle, skin soothing, whitening, moisturizing, etc. Extracts from herbal, aromatic and/or medicinal plants have been widely used as effective active ingredients in cosmeceuticals or nutricosmetics, especially in products for topical application and skin‐care formulations. However, over the past decade, there has been an increasing interest to plant cell culture – derived active cosmetic ingredients. These are “new generation” of high quality natural products, produced by the modern plan biotechnology methods, which usually showed stronger activities than the plant extracts obtained by the classical methods. In this review, the advantages and the current progress in plant cell culture technology for the production of active cosmetic ingredients have been summarized, and discussed in details within a presented case study for calendula stem cell product development.

Abbreviations

CMC

cambial meristematic cells

EGF

Epidermal Growth Factors

1. Introduction

Plants are the oldest source of natural compounds with medicinal and cosmetic properties explored by the mankind. Even nowadays, between 70 and 80% of the people worldwide relay on traditional herbal medicine to cover their basic healthcare needs 1. In fact, 11% of the existed essential drugs used in modern human medicine are still with plants origin 2. The percentage of plant‐derived natural products, applied as active ingredients in cosmetics, is much higher. In accordance with the continuous increase of world population, the global demand for natural plant ingredients is expected to continue its exponential growth 1. As result, we are already witnesses of mass overexploitation of natural habitats of some herbal and aromatic plants, placing them under threat or even leading to their extinction 3. This negative tendency is especially dangerous for medicinal plants, which in most cases are rare and endemic species. These species often grew under extreme climate, unique soil composition or specific latitude. Their populations usually have been characterized with low levels of genetic diversity, which makes these species extremely vulnerable to genetic erosion and decrease their chance to survive in case of environmental pollution 4. Moreover, plant derived active ingredients are usually very complex mix of bioactive molecules, which cannot be easily replaced by chemically synthesized analogs. To solve the above problems, plant biotechnology has provided the tool, which can secure more eco‐friendly and sustainable supply of valuable phytochemicals with health‐beneficial properties, to cover the growing demands of cosmetic industry. Over the last decade, the advantages of plant cells and tissues culture technologies have been widely explored in development of highly‐efficient platforms for more rapid production of pharmaceutically important molecules of plant origin or for heterologous expression of therapeutic proteins 5. Nowadays, there are many cosmetic products, including both cosmeceuticals and nutricosmetics, which have active ingredients, derived by plant cell culture technology. The aim of this paper is to review the recent progress in plant cell technology for cosmetic application and to provide short overview on commercialized plant cell culture – derived active cosmetic ingredients available on the market. The information in this review was retrieved by an exploratory electronic survey of plant tissue culture ‐ derived cosmetic ingredients and their technical specification datasheets, conducted by search in the technical websites of the major suppliers such as Prospector® search engine, SpecialChem, Cosmetic Design Europe, the academic search engines Scopus, ScienceDirect, Google Scholar, and PubMed, and the popular search engine Google. In addition, the main steps in development and characterization of plant cell culture – derived active ingredients are closely discussed in the presented case study for development of InnovaStemCell Calendula product.

2. Plant extracts as active ingredients in cosmetics

Plants are rich source of countless metabolites with potential application in cosmetic products. Since time immemorial, the mankind has used different plants and their extracts to create cosmetics with the aim to establishing a state of eternal youth 6. Nowadays, the plant extracts are becoming the most popular active ingredients of cosmetics due to the ever‐increasing demand for natural compounds which in addition to esthetic looks can provide additional health benefits. Development of such products, known as “cosmeceuticals”, reflects a most recent trend in the modern cosmetics and personal care industries. Indeed, plants are rich in endogenous bioactive metabolites with potential cosmetic and pharmaceutical applications 7, 8, 9. Most of these phytochemicals, such as polyphenols, phenolic acids, triterpenes, flavonoids, stilbenes, steroids, carotenoids, steroidal saponins, sterols, fatty acids, polysaccharides, sugars, peptides, etc., could be extracted with appropriate solvents and used as active ingredients present in cosmetic formulations 10. Because of this, huge numbers of plant sources have been explored by the cosmetics industry in search of innovative active ingredients which combine some specific pharmacological properties, such as antioxidant, antimicrobial, antiviral, anticancer, antifungal, anti‐inflammatory, anti‐allergy, etc., and also showing strong moisturizing, anti‐ageing, anti‐wrinkle and UV protective effects 11. However, the quality and phytochemical profiles of plant extracts varied in wide range, depending on climate, soil, latitude, seasonal factors, time of harvest, and the field management practice, which could be a challenge to standardize their activities 12. The search for novel natural phytochemicals has led to the gathering bioactive extracts not only by plants, but also from mushrooms, algae, and also, by utilization of by‐products of plant origins 13, 14, 15, 16, 17. Nowadays, with the growth in consumers’ interest to effective and safe natural products, a “new generation” of high quality bioactive phytochemicals, produced by the plant cell culture technology, have been introduced in the past decade, and now, their presence in the cosmeceuticals market have been steadily on the rise.

3. Plant cell culture technology: principles for production of active cosmetic ingredients

Plant cell culture technology is a technique for growing of plant cells under strictly controlled environmental conditions. Because plant cells are considered totipotent, they have the potential to express the full genetic machinery coded in the nucleus, and thus, they are able to produce the full spectrum of characteristic secondary metabolites, found in mother plants. Plant cells are amenable to good manufacturing practice procedures and can be easily propagated by using large volume bioreactors independently on climate or soil or field management practices 5, 18. Moreover, in vitro cultured plant cells are characterized with fast growth, and the ability to accumulate large amount of uniform biomass for a short period of time 19, 20. This is very important advantage especially for the production of rare bioactive compounds, as resveratrol, paclitaxel or terpenoids, which are usually found in low concentrations in plants and their isolation and purification requires the processing of large amounts plant biomass 21, 22, 23, 24, 25. Additionally, plant cell culture technology offers a reliable and powerful production platform for continuous supply of contamination‐free, phytochemically uniform biomass from herbal, aromatic, medicinal, and even from rare and threatened plant species 26. The perspective to obtain natural phytochemicals by using an environmentally sustainable biosynthetic platform made plant cell culture technique exceptionally attractive for the production of active ingredients for high added values “green” cosmetic formulations 27, 28, 29, 30. It should be noted, that active cosmetic ingredients, obtained by the plant cell culture technology are popularized among the customers under the name “plant stem cells”. Here it is important to understand that the term “stem cells” used in this phrase is not always referred to real plant stem cells. The most of the existed plant cell culture production platforms are in fact developed on the basis of the use of dedifferentiated plant cells rather than on the culturing of the true plant stem cells. Dedifferentiated plant cells are obtained by dedifferentiation of already differentiated mature plant cells from different specialized tissues, whereas the true plant stem cells should be never differentiated in their life cycle. Because raised from the dedifferentiation of differentiated cells, the dedifferentiated plant cell cultures could inherit some epigenetic modifications, characteristic for the type of the tissues they have obtained from, and thus, they could be very heterogeneous in their biosynthetic and growth properties. This fact made it possible to generate almost unlimited numbers of plant cell lines with unique phytochemical profiles and growth characteristics even from the same plant, used for their initiation. For this reason, the terms as “leaf stem cells”, “meristems stem cells”, “root stem cells” “rhizome stem cells”, “flower stem cells”, “fruit stem cells”, etc. could be often seen in INCI names of active cosmetic ingredients, but in fact all of these refers to dedifferentiated plant cell cultures. On the other hand, there are plant cell technologies existed, which are developed on the basis of the cultivation of true plant stem cells – the cambial meristematic cells (CMC). These cells are isolated by cambial layer and consist only of true meristem stem cells 31, 32. Cambial meristematic cells are characterized with fast and uniform growth, lack of epigenetic modifications and ability to produce predictable yields of secondary metabolites when treated with stimulating factors such as elicitors 33. This technology have been used by the Korean company “Unhwa Corp.” to produce several active cosmetic ingredients, by using the patent protected expression platform Ddobyul®, developed on the basis of propagation of cambial meristematic cells (Table 1). Another important fact, concerning the plant cells used for cosmetics, is that the term “plant stem cells” is often equally used for active ingredients, produced by either callus cultures, cell suspensions or hairy roots. It is important to understand, that the callus cultures are plant cells, cultured on solid medium, whereas the cell suspension cultures are single plant cells or small cell aggregates cultivated under submerged conditions in liquid medium. Both callus cultures and cell suspensions could consist of dedifferentiated or true stem cells. On the opposite, the hairy roots are organ cultures, obtained by genetic transformation of plant cells 23, 34. Some of the plant cell culture technology – derived active cosmetic ingredients, currently available on the market are reviewed in Table 1.

Table 1.

Some of the most popular plant cell culture technology – derived active cosmetic ingredients, currently available on the market

Market names of active ingredient	Plant species	Type of cell culture and extracts	Benefits	Company; country
Phyto‐Biotics Perilla®	Perilla frutescens	cell suspension extract	anti‐aging, antimicrobial, soothing effect	Active Concepts LLC; USA
Phyto‐Biotics Quercus®	Quercus alba	meristematic stem cells extract	antioxidant, soothing effect, antimicrobial, anti‐aging	Active Concepts LLC; USA
Phyto‐Biotics Acai®	Euterpe oleracea	cell suspension extract	antioxidant, anti‐aging, anti‐wrinkle, soothing effect, moisturizer	Active Concepts LLC; USA
AKOSKY® APIUM	Apium graveolens	callus culture extract	skin regeneration	Akott Evolution S.R.L.; Italy
VITADENIA®	Gardenia taitensis	callus culture extract	anti‐aging, anti‐wrinkle, regenerative, repairing	Biocosmethic; France
UrbanEthic®	Gossypium herbaceum	stem cell extract	anti‐photo‐aging, protection against atmospheric pollutants and heavy metals. protection from oxidative stress	Biocosmethic; France
NatureCells Hydragenesis	Vitis vinifera cv. Verdejo	liposomal complex of cell suspension extract	antioxidant, moisturizer, hydrating effect	Infinitec; Spain
NatureCells Antiaging	Vitis vinifera cv. Mencia	liposomal complex of cell suspension extract	anti‐aging, antioxidant, anti‐wrinkle	Infinitec; Spain
NatureCells Anti Stretch Marks	Centella asiatica	liposomal complex of cell suspension extract	skin‐firming, anti‐stretch mark effect, anti‐inflammatory, anti‐cellulite	Infinitec; Spain
InnovaStemCell Calendula W	Calendula officinalis	cell suspension extract	anti‐wrinkle, skin regeneration, moisturizer	Innova BM; Bulgaria
InnovaStemCell Calendula EM	Calendula officinalis	emulsified cell suspension	anti‐wrinkle, skin regeneration, deep hydration	Innova BM; Bulgaria
InnovaStemCell Rosa damascena W	Rosa damascena	cell suspension extract	skin regeneration, anti‐aging, antioxidant, anti‐inflammatory	Innova BM; Bulgaria
InnovaStemCell Rosa rugosa W	Rosa rugosa	cell suspension extract	antioxidant, antimicrobial, UV protective	Innova BM; Bulgaria
Plant C‐Stem™ Vigna Radiata	Phaseolus radiatus	meristem cell culture vacuole extract	anti‐aging, anti‐wrinkle, protection/repair of environmental damage	Innovacos Corp.; USA
Roseroot (Rhodiola rosea L .) Plant Stem Cell Extract	Rhodiola rosea	callus culture extract	antioxidant	In vitro Plant‐tech AB; Sweden
Milk thistle (Silybum marianum L.) plant stem cell extractat	Silybum marianum	callus culture extract	antioxidant	In vitro Plant‐tech AB; Sweden
Greater plantain (Plantago major L.) plant stem cell extract	Plantago major	callus culture extract	antioxidant	In vitro Plant‐tech AB; Sweden
PhytoCellTec™ Malus Domestica	Malus domestica	liposomal complex of cell suspension extract	anti‐wrinkle	Mibelle AG Biochemistry; Switzerland
PhytoCellTec™ Solar Vitis	Vitis vinifera cv. Gamay Teinturier Fréaux	cell suspension extract	anti‐photo‐aging, extending skin vitality	Mibelle AG Biochemistry; Switzerland
PhytoCellTec™ Alp Rose	Rhododendron ferrugineum	cell suspension extract	protecting longevity, increasing skin vitality, protection/repair of environmental damage	Mibelle AG Biochemistry; Switzerland
PhytoCellTec™ Argan	Argania spinosa	cell suspension extract	improve activity of human dermal stem cells, anti‐wrinkle	Mibelle AG Biochemistry; Switzerland
PhytoCellTec™ Symphytum	Symphytum officinale	cell suspension extract	increase skin renewal, rejuvenates skin epidermis	Mibelle AG Biochemistry; Switzerland
PhytoCellTec™ nunatak®	Saponaria pumila	cell suspension extract	increase skin elasticity, firmness and density, protection/repair of environmental damage, anti‐aging	Mibelle AG Biochemistry; Switzerland
RootBioTec HO	Ocimum basilicum	hairy root culture extract	reduce hair loss, inhibit 5α reductase activity, stimulate dermal papilla cells	Mibelle AG Biochemistry; Switzerland
EGF (Epidermal Growth Factor);	Oryza sativa	recombinant proteins expressed by rice cell culture	restore reduced activity with age, stimulate epidermal skin cells, anti‐aging, anti‐wrinkel, wound healing, stimulation on collagen synthesis in aged skin	Natural Bio‐Materials (NBM) Inc.; Korea
FGFb (basic Fibroblast Growth Factor)	Oryza sativa	recombinant proteins expressed by rice cell culture	anti‐aging, anti‐wrinkle, anti‐hair loss, hair growth	Natural Bio‐Materials (NBM) Inc.; Korea
IGF‐1 (Insulin‐like Growth Factor‐1)	Oryza sativa	recombinant proteins expressed by rice cell culture	growth support for skin cells with EGF, support young and healthy skin	Natural Bio‐Materials (NBM) Inc.; Korea
KGF (Keratinocyte Growth Factor, Fibroblast Growth Factor‐7)	Oryza sativa	recombinant proteins expressed by rice cell culture	specific growth factor for keratinocyte	Natural Bio‐Materials (NBM) Inc.; Korea
VEGF165 (Vascular Endothelial Growth Factor)	Oryza sativa	recombinant proteins expressed by rice cell culture	skin cell growth and support, wrinkel and skin aging improvement, wound healing after laser resurfacing or injury, hair cell growth and support, hair cycle regulation and anti‐hair loss	Natural Bio‐Materials (NBM) Inc.; Korea
Foreseen Shield Nopal	Opuntia ficus indica	powdered leaf cells	antioxidant, anti‐wrinkle, anti‐aging, UV protective, decrease melanine spots	Naolys, France
Initial E [PT+TMG]	Polianthes tuberosa	callus culture extract	anti‐aging, anti‐wrinkle, anti‐circle,
anti‐puffines, brightening, firming, soothing, moisturizing	Naolys, France
Power Extension [HSB+R]	Hibiscus syriacus	leaf cell extract with rutin	anti‐aging, anti‐wrinkle, antioxidant	Naolys, France
All Even Sweet iris	Iris pallida	glycerin based leaf cell extract	anti‐aging, anti‐wrinkle, firming	Naolys, France
All Fiber Booster Olive tree	Olea europea	glycerin based leaf cell extract	anti‐aging, restructurating	Naolys, France
All Fiber Booster Chinese hibiscus	Hibiscus rosa sinensis	glycerin based leaf cell extract	anti‐aging, restructurating	Naolys, France
All Fiber Booster Green tea	Camellia sinensis	glycerin based leaf cell extract	anti‐aging, firming, regenerating, softening, restructurating	Naolys, France
Fiber Booster Plus Sequoia and Vitis flower	Sequoia sempervirens and Vitis vinifera	glycerin based leaf and flower cells extract	anti‐aging, antioxidant	Naolys, France
Inside Heart Egyptian blue lily	Nymphaea caerulea l	glycerin based leaf and flower cells extract	anti‐aging, antioxidant	Naolys, France
Revive Commiphora and
Rose from Damas	Commiphora myrrha and Rosa damascena	glycerin based leaf cells extract	anti‐aging, energizing	Naolys, France
StandStill Rose from Damas	Rosa damascena	glycerin based leaf cells extract	anti‐aging, firming	Naolys, France
New ReGeneration Cocoa	Theobroma cacao	glycerin based leaf cells extract	anti‐aging, firming, regenerating, protective, restructurating	Naolys, France
Total Generation Sequoia and Egyptian blue lily	Sequoia sempervirens and Nymphaea caerulea	glycerin based leaf cells extract	anti‐aging, regenerating	Naolys, France
Total Generation Curry plant	Helicrysum italicum	glycerin based leaf cells extract	anti‐aging, regenerating	Naolys, France
Inside Light Poet's narcissus	Narcissus poeticus	callus culture extract	brightening, lightening and anti‐blemish, anti‐aging	Naolys, France
Bright Light Madonna lily	Lilium candidum	powdered leaf cells	brightening, regenerating	Naolys, France
LightWaves Defense [JS+M]	Jasminum sambac	sunflower oil based leaf cells extract	protective, anti‐aging, radiance, firming, anti‐redness	Naolys, France
Global Protect Common juniper	Juniper communis	glycerin based leaf cells extract	protective, anti‐pollution	Naolys, France
OxyRelax California poppy	Eschscholzia californica	glycerin based leaf cells extract	protective, antioxidant, anti‐aging	Naolys, France
OxyRelax Cherry tree	Prunus cerasus	glycerin based leaf cells extract	protective, antioxidant, cell relaxing	Naolys, France
Smooth Lightening White rose	Rosa alba	glycerin based leaf cells extract	protective, antioxidant, radiant, regenerating, detoxifying	Naolys, France
Sun Protect Date palm	Bombax costatum	glycerin based leaf cells extract	protective, soothing, sunscreen, anti‐aging	Naolys, France
Whole Protection Edelweiss	Leontopodium alpinum	sunflower oil based leaf cells extract	protective, soothing, repairing, anti‐aging	Naolys, France
Whole Protection Red‐flowered silk cotton tree	Bombax costatum	glycerin based leaf cells extract	protective, soothing	Naolys, France
Refine Ginger	Zingiber officinale	glycerin based leaf cells extract	astringent, firming, mattifying, moisturizing, antioxidant	Naolys, France
Pure Light Chinese peony	Paeonia lactiflora	powdered leaf cells	mattifying, radiance, moisturizing, soothing, relaxing	Naolys, France
Unwind Sacred lotus	Nelumbo nucifera	sunflower oil based leaf cells extract	soothing, relaxing, radiance	Naolys, France
Essential Being Indian jasmine	Jasminum sambac	glycerin based leaf cells extract	regenerating, detoxifying	Naolys, France
Full Detox Ylang Ylang	Cananga odorata	sunflower oil based leaf cells extract	radiance, mattifying, regenerating, antioxidant, anti‐pollution	Naolys, France
Purify Apothecary's rose	Rosa gallica	sunflower oil based callus extract	soothing, regenerating, radiance	Naolys, France
Purify Aloe vera	Aloe barbadensis	sunflower oil based callus extract	soothing, regenerating, radiance	Naolys, France
First Light Snow lotus	Saussurea involucrata	sunflower oil based callus extract	radiance, brightening, regenerating, antioxidant	Naolys, France
HydraSourcing [AM+PS]	Argemone mexicana	glycerin based callus extract	moisturizing, protective, regenerative, anti‐aging	Naolys, France
HydraGeneration Papyrus	Cyperus papyrus	powdered leaf cells	hydrating, protective, barrier function	Naolys, France
HydraGeneration Pale rose	Rosa centifolia	glycerin based leaf cells extract	moisturizing, regenerative	Naolys, France
HydraSoothing Indian olibanum	Boswellia serrata	sunflower oil based leaf cells extract	moisturizing, regenerative, antioxidant, soothing	Naolys, France
Soothing Light Apple tree	Malus domestica	powdered leaf cells	soothing, lightening, radiance	Naolys, France
Fragile Vitis flower	Vitis vinifera	glycerin based flower cells extract	soothing, antioxidant	Naolys, France
Fragile Cotton	Gossypium arboreum	glycerin based leaf cells extract	soothing, protective	Naolys, France
Fragile Japanese Cherry tree	Prunus serrulata	sunflower oil based leaf cells extract	soothing, antioxidant	Naolys, France
Soothing Light Apricot	Prunus armeniaca	glycerin based leaf cells extract	soothing, lightening	Naolys, France
OvernightEnhance [MJ+C]	Mirabilis jalapa	glycerin based callus extract	radiance, energizing, repairing, detoxifying, antioxidant	Naolys, France
Balancing Energy Asian ginseng	Panax ginseng	sunflower oil based leaf cells extract	energizing, antioxidant	Naolys, France
Light&Energy Coffee and Saffron	Crocus sativus and Coffea arabica	sunflower oil based callus and leaf cells extract	energizing, antioxidant, radiance	Naolys, France
Full Energy Vanilla	Vanilla planifolia	sunflower oil based leaf cells extract	energizing, antioxidant, regenerating	Naolys, France
Splint&Slim Great bougainvillea	Bougainvillea spectabilis	powdered leaf cells	firming, slimming, lipolitical, anti‐aging	Naolys, France
EtHAIReal Peppermint	Mentha piperita	sunflower oil based leaf cells extract	regulating, soothing, antioxidant, lightening	Naolys, France
Healthy Shine Lilac	Syringa vulgaris	glycerin based leaf cells extract	restoring shine and damaged hair, soothing, energizing, protective	Naolys, France
3HC Hair Stimulation Complex	Vitis vinifera	mix of meristem plant cell culture derived active ingredients and hydrolyzed plant extracts	increase the lifespan of hair and reduce hair loss	Phenbiox SRL; Italy
Soy Cell	Glycine max	callus culture extract	anti‐photo‐aging, antiradical, anti‐wrinkles	Phenbiox SRL; Italy
G‐Cell	Vitis vinifera	meristem cell culture from green unripe grapes extract	antioxidant, skin protective	Phenbiox SRL; Italy
P‐Cell	Capsicum annuum	fruit meristem cell culture extract	promote cell protein synthesis, increase skin elasticity	Phenbiox SRL; Italy
Citrustem™	Citrus aurantium dulcis	callus culture extract	anti‐aging, skin conditioner	Provital Group; Spain
Lingostem™	Vaccinium vitis‐idaea	callus culture extract	anti‐aging, anti‐wrinkle, make up treatments	Provital Group; Spain
RASTEM	Lpomoea purpurea	callus culture extract	skin‐renewal, anti‐inflammatory, anti‐wrinkle, antioxidant	Radiant Inc.; Korea
Carrot Stem Cell	Daucus carota	callus culture extract	antioxidant, skin‐nourishing effect	Radiant Inc.; Korea
Cucumber Stem Cell	Cucumis sativus	callus culture extract	skin cooling effect, moisturizer	Radiant Inc.; Korea
Ginseng Stem Cell	Panax ginseng	cell culture extract	skin regeneration	Radiant Inc.; Korea
Lotus Stem Cell Extract	Nelumbo nucifera	callus culture extract	controls excessive sebum secretion, absorbs sebum, maintains clean skin condition	Sandream Impact LLC; USA
Tomato Callus Stem Cell Extract	Solanum lycopersicum	callus culture extract	moisturizer	Sandream Impact LLC; USA
Rice Callus Stem Cell Extract	Oryza sativa	callus culture extract	antioxidant, anti‐inflammatory, whitening	Sandream Impact LLC; USA
Carrot Callus Stem Cell Extract	Daucus carota	callus culture extract	moisturizer	Sandream Impact LLC; USA
Rose Callus Stem Cell Extract	Rosa damascena	callus culture extract	protecting longevity, delaying senescence, increasing skin vitality, boosting epidermal skin cell regeneration	Sandream Impact LLC; USA
Grape Callus Stem Cell Extract	Vitis vinifera	callus culture extract	antioxidant, anti‐inflammatory	Sandream Impact LLC; USA
Orchid Callus Stem Cell Extract	Orchis spp.	callus culture extract	promote skin growth and proliferation, moisturizer, skin rejuvenation, soothing effect	Sandream Impact LLC; USA
Ginseng Callus Stem Cell Extract	Panax pseudoginseng	callus culture extract	restorative effect, tonic	Sandream Impact LLC; USA
Green Tea Callus Stem Cell Extract	Camellia sinensis	callus culture extract	antioxidant, anti‐inflammatory	Sandream Impact LLC; USA
Marrubium Stems GX™	Marrubium vulgare	cell suspension extract	protective effect, pollution defense, skin conditioner	Sederma (Croda Personal Care); United Kingdom
Buddleja Stems GX™	Buddleja davidii	cell suspension extract	anti‐photo‐aging, anti‐aging, protects from UV‐induced oxidative stress, anti‐inflammatory, matrix metalloproteinase activation	Sederma (Croda Personal Care); United Kingdom
Centella Stems GX™	Centella asiatica	cell suspension extract	reduces skin redness, straightening capillary structure, fights against rosacea	Sederma (Croda Personal Care); United Kingdom
Dermasyr 10™	Syringa vulgaris	verbascoside concentrate extracted from leaf cell culture	reduce skin blemishes and inflammatory hyper‐pigmentation, control skin redness and balances of seborrhea, soothing, purifying, skin conditioning	Sederma (Croda Personal Care); United Kingdom
Echinacea Stems GX™	Echinacea angustifolia	cell suspension extract	stimulates collagen synthesis, prevents collagen loss, reduces capillary permeability	Sederma (Croda Personal Care); United Kingdom
Gardenia Stems GX™	Gardenia jasminoides	cell suspension extract	reduces MMP‐1 synthesis, inhibits collagenase activity, stimulates collagen synthesis	Sederma (Croda Personal Care); United Kingdom
Leontopod Stems GX™	Leontopodium alpinum	cell suspension extract	prevents collagen loss, antioxidant, anti‐wrinkle	Sederma (Croda Personal Care); United Kingdom
Celtosome™ Eryngium Maritimum ST	Eryngium maritimum	dried stem cell	anti‐aging, improving skin firmness, skin renewal	SEPPIC (Air Liquide Healthcare); France
Celtosome Crithmum Maritimum ST	Crithmum maritimum	dried stem cell	skin lightening, wound healing, lightening effect, pigmentation regulation, anti‐wrinkle	SEPPIC (Air Liquide Healthcare); France
Ddobyul®	Panax ginseng	cambial meristematic cells suspension extract	anti‐aging	Unhwa Corp.; Korea
Ddobyul®	Taxus cuspidata	cambial meristematic cells suspension extract	anti‐allergic	Unhwa Corp.; Korea
Ddobyul®	Ginkgo biloba	cambial meristematic cells suspension extract	whitening	Unhwa Corp.; Korea
Ddobyul®	Solanum lycopersicum	cambial meristematic cells suspension extract	antioxidant	Unhwa Corp.; Korea
Vita SeneBlock	Citrus limon	mix of cell wall peptides and sugars from somatic embryos	inducing youth and longevity markers, protect DNA	Vitalab s.r.l. (Arterra Bioscience s.r.l); Italy
Vita iLux	Capsicum annuum	mix of peptides and sugars from cell cultures	firming effect, promoting bilirubin degradation, improves skin's regenerative process	Vitalab s.r.l. (Arterra Bioscience s.r.l); Italy
Hibiskin Vita	Hibiscus syriacus	cell culture extract	restore colagen, improve skin barrier function, moisturizer, improves skin's regenerative process	Vitalab s.r.l. (Arterra Bioscience s.r.l); Italy
Mythos Vita	Actinidia arguta	mix of stem cell culture derived extract and hydrosoluble plant fruit extract	anti‐photo‐aging, protect DNA	Vitalab s.r.l. (Arterra Bioscience s.r.l); Italy
Vita Nova	Lotus japonicus	mix of cell wall peptides and sugars from somatic embryos	extends cellular vitality and longevity, anti‐aging, reactivate skin's natural repair process	Vitalab s.r.l. (Arterra Bioscience s.r.l); Italy
Vita Genesis White	Brassica rapa	mix of cell wall fraction and hydro‐ethanolic extract derived from hairy root cultures	inhibits pigmentation process, reduce melanin synthesis	Vitalab s.r.l. (Arterra Bioscience s.r.l); Italy
Vita Freeze	Solanum lycopersicum	mix of peptides and sugars from cell cultures	prevent premature skin ageing, improve skin elasticity, extend cellular vitality and longevity	Vitalab s.r.l. (Arterra Bioscience s.r.l); Italy
Daphne VitaSense	Daphne odoracell	cell suspension extract	strengthens skin protecting barrier, accelerate healing of cutaneous micro‐lesions, reduce skin inflammation	Vitalab s.r.l. (Arterra Bioscience s.r.l); Italy
VitaLight	Cirsium eriophorum	cell suspension extract	reduce sebum production, reduce skin inflammation, increase skin desquamation	Vitalab s.r.l. (Arterra Bioscience s.r.l); Italy
VitaShape	Coleus forskohlii	cell suspension extract	anti‐fat properties, anti‐oedema effect, antioxidant, detoxifying capacity	Vitalab s.r.l. (Arterra Bioscience s.r.l); Italy
BerryFlux Vita	Rubus idaeus	cell culture extract	moisturizer, hydrating effect, improve production of ceramides	Vitalab s.r.l. (Arterra Bioscience s.r.l); Italy
FicuCell Vita	Opuntia spp.	cell culture extract	protect extracellular matrix, extend cellular longevity, delay premature skin ageing, improve skin firmness and elasticity	Vitalab s.r.l. (Arterra Bioscience s.r.l); Italy
DoliCos PhotoProtect	Dolichos spp.	cell culture extract	UV protective,detoxifie cells, soothing effect	Vitalab s.r.l. (Arterra Bioscience s.r.l); Italy
Cell Pulse	Coffea spp.	cell culture extract	antioxidant, cell energiser, anti‐wrinkle	Vitalab s.r.l. (Arterra Bioscience s.r.l); Italy
Lycoskin Defence	Solanum lycopersicum	water‐soluble fraction from cell cultures	extend cellular vitality, protect DNA, radiance effect	Vitalab s.r.l. (Arterra Bioscience s.r.l); Italy
BioNymph Peptide	Nicotiana sylvestris	mix of peptides and sugars from cell cultures	antioxidant, extend cellular vitality and longevity, anti‐wrinkle, improve skin smoothness	Vitalab s.r.l. (Arterra Bioscience s.r.l); Italy
Cellintegrity	Rubus idaeus	concentrated water‐soluble fraction from cell cultures	antioxidant, anti‐inflammatory, protect DNA, enhance cellular longevity, soothing effect, strengthening action	Vitalab s.r.l. (Arterra Bioscience s.r.l); Italy
SENSIA CAROTA PRCF	Daucus carota sativa	lysate of root stem cells culture	anti‐aging, anti‐inflammatory, antioxidant	Vytrus Biotech; Spain
LUMINIA GRANATUM PRCF	Punica granatum	powdered plant stem cells	antioxidant, reduces hyper‐pigmentation, enhances skin radiance and glow	Vytrus Biotech; Spain
ARABIAN COTTON PRCF	Gossypium herbaceum	lysate of plant stem cells culture	soothing, anti‐inflammatory, antioxidant, photoprotection, regeneration	Vytrus Biotech; Spain
TURMERIA ZEN PRCF	Curcuma longa	glycerin based lysate of rhizome cells culture	anti‐stress wrinkles, emotional hydration manager, modulator of Brain‐skin connection	Vytrus Biotech; Spain
SARCOSLIM RE‐SHAPE PRCF	Sarcocapnos crassifolia,	lysate of callus culture	skin regeneration and repair	Vytrus Biotech; Spain
CAPILIA LONGA PPF	Curcuma longa	plant growth factor peptides ‐ concentrated secretome of totipotent rhizome cells	hair follicle regeneration, hair growth re‐activation	Vytrus Biotech; Spain
CENTELLA REVERSA PPF	Centella asiatica	plant signaling peptides ‐ concentrated secretome of totipotent petioles cells	rebuilding the core skin structure	Vytrus Biotech; Spain

3.1. Plant stem cells

Plant cells can be propagated and used for continuous supply of fresh plant biomass for cosmetic formulations. However, it is of great importance to understand that we cannot introduce the entire plant cells in cosmetic products and keep them alive as active ingredient there 30, 35. Plant cells are extremely sensitive to environmental factors, nutrient medium composition, osmotic and mechanical (shear) stress, gas exchange and oxygen supply, temperature, light, ionic strength and water content, and thus, they can survive neither during cosmetic products preparation, nor during storage or application of cosmetics on skin. Even a delivery system, able to maintain and supply live stem cells could be developed, the size and the specific structure of plant cells will not allow them to attach or penetrate the skin surface. Because of all discussed concerns, the plant stem cells are rather used as a raw material for preparing different types of extracts, which then could be included in cosmetic formulations as active ingredients. However, there are various products, available on the market, which are based on a whole dried plant cells and standardized on the cells count per gram of active ingredient (Table 1). Such examples are the anti‐aging active ingredient Celtosome™ Eryngium Maritimum ST (based on sea holly cell culture with 100 million cells/g of active ingredient), and the tyrosinase inhibitory skin lightening active ingredient Celtosome™ Crithmum Maritimum ST (based on rock samphire cell culture with more than 1 billion cells/g of active ingredient) (Table 1).

3.2. Plant stem cells extracts

Most of the available active cosmetic ingredients, obtained by plant cell culture technology are marketed in the form of different extracts (Table 1). In contrast to the plant – derived extracts, the extracts obtained from plant cell cultures can be easily standardized, and perfectly complying with the strict safety requirements that the high end cosmetic market constantly demands 35. Plant cell extracts are free of pathogens, agrochemical, toxic substances, allergens and pollutants, because they are produced under controlled conditions, complying with the procedures of good manufacturing practice. Depending of the type of used solvent, the plant cell extracts used in cosmetics could be contingently divided to liposoluble (extracted with oils) and hydrosoluble (extracted with glycerin) extracts, dried extracts (conditioned with maltodextrin), plant cell wall extracts (rich in peptides and sugars), nanoemulsions or suspension extracts 35. However, most of the existed extracts have been developed on the basis on extraction of target compound or group of closely related bioactive compounds, and thus, not the entire health beneficial potential of the extracted plant cells could be utilized. Some exceptions could be found, where the entire plant cells were freeze dried and powdered for direct application in cosmetic formulations (see some products offered by “Vytrus Biotech” and “Naolys”, Table 1), or the entire cell suspensions have been, emulsified or encapsulated in liposomal complex (see some products offered by “Innova BM” and “Mibelle AG Biochemistry”, Table 1) by using high‐pressure homogenizers.

3.3. Molecular farming for production of recombinant proteins

Plants are excellent production matrixes for expression of recombinant proteins with important pharmaceutical properties 36. This powerful recombinant protein expression technique, known as “molecular farming” have been widely used for the production of vaccines 37, 38, 39, cytokines 40, and even for production of therapeutic protein for human use 5, 41, 42. The classical molecular farming technology is based on genetic modification of plants for recombinant protein expression. This could be realized by permanent integration of foreign genes into the host DNA to generate stable transgenic lines, or by transient transformation of intact plant leaves 43. However, both methods requires growing of whole plants under strictly controlled environmental conditions into contained greenhouses, complying with the strict regulatory standards 36. By using such technology, The Iceland Company “BIOEFFECT” has applied for the first time a plant‐based transgenic platform for large‐scale production of Epidermal Growth Factors (EGF) for cosmetic use, expressed in genetically engineered barley seeds. The company offers a wide range of skin‐care products containing this cellular activator, which contribute for healthier and younger‐looking skin. However, the recent interests in molecular farming have been focused on adaptation of plant cell culture technology for the production of recombinant proteins. This technique offers sustainable and continuous heterologous proteins production by plant cells growing under precisely controlled micro‐environmental in vitro conditions in bioreactors. Cultivation of genetically engineered plant cell cultures have been recognized as much powerful expression platform, when compared to the classical molecular farming techniques, relying on agricultural‐scale production by growing of genetically transformed plants 18, 41. Recently, the Korean company “Natural Bio‐Materials (NBM)”, has launched series of active cosmetic ingredients containing growth factors (Epidermal Growth Factor, Basic Fibroblast Growth Factor, Insulin‐like Growth Factor‐1, Keratinocyte Growth Factor, Fibroblast Growth Factor‐7, and Vascular Endothelial Growth Factor), expressed by recombinant rice (Oryza sativa L.) cell cultures (Table 1). However, even that the human recombinant proteins, produced by this technology have undeniable advantages and are characterized with high level of purity, animal‐free, virus‐free, bacterial‐free and exotoxin‐free production, there are still some consumers which have concerns to use such products because they are expressed by genetically modified organisms.

4. Case Study: Initiation, growth, phytochemical profile and physicochemical characteristics of exopolysaccharides in INNOVA StemCell Calendula

Calendula (Calendula officinalis L.), known as “marigold”, has been widely used in traditional herbal medicine and skin care cosmeceuticals for topical application 11. The pant was shown to be rich in phenolic acids, flavonoid, triterpenes, carotenoids, aroma compounds and unique mix of polyunsaturated fatty acids 44, 45, 46. Because of its high therapeutic value and the proved cosmetic effects, the Bulgarian company “Innova BM” has developed and released on the market two high quality active cosmetic ingredients, based on Calendula dedifferentiated cell culture (Table 1). The development of these products is schematically presented on Fig. 1. The technological steps include screening of calendula plants with superior phytochemical profiles, selection, sterilization and cultivation of plant explants on callus induction medium, selection of friable cell lines with appropriate phytochemical profiles, initiation of liquid cell suspension culture and optimization of cultivation conditions and nutrient medium composition. The optimization step is critical in our technology, since a significant increase in biosynthetic potential and accumulated biomass of selected cell line can be achieved (Table 2, Fig. 2). After optimization, the selected line was scaled‐up to large scale cultivation in stirred tank bioreactor. The produced cell suspension (cells and culture liquid) was then processed by high‐pressure homogenizer to produce glycerin extract (50 % wt.) or calendula emulsion (75 % wt. cell suspension) 47. The produced active ingredients have been found to have superior moisturizing, anti‐wrinkle, hydrating and regenerative effects, when applied on skin. These effects are due to the high content of secreted exopolysaccharides, during cultivation of calendula cells (Fig. 2D). The exopolysaccharides have been identified to belong to pectin type. The crude exopolysaccharides fraction contains 879 μg/mg neutral sugars and 50 μg/mg proteins. The unique combination of polysaccharides and peptides made this exopolysaccharide fraction almost perfect for application as emulsifier in cosmetic products for topical application. The potential synergistic interactions, when applied with other popular emulsifiers, as well as their emulsion stabilization properties are presented in Table 3. The polysaccharides have molecular weight of 6.7 × 10⁴ Da and contain 413 μg/mg uronic acids. The full monosaccharide composition of exopolysaccharide fraction was determined as: glucuronic acid (13.6 μg/mg), galacturonic acid (399.7 μg/mg), glucose (185.5 μg/mg), galactose (179.9 μg/mg), rhamnose (178.9 μg/mg), arabinose (166.7 μg/mg), fucose (0.6 μg/mg) and mannose (4.7 μg/mg). The presence of this exopolysaccharides fraction incorporated into Innova StemCell Calendula products, in combination with bioactive compounds from the released cell content (Table 2) made these products unique and one of the kind in the market of active cosmetic ingredients.

Figure 1.

Schematic presentation of technological steps in development of InnovaStemCell Calendula active cosmetic ingredient.

Table 2.

GC‐MS profiles of Calendula officinalis stem cell suspension before and after optimization of nutrient medium composition 47

Metabolites	Calendula cell suspension before optimization	Calendula cell suspension after optimization
	μg/g dry biomass	μg/g dry biomass
Amino acids
Glycine	62.77	106.74
Serine	21.09	175.87
L‐Proline	73.08	162.56
L‐Aspartic acid	10.83	138.41
L‐Glutamic acid	161.68	274.91
L‐Asparagine	19.19	146.38
L‐Glutamine	88.03	224.10
Organic acids
Succinic acid	6.40	130.88
Fumaric acid	7.23	172.30
Malic acid	5.41	149.20
GABA	50.02	184.38
Pyroglutamic acid	20.41	115.51
Saturated and Non‐saturated Fatty Acids
n‐Tetracanoic acid (Myristic acid) (C14:0)	26.68	83.07
n‐Hexadecanoic acid (Palmitic acid) (C16:0)	183.91	178.69
9‐(Z)‐Hexadecenoic acid (Oleic acid) (C 18:1)	39.33	95.36
Octadecanoic acid (Stearic acid) (C 18:0)	197.61	192.01
Sterols
Campesterol	59.27	59.07
Stigmasterol	65.34	89.14
β‐Sitosterol	94.82	112.77
β‐Amyrin	12.13	77.46
Phenolic acids
Quinic acid	13.49	81.92
Caffeic acid	‐	62.05
trans‐Cinnamic acid	‐	170.39
p‐Coumaric acid	‐	67.63

Figure 2.

Dynamics of changes in accumulated dry biomass and growth index (A), fresh weight to dry weight ratio (B), pH and specific electric conductivity of culture medium (C), and exopolysaccharides release (D) during 15 days of submerged cultivation in shaking flasks of Calendula officinalis L. cell suspension culture (115 rpm, 26°C, on darkness).

Table 3.

Synergistic effect of exopolysaccharides, released by Calendula officinalis stem cell suspension in relation to emulsion stabilizing properties in mixtures with cellulose gum, xanthan gum, guar gum, and sodium‐alginate

Model emulsion systems Oil/Water (1:1 v/v) with:	Light microscopy	Turbidimetric method
		Emulsifying activity index (EAI), m2/g	Emulsion stability index (ESI), min
0.6 % Calendula Cell Suspension Exopolysaccharides		175.0	16.8
0.6 % Calendula Cell Suspension Exopolysaccharides And 0.3 % Cellulose Gum		38.4	13.9
0.6 % Calendula Cell Suspension Exopolysaccharides and 0.3 % Xanthan Gum		47.6	22.1
0.6 % Calendula Cell Suspension Exopolysaccharides and 0.3 % Guar Gum		322.4	15.0
0.6 % Calendula Cell Suspension Exopolysaccharides and 0.3 % Sodium Alginate		1307.5	60.7

The emulsifying activity index (EAI) and emulsion stability index (ESI) were determined according to 48. Model emulsion systems were prepared by mixing polysaccharides, water and sunflower oil for 5 min at 10 000 rpm by homogenizer (Ultra Turrax IKA T18 Basic, Germany).

5. Concluding remarks

Following the ever‐growing demand for high quality natural products of plant origin for application as active ingredients in cosmeceutical formulations, the plant cell culture technology has developed powerful production platforms which can effectively supply the customers’ needs. We are witnesses of exponentially growing number of commercialized plant cell – derived ingredients, offered on the cosmetic industry market and the diversity of utilized plant species, used for their production has continue to increase every year. In fact, the observed interest to production of plant cell ‐ derived ingredients for cosmetic needs could be correlated with the recent advance in development and commercialization of plant cell culture technology in technologically advanced countries. Moreover, the new developed techniques of gene editing, metabolite engineering and synthetic biology could have a significant impact on improvement the yields and development of tailor made cosmetic products with desired activities. The advance in molecular farming has already leaded to commercial production of rare human activator peptides, cytokines and growth factors, which are the first step in development of cosmetic products with potential to extend skin life by using the body self‐repair mechanisms. However, till now there are many open questions, concerning regulatory standards and documentation, unification of health beneficial claims and the methodology for evaluation of pharmaceutical effects, which should be answered in order to help the consumers to make the right choice of their cosmeceutical product.

Practical application

The aim of this paper is to review the recent progress in plant cell technology for cosmetic application and to provide short overview on commercialized plant cell culture – derived active cosmetic ingredients available on the market.

The authors have declared no conflict of interest.